TRAINING MANUAL ON

SEED PRODUCTION AND FARMING OF

BRACKISHWATER FINFISHES

30th July – 3rd August, 2018

Funded by
NATIONAL FISHERIES DEVELOPMENT BOARD
HYDERABAD

ICAR-CENTRAL INSTITUTE OF BRACKISHWATER AQUACULTURE

75, Santhome High road, R. A. Puram, Chennai, 600028, India
Tel: 91-44-24618817, 24616948, Fax-91-44-24610311
Published by

Dr. K. K. Vijayan
Director

Compiled and Edited by

Dr. M. Kailasam
Dr. M. Makesh
Dr. S.N. Sethi
Dr. Krishna Sukumaran
Dr. Aritra Bera
Smt. Babita Mandal
Shri. T. Sivaramakrishnan
Shri.Tanveer Hussain
Shri. Dani Thomas
Shri. Pankaj Patil
 CONTENTS
SI. Page
Topic
No. No.

Recent advances of brackishwater finfish aquaculture in India- Milestones

1. 1
and the way ahead
2. An overview of induced breeding techniques of brackishwater finfishes 10

3. Biology of milkfish (Chanos chanos) 15

4. Seed production of Milkfish (Chanos chanos) – Global scenario 21

Induced breeding, larval rearing and seed production of Asian seabass

5. 23
(Lates calcarifer)

6. Nursery rearing and grow out culture of Asian seabass Lates calcarifer 31

Broodstock development, Induce breeding and larval rearing of milkfish

7. 39
(Chanos chanos)
8. Present status of milkfish Chanos chanos farming 46

9. Biology of grey mullet (Mugil cephalus) 51

Procurement, transportation and acclimatization of fishes for broodstock

10. 58
development
Broodstock Development and Induced Breeding in grey mullet (Mugil
11. 61
cephalus)

12. Nursery rearing and grow-out culture of grey mullet (Mugil cephalus) 67

13. Breeding and seed production of pearlspot, Etroplus suratensis (Bloch) 71

14. Brackishwater ornamental fish breeding techniques 75

Biology of Spotted Scat (Scatophagus argus) – A potential Brackishwater

15. 87
ornamental fish

16. Culture of live food organisms: microalgae, rotifers, artemia and copepods 90

17. Health management in brackishwater finfishes 99

18. Development of formulated feed for cultivable brackishwater finfishes 106

19. Genetic factors in broodstock management in finfish hatchery 111

20. Hormonal control of reproduction in fish 117

21. Synthetic hormones and their role in brackishwater finfish breeding 124

Soil and water quality management of finfish hatcheries and culture

22. 129
practices

23. Transportation methodology of brackishwater finfishes 140

 Recent advances of brackishwater finfish aquaculture in India-
Milestones and the way ahead
K.K. Vijayan, M. Kailasam

Introduction
The brackishwater aquaculture sector is a significant contributor to the economic
development of our country by earning valuable foreign exchange, providing direct and
indirect employment, livelihood opportunities and ensuring nutritional security. Indian
aquaculture sector has witnessed a 6.5 fold increase in the past two decades and contributed
4.8 million tonnes to the total fish production of 9.8 million tonnes in the year 2014 (FAO,
2016). India is endowed with vast areas of brackishwater aquaculture resources amounting to
1.2 million ha, thisis beside the 1.24 million ha salt affected soils in coastal areas of about 6.1
million ha inland saline soils, only a fraction of which is currently utilised (over 1.15 lakh
ha). These figures highlight the potential and scope for the development of brackishwater
aquaculture in our country.

Brackishwater aquaculture has been an age old practice in the traditional culture systems of
India in the form of traditional prawn filtration systems Pokkali fields of Kerala, the bheris of
West-Bengal, Gheris of Orissa, Khar lands of Karnataka and Ghazni fields of Goa. As early
as 1911 James Hornell suggested the development of salt water fish farming in Madras
Presidency which led to establishment of marine fish farm near Tuticorin by utilising few
lagoons in the area and stocking mullets and sand whiting (Mugil spp. and Sillago sp.).
Establishment of Narrakal fish farm at Kochi during 1940-42 symbolised the initiation of
brackishwater fish farming (mullets and milk fish) on a larger scale. Similarly Ayiramthengu
fish farm adjoining Kayamkulam lake in Kerala was a significant initial step in the
development of the culture of brackishwater fish (pearlspot, mullets and milkfish) (Tampi
1958). Another major milestone in the development of brackishwater aquaculture in our
country was the initiation of All India Coordinated Project (AICRP) on brackishwater fish
farming by ICAR in 1973 resulting in the development of many finfish and shrimp farming
technologies.

Despite all these efforts brackishwater aquaculture has been largely synonymous with shrimp
aquaculture owing to the high export potential of shrimps. (Rao and Ravichandran, 2001).
The early 1990’s was characterised by a boom in unregulated shrimp farming activities

1 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

dominated by the giant tiger shrimp Penaeus monodon. This however faced a set-back due to
the emergence white spot disease (WSD) caused by white spot syndrome virus (WSSV).
Since 1993 WSD has continued to pose a major challenge to the shrimp farming. The
consequent economic losses continues to affect the farmers and the shrimp farming sector
even today. This has led to the introduction of exotic SPF Penaeus vannamei in 2009-10, and
in a matter of 5 years, we witnessed a changed scenario of the shrimp farming sector being
dominated by white legged shrimp (P. vannamei), which has totally replaced the Indian tiger
shrimp, a species cultured in 1990’s to 2010. This major species shift took place after a
cautious risk analysis undertaken by the government with ICAR-CIBA playing a key role in
the introduction of this fast growing species with high export potential. Today again as we
take stringent steps to address the potential threat posed by EHP to the shrimp farming sector,
we are also vociferous in promoting diversification of aquaculture species and practices.
ICAR-CIBA has has believed that species diversification in aquaculture and developing need
based location specific technologies is the best roadmap for sustainable aquaculture sector for
our country.

The development of brackishwater finfish seed production technology in India

The prime brackishwater finfish species having high consumer preference and market
demand in India include Asian seabass Lates calcarifer (Bloch, 1790), grey mullet Mugil
cephalus (Linnaeus, 1758), milkfish Chanos chanos (Forsskal, 1775), pearlspot Etroplus
suratensis (Bloch 1790), and red snapper Lutjanus argentimuaculatus. In the year 1997, a
significant milestone achieved with respect to brackishwater finfish aquaculture in our
country with the successful breeding of Asian seabass in captivity, at the Central Institute of
Brackishwater Aquaculture. The R&D in seabass breeding thus led to the establishment of
the first brackishwater/ marine finfish hatchery of our country located at MES, CIBA,
Chennai.

After the successful induced breeding of freshwater carps in 1950’s, it was after a
spell of four decades that a breakthrough was achieved in successful induced breeding and
larval production of seabass, a brackishwater/marine foodfish in our country. Subsequently
seabass hatching and rearing technology, developed by CIBA was transferred to Rajiv
Gandhi Centre for Aquaculture (RGCA) during 1999-2000 (CIBA Annual Report 2000-01).
The hatchery produced seeds are being produced and supplied to farmers, which is further

Training Manual on Seed Production and Farming of Brackishwater Finfishes I 2

spreading the seabass farming in country. Today private entrepreneurs are taking interest in
the farming of seabass, and the demand for hatchery produced seeds of finfish species is on
the rise, reflecting the increasing interest of brackishwater farmers in adopting seabass
nursery rearing and farming technology. Significant progressive efforts towards the breeding
of other candidate species such as cobia Rachycentron canadum (Linnaeus, 1766) and silver
pompano Trachinotus blochii have been made in our country by institutions such as Central
Marine Fisheries research Institute (CMFRI), Central Institute of Brackishwater Aquaculture
(CIBA) and Rajiv Gandhi Centre for Aquaculture (RGCA). For these species the hatchery
based seed production and aquaculture are in their formative stages.

The declaration of pearlspot, “Karimeen” as the state fish of Kerala in 2010 came as a
boon to this indigenous cichlid which has been an important brackishwater food fish of
Kerala, with great local demand with a market price in the range of Rs. 300 to 600/kg. The
policy helped to lay a new focus on the species leading to the revival the aquaculture for
improving the overall pearlspot production in the state. The year 2015 will go down in the
history of Indian brackishwater aquaculture for another significant milestone achieved- the
first successful captive breeding of the marine herbivorous fish species, the milkfish Chanos
chanos by ICAR-CIBA. The herbivorous species forms the mainstay of brackishwater finfish
aquaculture of many south-east Asian countries such as Indonesia and Philippines. Farming
of milkfish can play a significant role in the development of sustainable models of
brackishwater aquaculture in India, in the days to come. In the year 2016, the captive
breeding of milkfish by CIBA was followed up by successful larval rearing and fry/fingerling
production and its distribution to farmers for growout culture, for the first time. Successful
results came from the efforts for transporting the batches of fertilized eggs of milkfish to
private entrepreneur and the subsequent rearing of milkfish larvae to fry stage and its sale to
growout farmers. Grey mullet Mugil cephalus by virtue of occupying lower trophic levels of
the food chain and at the same time having a high market value is a species of significance
for sustainable aquaculture systems like the IMTA (Integrated Multi-trophic Aquaculture)
and finfish polyculture. We are still reliant on wild seed resources for its aquaculture the
availability of which is today getting limited due to deteriorating nursing grounds and
emerging regulations on wild seed collection by states like Kerala. Hence ICAR-CIBA has
laid high impetus on developing a technology for grey mullet captive seed production.

3 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

Brackishwater ornamental aquariculture is also given a special focus and being developed as
new area on account of its potential to play a significant role in providing livelihood to small
scale farmers and Self Help Group’s, by adopting the homestead rearing model. CIBA has
bred spotted scat, Scatophagus argus, Moon fish Monodactylus argenteus for the first time in
the country. Species such as orange chromide, crescent perch, pearlspot are also being
developed as ornamental fish production models, suitable to get regular monthly incomes to
farmers. CIBA has been successful in breeding and successfully developing farming models
of the catfish Mystus gulio which commands good market value in the states such as Bengal.

Brackishwater Finfish aquaculture- The Present Scenario

Asian seabass has been mooted as a prime candidate for diversification to finfishes on
account of its high market demand (Rs.300-400 per kg), and availability of hatchery
produced seed and formulated extruded feeds. CIBA has achieved an average larval
survival rate of 30% and a maximum survival of 62% (Arasu et al., 2012) which is at par with
global standards. The global Asian seabass production from aquaculture was to the tune of
71,851 t in 2014. Globally seabass is cultured in different culture systems like cage, ponds,
pens and tanks showing the adaptability of the fish to the different culture methods. In India
seabass is primarily cultured in brackishwater pond systems where it exhibits growth rates
ranging from 350 g- 1.5 kg, in a period of 6 month to 1.2 years. Partial harvesting of larger
fish is carried out periodically in this extensive farming practice. Monoculture of seabass is
practiced at stocking densities of 10000-20000 ha-1 (initial wt.-2.0 g) and a production of 2.5-
5 t ha-1 with a survival of 30-70 % is observed in 10 months. The cost of production is around
Rs. 175-225/kg at a fish sale price of Rs 350-400. To develop a holistic model of seabass
culture, a formulated seabass feed, ‘Seabass Plus’ has been developed by CIBA for nursery
and growout culture which gives an FCR of 1.5-1.7 and the feed cost is approximately Rs.
55/kg. Presently, seabass is considered as one of the most potential finfish aquaculture
candidates for those looking for diversification from shrimp culture.

Herbivorous species form the backbone of sustainable fish farming models due to the
lower cost of production- milkfish, grey mullets and pearlspot have been traditionally
cultured by enhancing the ponds natural productivity and through low cost supplementary
feeding using agro-by-products. CIBA’s effort on the seed production of these fish species
bore fruit with the captive breeding of milkfish at its Muttukadu Experimental Station (MES).

Training Manual on Seed Production and Farming of Brackishwater Finfishes I 4

Today hatchery produced milkfish seeds distributed to brackishwater farmers across the
country are being cultured using growout formulated feeds specially developed for milkfish.
Interventions in parental care of pearlspot has assured enhanced seed production and given
rise to a modular tank based system for seed production for adoption by small scale farmers.

Few models of finfish culture being given key focus by CIBA

In keeping with its slogan “Brackishwater aquaculture for food employment and prosperity”,
icar-CIBA recognizes that brackishwater aquaculture is a powerful tool to bolster livelihood
and nutritional security. Hence, focus has been placed by CIBA to develop location specific
need based models for different stakeholders.
i) Satellite based nursery rearing of seabass - One of the interesting innovative
approaches adopted by CIBA for developing seabass as a separate livelihood
activity is the hapa based nursery rearing model where hatchery reared fry are
cultured for a duration of about 45 days to fingerlings size. This model is being
mooted both as a livelihood activity for farmers for giving returns in a short
duration and also for developing satellite seabass seed rearing centres to facilitate
widespread adoption of seabass aquaculture. This helps in saving space and time
during the grow-out culture of seabass and helps farmers to tide over the phase
which requires physical labour for frequent size grading of the fry. Adoption of
nursery rearing of seabass fry by farmers involved in low volume cage culture of
Asian seabass Lates calcarifer is encouraged by ICAR-CIBA. The activity helps
farmers get a better control over the initial size used for stocking in cages, a factor
critical for getting optimum survival rates and fish production. A farmer having a
1 ha pond, can stock 50,000 numbers of seabass fry in 50 hapas (2x1x1m).
Considering a seed cost of Rs 2,50,000 (Rs 5 per unit), feed cost- Rs 50,000,
installations and labor Rs 1 lakh and a final survival rate of 70 %, amounting to a
revenue of Rs 5.25 lakh (fingerling sale price Rs 15 per unit) , a benefit cost ratio
of 1.31 is worked out for the seabass nursery rearing activity.
ii) Low volume cage culture - The access of small scale farmers to diverse
opportunities offered by different aquaculture initiatives is often limited by
ownership or access to water resources, access to simple and adoptable technology
and high investment costs. Production of high value fish using low volume cages
set in brackishwater bodies can thus be a potential livelihood option to the poor.
Low volume cages can be fabricated by the farmers themselves. Species like

5 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

 seabass stocked at rates of 30-40 fry per cubic-m has shown to yield a production
upto 20kgm-3. For example, in a demonstration using 3 cage units of 8 m3 each, a
production between 450-500 kg of seabass was obtained Partial harvest of the fish
from the cages can thus provide the family with sustained monthly income of Rs
10,000-15,000 by sale of the fish at Rs 400/kg. Construction and setting up of
these low volume cages is also being mooted as a skill development activity for
small farmers.
iii) Integrated Multi-Trophic Aquaculture (IMTA) - IMTA is the farming of
aquaculture species from different trophic levels and with complimentary
ecosystem function. For laying a roadmap for sustainable aquaculture, CIBA is
developing economical polyculture models and adopting IMTA (Integrated Multi-
Trophic Aquaculture) approach. These models have been successfully
demonstrated at Kakwdip, West Bengal and Sindhudurg, Maharastra with farmers
participation. The benefit cost ratio of pond based IMTA was worked out to be 1.5
as compared to 1.4 in monoculture of shrimps at the culture demonstrated at
Sindhudurg district of Maharastra. Culture demonstrations at Kakwdip centre of
ICAR-CIBA has shown the environmental and economical benefits of IMTA over
conventional culture practices.
iv) Zero stocking models - CIBA also realises the relatively longer culture duration of
finfish compared to shrimps as a possible stumbling block for adoption of finfish
aquaculture. For this, CIBA advocates adoption of a multi-phased model for
finfish culture. A model has already been tested with seabass where the long
culture duration has been split into three phases- the nursery, pre-growout and
growout phase.
v) Finfish seed production from egg stage – Realising the need for elaborate
investment and infrastructure for maintenance of broodstocks of finfishes, private
entrepreneurs are encouraged to transport fertilized eggs for hatching and
subsequent larval rearing in their hatcheries. This model has been successfully
adopted in case of candidate species like seabass and milkfish by private
entrepreneurs.
vi) Ornamental fish seed rearing as a household activity - CIBA is mooting the
adoption of nursery rearing of ornamental fish like orange chromide, pearlspot,
and crescent perch as an activity to be adopted at household levels for getting

Training Manual on Seed Production and Farming of Brackishwater Finfishes I 6

 regular monthly income. As a part of the Mera gaon mera Gaurav program of
CIBA, pearlspot nursery rearing as alivelihood activity is being successfully
adopted by tribal women groups.
Way forward for brackishwater finfish aquaculture development and CIBA’s
interventions
For the development of brackishwater finfish aquaculture, mass availability of
hatchery produced seed during at the right time in the required quantity continues to be the
single major constraint. For this CIBA has placed a major thrust on developing economically
viable seed production technologies of the prime brackishwater candidate species such as
seabass, cobia milkfish and grey mullet. The development of hatchery technologies are also
being followed by partnership with entrepreneurs for facilitating technology adoption in
different states in PPP mode, this is witnessed in the partnerships both for shrimp and finfish
species. There is a need of private sector hatcheries for the scaling up of seabass seed
production for catering to the increasing demand of seabass and milkfish fry among
brackishwater fish farmers.
For the development of large scale economical finfish aquaculture practices,
development of efficient, eco-friendly and low cost feeds is perceived as the next major
challenge. The problem has aggregated with our dependence on fishmeal as a primary protein
source in feeds. CIBA has developed feeds for Asian seabass and other major species are
being tested. By entering into partnerships CIBA is also facilitating the development of feed
mills and developing feeds for different brackishwater foodfish and ornamental species.
Considering the significance of aquaculture as a tool to alleviate poverty, provide
livelihood and nutritional security, CIBA lays a major thrust on developing family farming
models for widespread adoption in different states. Thrust is also being laid for developing
suitable marketing models for getting the best price for the farmer.
CIBA visualises and strives for a holistic sustainable development of the
brackishwater aquaculture sector of India with an underlying thrust on sustainability,
economic viability and livelihood provision. For this CIBA reaches out for active
partnerships with the state government, the private sector, other research organisations and
Self-Help-Groups to develop and advance the brackishwater aquaculture technologies for the
betterment of the Indian brackishwater aquaculture sector.

7 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

References
Ali, S.A., Ambasankar, A., Syama Dayal, J., Thirunavukkarasu, A.R., Kailasam, K.,
Sundaray, J.K. Ponniah, A.G. (2012) Seabass feed technology-CIBA Bhetkiahar CIBA
Technology Series No. 6.
Arasu A.R.T, Kailasam, M., Sundaray, J.K., Subburaj, R., Thiagarajan, G. and Karaiyan, K.
(2008) Improved hatchery technology for Asian seabass lates calcarifer (Bloch). CIBA
Special Publication. No. 34.

Baiju A. (2010) The impact of technology adoption and policy interventions in the
popularisation of brackishwater cage culture in Kerala. M.Phil. Thesis (Kerala University).
Biology, fishery, culture and seed production of the pearlspot Etroplus suratensis (Bloch)
(1995). CIBA Bulletin No. 7.
Biswas and Sundaray (2009). Status of Asian seabass Lates calcarifer culture. In Training
manual on Seed production and culture of brackishwater finfishes with special reference to
Asian seabass Lates calcarifer. pp:157-160.
Cadalmin CMFRI Newsletter No. 130: 7. Pearlspot - the State Fish of Kerala: Mass seed
production to cage culture - A glance.
CIBA annual report 2002-03 Central Institute of Brackishwater Aquaculture (ICAR),
Chennai. pp:22-23.
CIBA annual report 2007-08 Central Institute of Brackishwater Aquaculture (ICAR),
Chennai. pp: 23-25.

CIBA Annual report 2011-12- Central Institute of Brackishwater Aquaculture (ICAR),

Chennai pp: 21-28.
CIBA Bulletin No. 7. (1995). Biology, fishery, culture and seed production of the pearlspot
Etroplus suratensis (Bloch)

CMFRI, Kochi (2013) CMFRI Annual Report 2012-2013. Technical Report. CMFRI, Kochi.
pp-104.
George, KC (1971) Salt water fish farming. Seafood Export Journal, 7(11): 7-14.

J.B. Kaiser, J.G. Holt. (2007) Cultured Aquatic Species Information Programme.
Rachycentron canadum. Cultured Aquatic Species Information Programme. Text by. In: FAO
Fisheries and Aquaculture Department [online]. Rome.

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Kaiser, J.B. and Holt. JG (2007) Cultured Aquatic Species Information Programme.
Rachycentron canadum. Cultured Aquatic Species Information Programme. In: FAO
Fisheries and Aquaculture Department [online]. Rome.

Krishna Sukumaran, Arasu A.R.T., Natarajan, M., Kailasam, M., Sundaray, J.K., Prem
Kumar, Subburaj R., Thiagarajan G., Venu S. (2013).Seed production of pearlspot Etroplus
suratensis (Bloch) in small net cages. Book of Abstracts: National seminar on emerging
trends in Indian Aquaculture, 28-30 March 2013. pp:12-13.

Krishnan, R., Neetha Susan David, Krishna Sukumaran, Pramod Kiran R.B. (2013). Milk fish
culture- an emerging brackishwater aquaculture practice in Kerala. National seminar on
Emerging trends in Indian aquaculture. 28-30 March 2013, Dept. of Aquatic Biology,
University of Kerala, Trivandrum, pp:20
Padmakumar K.G. , Bindu L., Manu P.S. (2012) Etroplus suratensis (Bloch), the state fish of
Kerala. Journal of Biosciences 37(1): 925-931.

Padmakumar, K.G.; Manu, P.S.; Bindu, L. (2009) Open water farming of pearlspot Etroplus
suratensis (Bloch) in low-volume cage. Asian Fisheries Science 22( 2): 839-847.
Pillai SM, Krishnan L., Venugopal N., Sasidharan CS (2002). Traditional system of
brackishwater aquaculture of Kerala. CIBA Bulletin No 14.
Rao, G.R.M. and Ravichandran,P. Sustainable brackishwater aquaculture. In Sustainable
Indian Fisheries, Pandian T.J. (ed.) 2001 pp-134-151.
Thirunavukkarasu ART, Kailasam M, Sundaray, J.K. (2012). Nursery rearing. In Training
manual on Seed production and culture of brackishwater finfishes with special reference to
Asian seabass Lates calcarifer. pp:47-48.

Thirunavukkarasu ART, Kailasam M, Sukumaran K., Prem Kumar, Subburaj, R.,

Thiagarajan, G., 2012. Diversification of species for sustainable aquaculture. In: Sustainable
shrimp farming through adoption of better management practices and biosecurity pp: 22-29.
Thirunavukkarasu,A.R. A, Mathew and M.Kailasam 2004. Hand book of seed production and
culture of Asian seabass Lates calcarifer (Bloch), CIBA Bulletin, 18 : 1-58.

9 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

 An overview of induced breeding techniques of brackishwater finfishes
M. Kailasam, S.N.Sethi, G.Biswas, K. Sukumaran, Prem Kumar, Aritra Bera, Babita Mandal

Introduction

Successful aquaculture largely depends on the availability of sufficient quality seed at

the required time. Availability of quality seed from natural sources is always erratic and
undependable. Moreover collection of wild seed will deplete the natural fishery. Almost all
of the cultivable brackishwater finfishes do not breed in captivity even though they attain
gonadal maturity. Hence it has become necessary to go for induced breeding either by
reproductive hormonal or environmental manipulation. Artificial spawning was first
achieved in Italy during 1930 in striped mullet. Use of hormones to induce fish to spawn was
started in Brazil in 1932. Compared to the advancement made in the breeding and seed
production of freshwater fishes, the technology development in brackishwater fishes
especially in India is far behind and this is to some extent are due to the non-availability of
facilities for the development of captive broodstock and lack of expertise.

Selection of breeders

Breeders can be obtained either from wild or from broodstock developed in captivity.
One of the problems faced in induced breeding is that variations occur in the gonadal
development among individual fish both in the wild and in broodstock developed in captivity.
Successful induced breeding depends upon the selection of the recipient fish at the proper
stage of the gonad development. Normally, the external characters like fullness of belly,
colour and state of swelling of genital opening such as protruding pinkish/reddish, genital
papilla, softness and resilence of the belly (in females), roughness of pectoral fins, presence
of hard tubercles (in males) etc. were considered for the selection of breeders. However,
many of these parameters are not absolutely reliable. For example, enlargement of belly can
be due to presence of food in the intestine and stomach. The more reliable method to assess
the maturity of females now being used is through ovarian biopsy taking a sample of the ova
using a catheter and to examine them under microscope. The mature ova will have round
shape and non-adhesive. The average ova diameter has to be determined and this is used as
an important criterion in the selection of females for induced spawning. In the case of males,
maturity is ascertained by applying pressure on either side of the belly. In the case of fish in

Training Manual on Seed Production and Farming of Brackishwater Finfishes I 10

mature condition, milt will be flowing through the genital opening on application of gentle
pressure.

Sex determination

Majority of sea bass in the size range of 1.5 to 3.0 kg are males and as they attain a
size of 3.5 to 4.0 kg, majority of them undergo sex change and become females. So, the size
of the fish is commonly used for the identification of the sexes. Otherwise sexual
dimorphism is not well marked and sex can be determined accurately only when they are in
mature stage. In mature males, milt will be extruding on application of pressure on the
abdomen. Females can be identified from the comparatively big soft round belly with pinkish
genital papilla. In fully mature female, eggs will be even visible when the abdomen is
pressed. There are some other minor identification marks. In males the snout is slightly
curved while that of the female is straight. The scales near the cloaca of males are thicker
than the scales in females during the spawning season. The body of males is comparatively
slender compared to females. In the case of other fishes like Cobia and mullet, females
appear with bulged soft belly with genital papilla. Males will be oozing while pressing the
abdomen in both the species and also in milkfish.

Methods of Breeding

There are three methods by which fertilized eggs are obtained and seed production is
done. They are artificial fertilization by striping of mature females and males, induced
breeding by reproductive hormone administration and breeding by environmental
manipulation.

Artificial fertilization by striping

In this method spawners are obtained from wild during the natural breeding season.
In seabass breeding is related to lunar cycle. Again breeding occurs before midnight during
high tide. Even though the fish breeds both during the new moon and full moon phases,
quality of eggs released during full moon phase is better and the number of eggs released also
will be more. Fishes caught during full moon and new moon phases and during high tide are
examined for maturity. Both males and females that are in oozing stage can be striped and
fertilized artificially. In oozing females the diameter of the eggs will be around 0.7 to 0.8
mm with large oil globule. The eggs will be almost transparent. The ripe eggs will scatter

11 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

individually whereas unripe eggs tend to group together in water. In water having salinity 28
– 30 ppt the ripe eggs will float.

For easy handling the selected females and males are anaesthetized. Eggs and milt
are stripped into a dry clean tray and mixed thoroughly with a feather. After 1 – 2 minutes,
fresh clean seawater of salinity around 30 ppt is added to keep all eggs floating and mixed
well for 2 – 3 minutes. Then the eggs are washed 3 to 4 times using a strain to remove all
mucus and other tissues. Thereafter the fertilized eggs are distributed to incubation tanks.

Environmental manipulation

This technique is usually followed in broodstock developed in captivity. About a

month prior to the spawning season, the mature females and males are transformed to
spawning tanks at a density of 1 kg/m3. The salinity of the broodstock tank and spawning
tank should be same. After 2 to 3 days when the fish got acclimatized to the spawning tank
conditions, the salinity of the water is reduced to around 24 ppt. The fishes are maintained in
this condition for about a week and then the salinity is gradually increased to 30 to 32 ppt by
daily water exchange over a period of 10 days. This increasing of salinity simulates the
condition similar to that of the migration of the fish from low saline feeding ground in the
brackishwater to the high saline spawning ground in the sea and stimulates breeding.

On the ensuing full moon/new moon day, the water level is reduced to about 30 cm
during noon time and the water temperature is allowed to go up to above 30 0C. By dusk
fresh sea water is added to the spawning tank to simulate the rising tide conditions and
simultaneously water temperature also declines to around 270C. The fish that is in right stage
and good condition will spawn in the same night or during the subsequent night. The fish
would continue to spawn for 3-5 days after the first spawning provided the environmental
factors remain conducive. Seabass being an intermittent spawner releases eggs in batches;
the same spawner will continue to spawn during full moon or new moon for the next 4-5
months. The fish that have not spawned can be subjected to induced spawning by hormone
administration.

Induced Spawning

Seabass does not spawn in the broodstock tanks normally. Administration of

reproductive hormones becomes necessary for inducing them to spawn. Human chorionic

Training Manual on Seed Production and Farming of Brackishwater Finfishes I 12

Gonadotropin (HCG), Puberogen, Pregnyl and Luteinizing hormone – releasing hormone
analogue (LHRH-a) are the main reliable synthetic hormones that are used for induced
breeding.

The fishes that have to be induced are transferred from broodstock tanks to pre-
spawning tank 2 months before the breeding season. These fishes are checked at fortnightly
intervals to assess the maturity condition. The maturities of females are examined by taking
out a sample of the eggs using a polyethylene cannula of 1.2 mm diameter. To avoid any
handling stress, the fish is anaesthetized before the eggs sample is taken. Otherwise the head
of the fish is inserted in a loose perforated plastic hood. The hood will extend upto the
middle of the body. The fish is kept upside down keeping the head in water and the cannula
is inserted into the oviduct. Since seabass releases 3-4 batches of eggs during the spawning
process at definite intervals, it is clear that all the eggs in the ovary will not be in the same
stage of maturity. Since the eggs in the posterior end of the ovary will get released first they
will be in a more advanced stage of maturity compared to the eggs in the anterior region.
Hence it is essential that the eggs in the posterior end are sampled while examining the
maturity condition by inserting the cannula for a distance of 3-4 cm from the cloaca. The
other end of the cannula is held in the mouth of the operator and the eggs are aspirated into
the tube by the operator. When the eggs enters the cannula, the cannula is slowly withdrawn
and empty the eggs slowly by the operator to a clear petri dish containing clean seawater and
the diameter of the eggs are measured under a microscope using an ocular micrometer.
Mature eggs get scattered around once it is transferred to a petri dish having water. Females
that are having eggs of 0.4-0.5 mm average diameter can be given hormone treatment for
induced breeding. Males with oozing milt are taken for breeding.

At Central Institute of Brackishwater Aquaculture, Chennai, des – Gly 10 (D-Ala 6)

luteinizing hormone releasing hormone ethylamide acetate salt (LHRH-A) hormone is used
for the induced breeding of seabass. Breeding is normally taken up on new moon or full
moon nights. Female and male breeders are selected in the ratio 1:2 in the broodstock tanks
and transferred to the hatchery. Their total length and weight are recorded and also
ascertained that they are in good health condition. LHRH-A is administered to females and
males @ 60 – 70 ug/kg body weight and 30 – 40 ug/kg body weight respectively and
transferred to the spawning tank. Water salinity 30 – 32 ppt was found to be optimum for
spawning. The breeders should be free from disturbances like excess noise and human

13 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

movements. They spawn after 30 – 36 hrs of hormone administration. The spawning may
continue for a week releasing 3 – 4 batches of eggs.

In the case of grey mullet Mugil cephalus, the first maturity can be observed in 2-3
years old fish. In natural condition, mullet maturation and spawning noticed during October
to January in the east coast of India and during June-July in the west coast. Longer darker
period and low temperature directly linked with the maturation of M. cephalus. Females
with initial oocyte diameter of 600 µm and oozing males can be selected for induction of
spawning through hormonal manipulation. Carp Pituitary extracts and LHRHa @ 20mg/kg
and 200µg/kg body weight are used as priming and resolving doses for spawning. After
ovulation, stripping of ovulated eggs is common practice followed. The stripped eggs are
fertilized by mixing with milt obtained from males using bird feather by dry method. The
floating fertilized eggs can be stocked in the incubation tanks for hatching. The newly
hatched mullet larvae can be stocked in the larval rearing tanks to grow them to fry size in the
hatchery

Milkfish mature in seawater at the age of 3 years. However, broodfishes with age of 5
plus years are usually selected for breeding purposes. Milkfish require higher temperature
and longer day period for maturation, which is usually coincide with summer period.
Milkfish can be bred through LHRHa hormone treatment @ 50 µg/kg body weight either
with pellet implantation and intramuscular injection. The hormone treated milkfish spawn
spontaneously in the tanks and fertilized eggs are pelagic and float in the water. The fertilized
eggs hatch out between 22-24 hours of incubation period and the newly hatched larvae can be
stocked in the larval rearing tanks for fry rearing.

Cobia is one of the most preferred marine fishes in the cages because of its rapid
growth rate. The fish can grow 4-6 kg in one year under ideal condition in the cages. It can be
cultured in deeper ponds with good water exchange. Cobia tolerates the salinity range from
15 to 35 ppt. It is widely farmed in Vietnam, Mexico, USA, Taiwan, China and other South
East Asian countries. Cobia matures after attaining the age of 3 years. Sexes are separate. The
females, which are having the initial oozyte diameter of 700 µm are considered ready for
hormone induction. By applying hormone treatment with HCG @ 250-500 IU/kg body
weight, cobia can be induced to spawn. Cobia larvae reach to three inch size fingerlings in 45
days period rearing in the hatchery and these fingerlings can be stocked in the cages or ponds
for grow out culture.

Training Manual on Seed Production and Farming of Brackishwater Finfishes I 14

 Biology of milkfish (Chanos chanos)
Aritra Bera, Babita Mandal, Tanveer Hussain, T. Sivaramakrishnan
Introduction
Milkfish Chanos chanos) is one of the most popular cultivable brackishwater finfishes in the south
east Asian countries and widely distributed in the Indo-Pacific region. The maximum weight and age
of this fish were reported as 14kg and 15 years respectively. In India, it is named as Paal Meen in
Tamil, Pala Bontha and Tulli Chepa in Telugu, Poomeen in Malayalam, Hoomeenu in Kannada,
Golsi in Goa and Seba khainga in Oriya. Being herbivore, milkfish feeds on plankton, benthic algae,
detritus matter in the natural condition and easily accepts the pellet feed under culture condition. It
can tolerate and live in extreme salinity ranging from 0-100 ppt but growth is optimal between 0.5-40
ppt. Milkfish can attain the table size weight from 400 to 500 gm in 5-6 months under culture
condition. Milkfish having tiny bones resemble with Hilsa and can be considered as a
‘Decan Hilsa’. Milkfish can be produced in the farm with the production cost of Rs.80-90/kg
by feeding with low protein pellet feed.

Kingdom – Animalia
Phylum – Chordata
Order – Gonorynchiformes
Family - Chanidae
Genus – Chanos
Species - Chanos chanos

Adult milkfish (50-150 cm total length) are powerful swimmers and mostly found in open sea. During
the breeding, it migrates towards reefs and sandy-rocky shores for breeding. Milkfish fertilized eggs
appear slightly yellowish (size 1.10 - 1.25 mm). Incubation period is ranged 24-26 h. Milkfish fry are
most abundant in shore waters. Large juveniles and sub-adults return to sea. During 1986, the
Southeast Asian Fisheries Development Center (SEAFDEC) at Phillipines under National Bangus
Breeding Program successfully established milkfish seed production technology. Global production
of milkfish is estimated to be 9 lakh metric ton during 2012 Philippines is the leading producer of
milkfish with 391,983 MT during 2014 apart from Indonesia and Taiwan.

Wild milkfish seeds are collected during March to May from Andhra Pradesh and Tamil Nadu coasts
by traditional methods and farmed in the coastal ponds along with other finfish and shell fishes.
ICAR-CIBA has made major breakthrough on captive breeding of milkfish for the first time
in India during June 2015 and developed comprehensive technology package for seed
production of milkfish. Captive breeding of milkfish involves development of land based

15 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

captive broodstocks (6+ years old) and application of calculated dose of slow release
hormone (LHRH-A) pellet. Hatchery produced seeds were distributed among farmers for
promotion and demonstration of milkfish farming in coastal states. To educate the farmer
community about this technology, routine trainings are being organized by ICAR-CIBA on
need-based.

Life history and habitat

Adult milkfish (50-150 cm total length) are found in open sea, swift and powerful
swimmers. During the breeding season, they migrate towards coasts where reefs and sandy-
rocky shores for breeding. Spawning takes place in the open sea and the eggs are pelagic.
Milkfish fertilized eggs appear slightly yellowish (size 1.10 - 1.25 mm). Eggs float at 34 ppt
and tend to sink at <30 ppt salinity. Incubation period is ranged 20-35 h at temperatures of
26-32°C and salinities 29.5-34 ppt. newly hatched larvae measure 3.5 mm TL at hatching,
have a large yolk sac (volume, 0.5 μl), unpigmented eyes, and no mouth. They grow to about
5 mm in 36 h after consuming about 90% of the yolk. Milkfish fry of 10-17 mm are most
abundant in shore waters. The larvae and juveniles spend their lives in inshore estuarine areas
and then migrate into rivers in the direction of fresh water (Lin et al., 2003; Pillay and Kutty,
2005) for further growth. Milkfish juveniles larger than 20 mm have the characteristic shape
and morphology of the adult fish.

Source: Life cycle and habitat of milkfish in wild (Martinez et al., 2006)

Training Manual on Seed Production and Farming of Brackishwater Finfishes I 16

Distribution worldwide

Milkfish can tolerate wide ranges of salinity and temperature fluctuations (Gordon and Hong,
1986). Milkfish occurs near coasts and islands in the tropical Indo-Pacific area and is
centered around the Philippines and Indonesia. It inhabits areas of the Pacific Ocean which is
affected by warm ocean currents. Their distribution can be found near to coral reef areas
where the water is clear, shallow and warm (more than 20°C). Milkfish has been recorded in
the Red Sea, off the coast of East Africa, including Zanzibar and freshwater lakes in
Madagascar, Mexico, coastal parts of India, Sri Lanka, Malaysia, Vietnam and Thailand.
Milkfish is abundant in Kiribati, Fiji, Tonga, and New Caledonia. In Kiribati milkfish has
been produced mainly as bait fish for supporting the tuna fishing industry.

Fig: Worldwide Milkfish Distribution

Distribution in India

Initial observations on the fry of milkfish in India were made by Panikkar et al. in 1952.
Adult milkfish from has been reported from CoroMandal Coast, Palk Bay, Gulf of Mannar
andMalabar Coasts (Chacko and Thomas, 1962), Salt pans of Kakinada (Dwivedi and Reddy,
1977), Pulicat lake (Rao, 1970), Vishakhapatnam, Krusadai and Rameswarm Islands (Chacko
and Mahadevan, 1956), Bakkhali region of lower Sunderbans (Basu and Prasad, 1976). On
the basis of occurrence of adult milkfish, spawning ground has been classified as Madras,
Mandapam, Tuticorin, Vizhinjam, Cochin, Calicut zone (Silas et al., 1985). Estuaries and
mangroves areas were recorded with abundance of fry after spawning season.

17 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

Source: Proceedings of the symposium on coastal aquaculture (Silas et al., 1985).

Food and feeding habits

The morphology of the digestive system of adult and juvenile milkfish suggests that it
is mainly an herbivore (Chandy and George, 1960; Kinoshita, 1981). It has a toothless mouth
with fine closely laid gill rakers and a pair of muscular rakerlined with epibranchial organs.
The esophagus is long and thick-walled. The stomach is large and the cardiac region is
characteristically bent (doubled-over). The pyloric region has a spherical gizzard with a
mucus membrane and very thick walls. The cardiac stomach has gastric glands. The gizzard
seems to function in trituration of coarse food materials. Numerous pyloric caeca clusters can
be found behind the gizzard. The intestine is extremely long and convoluted. Adult milkfish
can be kept in captivity on a diet of commercial pellets with about 42% protein given at 1.5-
2% of body weight twice daily (Marte and Lacanilao, 1986). After full pigmentation of
milkfish fry eye, mouth opens at 54 Hour after hatching. The yolk is completely adsorbed
within 120 hour post hatching. After three days of hatching, esophagus and intestine can be
differentiated. Mouth size is 200 μm, after opening the mouth of milkfish larvae (225 μm
body sizes). Copepods and Diatoms are the main feed for milkfish fry in coastal waters.
Larvae are visual feeders. In hatcheries larvae are fed with rotifer and Artemia nauplii and
later weaned on artificial larval diets. Milkfish fry cannot digest rigid cell wall of chlorella in
juvenile stage. After 2 weeks, Non-live feed can be given to larvae. 40% weaning can be
achieved with finely ground artificial diets at juvenile stage. In nature, juvenile milkfish is a
bottom feeder (iliophagous). It ingests the top layer of bottom sediments. Digestibility of fish
meal and soybean meal is lower in seawater as compare to freshwater. Therefore, food

Training Manual on Seed Production and Farming of Brackishwater Finfishes I 18

conversion efficiency in milkfish got decrease and protein requirements increase in seawater
conditions. Being an opportunistic filter feeder, adult milkfish mainly feed on zooplankton,
larval and juvenile clupeiods in wild.

In artificial intensive culture systems, milkfish require 30 % or more protein and a

lipid level of not less than 7%. Milkfish broodstock requires 1000 mg Vit. C/kg feed for good
egg and larval quality. Adult milkfish accepts a variety of diets like sinking particles and
floating pellets. Use of pellets improves the feeding efficiency. Pellets are having physical
characteristics such as better stability which prevents feed particles from dissolving and
leeching of nutrients in the water. Adult fishes are fed with pellets of approximately 4-5 mm
(dia) and 6-8 mm long.

Growth

However, growth rates in ponds and in pens vary considerably depending on initial
fish size, food, stocking density, climate, season, locality, water turnover rate, pond area and
depth, pests and predators. The growth curve of milkfish larvae is sigmoid. Some reports
have found that hatchery-bred and reared milkfish fry are generally heavier and
morphologically more advanced (heavy pigmentation, pelvic fins present) than shore-caught
fry of similar length. Growth rates of wild and pondreared juvenile milkfish vary from 7.0 to
8.7 mm weekly.

Growth and Reproduction:

Larval stages of Milkfish exhibit sigmoid growth curve. Hatchery-reared milkfish fry tend to
be growing fast and morphologically more advanced than wild caught fry of similar length.
Adult Milkfish grows very well in ponds, compared to other cultured finfish and shellfish
species. Growth rates in ponds and in pens vary considerably depending on initial fish size,
food, stocking density, climate, season, locality, water turnover rate, pond area and depth,
pests and predators. Sexes are separate in milkfish but morphologically identification is
difficult. Ripe male oozes milt when pressure is applied to the abdomen. In case of gravid
female fishes, abdomen may appear distended. Mature ovary of female fish grows 10 – 25 %
of body cavity. Milkfish do not spawn in freshwater. Milkfish have been observed to spawn
at water temperatures of 24-33°C. In nature, spawning of milkfish has been observed during
the first quarter and full moon periods. Peak season of spawning lies during March – June
which again varies according to geographic regions. A 5-13 kg female can produce 3, 00,000
– 5, 00,000 eggs/kg body weight.

19 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

Larval Stages
Milkfish larvae are constituents of open sea plankton. Newly hatched larvae measure 3.5 mm
TL at hatching, have a large yolk sac (volume, 0.5 ml), unpigmented eyes, and no mouth.
They grow to about 5 mm in 36 h, consuming about 90% of the yolk, and grow very little
until day 5 when the yolk is completely exhausted . Egg size, larval size, amount of yolk, and
mouth size are greater in milkfish than in many other tropical marine fishes (such as the sea
bass, Lates calcarifer and the rabbitfish, Siganus guttatus. This size advantage is probably
one reason for the relative ease in rearing milkfish larvae in the hatchery and for the
abundance of milkfish fry in the wild. Total lengths are given for larvae from the plankton
measured in the preserved condition and for larvae reared in the laboratory and measured in
the fresh state (in parentheses).

Fry and the Transformation Stage

Milkfish fry are most abundant in shore waters and constitute a valuable fishery that supports
the centuries-old pond culture industry in southeast Asia and India. Fry caught in shore
waters are all within a narrow range of 10-17 mm TL. The transformation stage, also termed
the "metamorphic stage," is a complex of morphological developments accompanied by
many physiological and behavioral changes

Juveniles
Milkfish larger than 20 mm acquire the characteristic shape and morphology of the adult of
the species and are considered juveniles. Juveniles <10-cm long arc usually called
fingerlings. They bear complete fin-ray complements, forked caudal fin, scales, and silvery
coloration. Juvenile milkfish have been found in such diverse habitats as coral lagoons,
mangrove lagoons, estuaries, marsh flats, tidal creeks, and tide pools that share the common
characteristics of rich food deposits and protected, relatively shallow waters

Training Manual on Seed Production and Farming of Brackishwater Finfishes I 20

 Seed production of Milkfish (Chanos chanos) – Global scenario
Babita Mandal, Aritra Bera, G. Biswas, Prem Kumar, M.Makesh,
S.N. Sethi, Tanveer Hussain, Dani Thomas, Pankaj Patil

Brackishwater finfish culture depends upon the availability of fry or fingerling for
stocking. In past, these seeds were collected from wild sources but with the domestication
and fruitful research experience, breeding and seed production was achieved for few finfish
species to produce reliable source of seeds. Seed production is an important activity of any
hatchery which needs knowledge of fish biology, its spawning behavior, nutritional
requirement, heath aspects etc. It also includes activities related to larval biology, nutritional
requirement and culture till it attains stock able size.

Culture of Milkfish (Chanos chanos) is an age old practice among south East Asian
country and in few states of India also. Characteristics like euryhaline nature, rapid growth,
herbivores feeding nature are the major reasons which make it an ideal candidate species for
culture. Maturation has been reported in varying salinities (0 – 125 ppt). In traditional culture
practices, wild caught fry were cultured in different culture systems of fresh water,
brackishwater and marine waters. Initial observations on the fry of milkfish in India were
made by Panikkar et al. (1952). Adult milkfish from has been reported from CoroMandal
Coast, Palk Bay, Gulf of Mannar and Malabar Coasts, Salt pans of Kakinada, Pulicat lake,
Vishakhapatnam, Krusadai and Rameswarm Islands, Bakkhali region of lower Sunderban.
On the basis of occurrence of adult milkfish, spawning ground has been classified as Madras,
Mandapam, Tuticorin, Vizhinjam, Cochin, Calicut zone. Estuaries and mangroves areas were
recorded with abundance of fry after spawning season.

Till 1970’s source of milkfish seed for nursery rearing and growout culture was wild
sources. But Rapid increase in milkfish farms created the need of controlled breeding units
for healthy and reliable seed supply. Since that time research was focused in South East
Asian countries for breeding in sea cages with the help of hormone manipulation in adult
fishes. As the fishes mature in +5 years in case of males and +7 years in case of females,
creating a broodbank has been a challenged task. Broodfishes which were captured from sea
were maintained in sea cages for spawning.

Adult milkfish are found in open sea are powerful swimmers. During the breeding
season, they migrate towards coasts where reefs and sandy-rocky shores for breeding.
Spawning takes place in the open sea and the eggs are pelagic. Milkfish fertilized eggs appear

21 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

slightly yellowish (size 1.10 - 1.25 mm). Eggs float at 34 ppt and tend to sink at <30 ppt
salinity. Incubation period is ranged 20-35 h at temperatures of 26-32°C and salinities 29.5-
34 ppt. newly hatched larvae measure 3.5 mm TL at hatching, have a large yolk sac,
unpigmented eyes, and no mouth. They grow to about 5 mm in 36 h after consuming about
90% of the yolk. Milkfish fry of 10-17 mm TL are most abundant in shore waters. The larvae
and juveniles spend their lives in inshore estuarine areas and then migrate into rivers in the
direction of fresh water for further growth. Milkfish juveniles larger than 20 mm have the
characteristic shape and morphology of the adult fish.

Seed production in South East Asia (Philippines & Taiwan):

Milkfish aquaculture in the Philippines has expanded from the traditional culture in
brackishwater ponds and freshwater pens to marine cages and pens. Natural spawning using
pond-reared milkfish was achieved in Taiwan. The first instance was in August 1980, when
the milkfish broodstock in floating cages at SEAFDEC/AQD spawned spontaneously.
Broodstock are kept in deep concrete tank fitted with a recirculating system. An airlift PVC
pipe (inset, arrow) fitted with a fine mesh net bag collects eggs naturally spawned by milkfish
broodstock. Sex ratio of 2:1 (M : F) is maintained in tanks. Fertilized eggs are represented by
1.1-1.23 mm oocyte dia. Salmon or Carp pituitary homogenate in combination with HCG, or
HCG are used to induce the fishes for spawning. Numbers of hormone injections may be 1-5
(mostly 2) with injection interval of 6 - 24 hour (mostly 8-12 hour). Time to stripping is
commonly 6-17 hour (12 hour appears best) in case of oozing female and male. A 5 - 13 kg
female can produce 300000 eggs/kg body weights. In wild, milkfish spawns more than once a
year. Spontaneous spawning without hormone treatment has also been achieved with captive
broodstock maintained in floating net-cages in the Philippines.
Larval rearing of milkfish can be done in extensive/intensive systems for 20 – 30 days
before initiating nursery rearing. In hatcheries, Indoor larval rearing facilities are utilized for
rearing the larvae in green water systems. Rotifer and Artemia nauplii were provided upto 15
– 20 days and weaned to artificial diets later on.
Conclusion

Seed production of Milkfish in hatcheries was achieved in Taiwan which was adopted
by Philippines and Thailand for the production of milkfish larvae. In other countries, seed
were collected from natural sources. In India, first successful captive breeding was achieved
in the year of 2015 which has paved the way for hatchery production in India. Lack of
availability of hatchery reared seeds was felt since long back yet the seed production in
hatchery has been realized recently.

Training Manual on Seed Production and Farming of Brackishwater Finfishes I 22

 Induced breeding, larval rearing and seed production of Asian seabass
(Lates calcarifer)

R. Subburaj, M. Makesh, Aritra Bera, G. Thiagarajan, Raja Babu

Introduction

Successful aquaculture largely depends on the availability of sufficient quality seed at the
required time. Availability of quality seed from natural sources is always erratic and
undependable. Moreover collection of wild seed will deplete the natural fishery. Almost all of the
cultivable brackishwater finfishes do not breed in captivity even though they attain gonadal
maturity. Hence it has become necessary to go for induced breeding either by reproductive
hormonal or environmental manipulation.

Asian Sea bass (Lates Calcarifer) known as Bhetki is one of the commercially important
finfish species caught from inshore areas, estuaries, backwaters, lagoons and fresh water ponds.
Sea bass is a fast growing species with ability to tolerate wide fluctuations in environmental
conditions. It fetches fairly good price in the domestic market and there is a scope for export to
Middle East and some of the European countries. It is considered as a potential candidate species
for farming in saline or freshwater environments in ponds and cages. It is extensively cultured in
South East Asian Countries like Thailand, Malaysia, Singapore and Indonesia and in Australia.
However in India sea bass farming is very much limited. It is done traditionally in the coastal
ponds, this method of farming is neither advisable nor dependable and the availability of quality
seed in adequate quantity is one of the major reasons hampering seabass farming in India in a
large scale.

Seed being one of the major inputs for a sustainable farming, development of techno economically,
viable indigenous technology for large-scale seed production of sea bass has been taken up on priority at
the Central Institute of Brackishwater Aquaculture, Chennai and a technology package has been developed.

Biology of Sea bass

Sea bass is an euryhaline species capable of withstanding wide salinity fluctuations. It is a

catadromous fish migrating towards sea for breeding. It is a fast growing fish and suitable for culture
in marine, brackish and fresh water bodies.

Seabass grows in salt water and fresh water, it migrates to sea in the reproductive phase.
Seabass spawns in the sea not far away from the shores but of about 20-30 kms. The newly hatched
larvae are drifted into coastal waters and even enter fresh water areas through water currents

23 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

 .

In sea bass, sexes are separate; however, they are protandrous hermaphrodite. Most of
the fishes are males when they are small less than 3 kg and change to females when they grow
more than 4.0 kg. Seabass is a protracted spawner (i.e an individual fish can spawn two or three
times in a season).

SEED PRODUCTION TECHNOLOGY

For sustainable captive seed production the following are important requirements:

 Broodstock fishes
 Broodstock holding facilities
 Facilities for pumping quality seawater
 Maturation and spawning facility
 Egg Incubation
 Live feed culture system-Stock culture and mass rearing facilities
 Larval rearing facilities and
 Nursery rearing

Broodstock Development:

The success of seed production depends upon the development of viable captive
Broodstock and for a sustainable hatchery operation a viable land based brood stock has to be
developed.

Fish Collection and transportation

Fishes in the size range 1.5 kg to 3.0 kgs for males stock and 4 to 10 kg for female stock
can be collected either from wild sources or from the culture ponds. The fish should be healthy
without external or internal injuries and free from pathogens. If the sources of collection is very
near this hardy fish can be transported safely using open containers and from far away places,

Training Manual on Seed Production and Farming of Brackishwater Finfishes I 24

water tanker with oxygen bubbling is advisable with biomass @ 3-4 kg/m3 for a transportation
duration of 8-9 hrs under water temperature of 27-28oC.

Acclimatization

Seabass though in general a sturdy fish capable of withstanding environmental variations,

the broodstock fish should be kept captive condition with minimal stress to the extent possible.
As they are predaceous preferring to feed on live fishes and crustaceans should be acclimatized to
feed on inert diet. Fishes either procured and transported have to be ‘conditioned’ before they are
transferred to holding tanks/cages. Prophylactic treatments with fungicides, bactericides are done
to avoid manifestation of diseases. Fishes should be screened for pathogens and quarantined for
a period 7-10 days before they are transferred to brood stock holding facilities.

Broodstock Holding Facilities

Broodstock can be maintained in larger tanks or in cages or earthen ponds where good
quality seawater can be supplied. These facilities should have adequate water exchange system
and aeration. RCC circular or rectangular holding tanks of 12 x 6 x 2 m are desirable.

Broodstock Holding Tanks

25 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

Broodstock Maintenance

Under captive conditions Broodstock fishes can be maintained at a density of 1 kg/m 3.

Fishes in the size group of 1.5 to 3.0 kg to cater the need of males and 4 to 10 kgs for females are
maintained.

Water Quality Management

In the Broodstock holding tanks, adequate water exchange to an extent of 70-80% daily is
desirable. Since, Broodstock fishes are maintained specifically for maturation and spawning
required parameters stimulating the conditions prevailing in the sea would be very much helpful
to obtain better response. The following water quality parameters are desirable for maturation
and spawning.

TABLE: 1 WATER QUALITY PARAMETERS REQUIRED FOR MATURATION

1. Water Temperature 28 – 30oC

2. Salinity 28-32 ppt

3. PH 7.8 – 8.2

4. Dissolved Oxygen < 5 ppm

5. Ammonia > 1 ppm

6. Phosphate > 60mg/l

7. Alkalinity >100 ppm

Management of Feeding For Broodstock

Nutritionally balanced diet is an essential component in broodstock development since

the health of brood stock will only determine the health of their offsprings. Seabass is being a
voracious carnivorous fish in feeding namely in live fishes/shrimps seabass it may be reluctant to
feed on inert feed. However, they can be weaned to feed an frozen fish. To start with fishes are
fed with live and frozen fishes and slowly live fish is replaced completely with frozen feed.
Frozen fishes like oil sardines and Tilapia are fed @ 5% of the body weight of the biomass daily.

Training Manual on Seed Production and Farming of Brackishwater Finfishes I 26

Health Management of Broodstock

The most common problem that would be associated will be the ‘pathogens’. Common
pathogens like caligus sp, Lernonthropsis sp and the monogenic trematode Diplectnum latesi are
encountered in fishes during months of October to February.Broodstock fishes maintained in
captivity will be susceptible for stress. Pathogens inside the holding tanks will infect leading to
mortality. Treatment with 1 ppm organo-phosphorus Dichlorvos for one hour if the parasitic
infestation is high or 100 ppm formalin in the case of lesser infestation is found to be effective in
controlling the parasite infestation.

Maturation and Spawning

Fishes maintained with proper water quality management, feeding and health management
attains maturity under captive conditions normally during breeding season. For eg. at Chennai
fishes attain full maturation during the months of May to November provide the salinity regime is
more than 28 ppt. If the salinity reduces resorbtion of gonad is observed under captive conditions
indicating salinity is the major factor influencing the maturation. To evaluate the maturity stages
ovarian biopsy using cannula is done. The polythene cannula of 1.2 mm dia is inserted into the
oviduct and gently aspirated. The ova is then observed under microscope.
Seabass is a protracted spawner. An individual fish will have in the ovary eggs in
different diameter. This facilitates the fish to spawn more than once in the spawning season.
Females with egg size more than 0.450 mm is selected for induction of spawning. In the case of
males, if thick white viscous milt is seen by gently pressing abdomen such fishes are selected.

Induced Spawning

Seabass fish spawns spontaneously if the conditions could be stimulated as prevailing in

the sea. But it may not be that simple. In such cases, the hormone responsible for ovulation in
administered extraneously. For Seabass breeding the hormone LHRH-a, a product of SIGMA is
highly effective. Female fishes with ova size more than 0.450 mm and oozing males are selected
and a single dose of LHRH-a hormone is administered intramuscularly @ 60-70µg/kg body
weight for females and 30 -35µg/kg body weight for males preferably in the early hours of the
day.

27 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

 For Example

If the 1mg vial is dissolved in 5ml; Each ml will have hormone concentration of 200
µg. If the selected female fish weighs 6 kgs, at the dosage level of @ 70 ug/kg body weight,
the hormone requirement will be

70 x 6 = 420 µg for female fish

go from the vial _1_ x 420 = 2.10 ml has to be drawn for giving injection

200

For the males – if the weight of male is 3 kg each the requirement at a dose rate of 35 µg/kg
body weight will be 35 x 3 = 105 µg.

Then _1_ x 105 = 0.525 ml has to be injected to each male.

200

Hormone is taken in separate syringes for injection to female and male fishes.

The sex ratio is maintained Female : Male @ 1:2. spawning takes place simultaneously
sly after 30-36 hrs after the injection. Spawning takes place normally in the evening hours. For
better spawning and fertilization full moon or new moon days are preferable. Being a batch
spawner seabass spawns during subsequent days also. The fecundity normally ranges from 0.7 to
3.0 million under captive conditions in a single spawning. The fertilized eggs will measure about
0.78 to 2.0 mm in size and float on the surface and the unfertilized opaque eggs will sink at the
bottom. The fertilization rate depends upon the conditions of the spawner, which is upto 90%
under captive condition.

Egg Collection and Incubation

The floating fertilized eggs can be collected by air drifting method or by scooping method
or by over flowing method using suitable mesh size cloth. The fertilized eggs are transferred to
the incubation tanks for further development and hatching and lesser at density of 70-100 nos
eggs/l in the incubation tanks. The eggs will hatch out in 17 – 18 hours after fertilization
undergoing developmental stages as follows

Training Manual on Seed Production and Farming of Brackishwater Finfishes I 28

 Embryonic development Stages Duration
One Cell stage 30 minutes

Two Cell stage 40 minutes

Four Cell stage 45 minutes

Eight Cell stage 60 minutes

Thirty two Cell stage 2 hrs

Sixty four Cell stage 2 hrs 30 minutes

128 Cell stage 3 hrs

Blastula stage 5 hrs 30 minutes

Gastrula stage 6 hrs 30 minutes

Neurula stage 8 hrs

Early embryo 11 hrs

Heart functional and tail movement 15 hrs

Hatching 17 – 18 hrs

Undergoing different stages of development, the fertilized eggs hatch out after 15-17 hrs. The
hatched out larva of seabass measures about 1.2 to 1.4 mm in size with comparatively small yolk sac. The
hatching rate depends upon many factors like health of the spawners, fertilization rate, water quality etc.
Normally it is observed to be up to 85% under captive condition.

Larval Rearing

Healthy larvae are transferred to rearing tanks and the density is maintained around 20-30 nos/l.
The larvae subsist on the yolk for 3 days. From day 4 larvae have to be fed with quality feed. The most
preferred feed for early larval stages, is the rotifer (Brachionus plicatlis or Brachionous rodentiformis ),
the details of live feed to be given for seabass larvae is given below:

29 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

Algae Green unicellular algae like Chlorella sp Tetraselmis sp Nannochlorpsis or
Isochrysis sp are needed for feeding the live feed (zooplankton), Rotifer and for
adding to seabass larval rearing tanks for water quality maintenance.

Rotifer Rotifer (Brachionus plicatilis) or B.rotundiformis is the most prefereed diet for
the fish larvae in their early stages. The size of the Rotifers vary from 50 – 250
µm. The early stage larvae (upto 7 days) are fed with small sized rotifer i.e. less
than 120µm and later assorted size rotifer can be fed.

Artemia Brine shrimp, Artemia in nauplii stage are required for feeding the larvae from 9th
day. Artemia with its natural nutrient profile required for larval development of
fish is used in all the hatcheries. Artemia cyst are kept for hatching and the
freshly hatched nauplii are given as feed for fish larvae upto 21 days and
afterwards Artemia biomass can be given.

During the process of rearing the most important aspect to be looked in to the grading seabass by
nature is showing differential growth and hierarchy is maintained. The larger ones (shooters) always
choose the small and pray upon because of the cannibalistic behaviour. Few large ones will finish of many
smaller ones. Hence from day 15 th onwards as frequent as possible ‘grading’ should be done so that
uniform size fry are reared together. Water exchange is to be done to the extent of 30-40%, the bottom
debris should be removed periodically, appropriate water quality should be maintained in the larval rearing
tank, with technology available with CIBA a survival rate up to 47% could be achieved with average
survival rate around 10% during larval rearing phase. A 25 days old fry will be around 1.2 cm in size.

Nursery Rearing

Nursery rearing is the intermediate phase between hatchery and grow out system. Sea bass
(Lates Calcarifer) can be cultured in ponds or net cages. Before stocking in growout culture system
seabass larvae reared in the hatcheries have to be further reared for a period of 30 – 45 days till they
attain a size that can withstand changes in the culture systems. In the nurseries, the fry can be
stocked in higher densities and reared. This would save space and time in growout phase. The
stockable size of the seed desirable is 5 – 10 gm. Nursery can be done in hatchery using FRP or
cement tanks, fixing hapas in ponds and stocking in small nursery ponds. The nursery rearing phase
will be 30-45 days, during the phase of nursery rearing in tanks and hapas grading of larvae should be
done to avoid cannibalism and better survival rate.

Training Manual on Seed Production and Farming of Brackishwater Finfishes I 30

 Nursery rearing and grow out culture of Asian seabass Lates calcarifer

M.Kailasam, Krishna Sukumaran, G.Biswas, Premkumar, K.Ambasankar and R.Subburaj

Seabass nursery rearing can be done in three different systems such as tank based, hapa net
based and pond based systems depending upon the resource available

Nursery Rearing in tanks

Seabass fry of 25 – 30 days old in the size of 1.0 – 1.5 cm can be stocked in the
nursery tanks of 5 – 10 tonn capacity circular or rectangular (RCC or FRP) tanks. Outdoor
tanks are preferable. The tanks should have water inlet and outlet provision. Flow through
provision is desirable. In situ biological filter outside the rearing tanks would help in the
maintenance of water quality. The water level in the rearing tanks should be 70 – 80 cms.
Good aeration facility should be provided in the nursery tanks. Nursery tanks are prepared a
week before stocking. After filling with water 30 – 40 cm and fertilized with ammonium
sulphate, urea and superphosphate @ 50, 5 and 5 gm (10 : 1 : 1 ratio) per 10 tonne of water
respectively. The natural algal growth would appear within 2-4 days. In these tanks freshly
hatched Artemia nauplli @ 500 – 1000 l are stocked after leveling the water to 70 – 80 cm.
The nauplii stocked are allowed to grow into biomass feeding with rice bran. When
sufficient Artemia biomass is seen, seabass fry are stocked @ 800 – 1000 nos/m3. The pre-
adult Artemia would form good food for seabass fry. Along with ‘Artemia biomass’
available as feed inside the tank, supplementary feed mainly minced fish/shrimp meat is
passed through a mesh net to make each particle of size of around 3 – 5 mm and cladocerans
like Moina sp can also be given. The fish/shrimp meat feeding has to be done daily 3 – 4
times @ 10-20% daily.

Regular water change to an extent of 70% is to be done daily. The left over feed and the
metabolites have to be removed daily and aeration should be provided. In a rearing period of
6-8 weeks in the nursery rearing, the seed will be in the size of 5-10/ 6-8 cm with survival
rate of 65-75%. For better survival, regular `Grading’ should be done. Vessels/trough placed
with different mesh sized nets can be used for grading. When the seed are left into the
containers the seeds will be sieved in different grades according to the mesh size and seed
size. Care should be taken that the fry are not injured while handling. If the number is less it
could be manually done.

31 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

Nursery Rearing in Ponds

Nursery ponds can be around 200-250 m2 area with provision to retain at least 70 –
80 cm water level. Adequate provision for water inlet and water drainage should be
provided. Towards drainage side there should be slope. Suitable sized (normally 1 mm)
mesh screen nets should be provided in the inlet side and outlet side to avoid entry of
unwanted fishes and escape of the stocked fish respectively.

The pond is prepared before stocking. If there are any predator/pest fishes they have
to be removed. Repeated netting, draining and drying the pond are done. In case where
complete draining is not possible, water level is reduced to the extent possible and treated
with Derris root powder @ 20 kg/ha added or mahua oil cake @ 200-300 kg to eradicate
unwanted fishes. After checking the pond bottom quality water is filled. If the pond bottom
is acidic, neutralization is done with lime application. In order to make the natural food
abundant, the pond is fertilized with cow dung manure @ 500 kg/ha keeping the pond water
level 40-50 cm. The water level is gradually increased. After 2-3 weeks period when the
natural algal food is more, freshly hatched Artemia nauplii are introduced. Normally 1 kg of
cyst is used for 1 ha pond. These stocked nauplii grow and become biomass in the pond
forming food for the seabass fry. Seabass fry is stocked @ 20-30 Nos/m2. Stocking should
be done in the early hours of the day. Fry should be acclimatized to the pond condition.
Acclimatization for the pond condition is done as follows:

Supplementary feeding is done with chopped, cooked fish/shrimp meat. The larvae
can be weaned to artificial feed at this stage. The feeding rate can be as mentioned in earlier.
Excessive feeding should be avoided since excess feeding would deteriorate the pond
condition and also promote filamentous algal growth. The excessive algal growth would
deplete dissolved oxygen level in the early hours of the day leading to fish mortality. Hence,
excessive algae if any should be removed. In a rearing period of 4-6 weeks in the nursery
rearing, the seed will be in the size of 4-12/ 5-10 cm with survival rate of 35-45%.

Nursery Rearing in Cages/Hapas

Rearing seabass fry in cages/hapas is commonly practiced. This method is

advantageous than other methods since the management is easier and installation of rearing
facility requires less space and capital investment. It can be also extended to any scale
depending on the necessity and the capability of the farmer. It can be maintained in one

Training Manual on Seed Production and Farming of Brackishwater Finfishes I 32

corner of the growout pond or near the growout cages itself. Since cages or hapas are in in
situ condition this will provide conducive environmental condition. The water flow in the
cage site would give the fish natural condition. The metabolites and the excess uneaten feed
will be washed away by the flow of water.

Floating net cages/hapas can be in the size of 2 x 1x 1 to 2x 2x 1 m depending upon

necessity. Cages are made with nylon/polyethylene webbing with mesh size of 1 mm. Fry
can be stocked @ 400 – 500/m2. Feeding rate can be as that described to tank nursery. The
net cages have to be checked daily for damages those may be caused by other animals like
crabs. The net cages will be clogged by the adherence of suspended and detritus materials
and siltation or due to foulers resulting in the restriction of water flow. This would create
confinement in the cages and unhealthy conditions. To avoid this, cages/hapas should be
cleaned everyday. Regular grading should be done to avoid cannibalism and increase the
survival rate. Even in higher stocking density @ 500/m2 farmer could get survival of 80% in
the farm site when the fry were reared in hapas adopting the trash fish feeding and other
management strategies mentioned above.

Seabass nursery rearing can be done by the self help group. For this purpose, each hapa
(2m2) can be given to two members to take care. Stocking of seabass fry can be done @
1000 nos/ha The farm made seabass nursery feed made in dough form and mixed with
cooked fish meat (Tilapia, sardines mackerel etc.,) and can be kept in the feed tray for
feeding twice a day @ 10-20% body weight. Seabass fingerling can be totally weaned to the
artificial feed with gradual with drawl of mixing minced fish meet meat with the farm made
feed. Grading done mostly by 3-4 people taking help from other groups every 4 days interval
to segregate the same size fishes and to avoid the cannibalism. During this time, hapa also
removed and checked for the holes/damage if any and tied back after cleaning using brushes.
As the fish grow to bigger (above 3 cm), aeration provided in all the segregation tubs using
battery aerators during grading and frequent water exchange also done. Feed supplied to the
fish two times morning (before 8.00 AM) and evening around 5.00 PM (When temperature is
low fish comes to surface and consume feed comfortably) Once, seabass reached to above 5
cm, they can be sold. Seabass fingerlings in the size group between 5-8 cm sold in two and
half months culture period (seed will be ready for sales once reach 5 cm size especially after
45 days and keep going up to 75 days). Survival rate can be obtained from 65 to 75%. The
seed sold @ Rs.2/cm. The production cost will be approximately around 50-60 paise per cm.

33 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

Pre-grow out culture technology

Pre-grow out culture is the stage between nursery and grow out culture, in which the
5-10g size fingerling seabas is grown to 80-100g stockable size for grow out culture for the
period from 60-75 days culture under pond condition wih the expected survival rate from 45-
60%. In this stage, seabass fingerlings (5-10g) can be stocked @ 10000-15000 nos/ha. Small
size ponds in the size of 0.1 to 0.2 ha are highly preferable for easy operation. Feeding can be
done @ 8-15% body weight daily basis in two rations per day. Total fish biomass can be
measured fortnightly basis by taking the random sampling of fish weight and accordingly
feed quantity can be decided. Care must be taken to maintain the optimal feed quantity while
feeding daily since under feeding may cause differential growth which may leads to
cannibalism and reduction in the survival rate.

Grow out culture technology

Traditional Culture

Seabass is cultured in the ponds traditionally as an extensive type culture. In low

lying excavated ponds, whenever the seabass juveniles are available from the wild (For eg.
April June in West Bengal, May-August in Andhra Pradesh, Sept-Nov. in Tamil Nadu), May
to July in Kerala and June-July in Maharashtra) they are collected and stocked in the pond
along with some species of fish, shrimps etc., The juvenile seabass introduced in the pond
will prey upon the available fish or shrimp juveniles as much as available and grow. Seabass
are allowed to grow for 6-7 months of culture period till such time water level is available in
these ponds and then harvested. In this manner, production upto 2 ton/ha/7-8 months have
been obtained. However, this practice is highly unorganized and without any guarantee on
production or return for the aquaculturists. With the advances in the technology in the
production of seed under captivity assuring the supply of uniform sized seed for stocking and
quality feed for feeding, the seabass culture can be done in more organized manner.

Improved Seabass Culture Methods

In this method, stocking of uniform sized seed at specific density and fed with low
cost trash fishes/formulated feed of required quantity. Water quality is maintained with
exchange periodically. Fishes are allowed to grow to marketable size,harvested and marketed
for high unit price. Seabass culture can be done in more organized manner as a small-

Training Manual on Seed Production and Farming of Brackishwater Finfishes I 34

scale/large scale aquaculture in brackishwater and freshwater ponds in cages. Seabass is
cultured in the ponds adopting poly culture method or feeding with low cost fishes like
Tilapia/oil sardines or with extruded floating pellets.

Pond preparation

The pond proposed for seabass seed stocking is dried, tilled and levelled, manured
with raw cow dung @10,000 kg/ha. Fertilizers like Urea @100kg/ha, Super phosphate
@50kg/ha can also be added to enhance the algal bloom in the pond. Water from the
adjoining water sources

seawater /freshwater is filled to a depth of 60-70cms in the pond. After sufficient algal
bloom (light green in colour) is observed, in the pond forage fishes like Tilpia are introduced.

Forage fish/Feed fish stocking and introduction of seabass

Fishes like Tilapia can be introduced @ 20-25 thousand per ha. into the pond.
Stocking of forage fishes should be done in the seabass culture pond a month ahead of
seabass stocking. The water level in the pond may be maintained around 80-90 cm.
Subsistence level of feeding with cheaper ingredients like Rice bran and ground nut oil cake
@ 2-3% of the biomass can be done. Being a prolific breeder in habit, the forage fish Tilapia
will start breeding in the pond and at given point of time in the proposed seabass culture
pond, there will be different size groups of Tilapia will be available. In the pond with natural
forage feed, seabass of 80-100 gms can be stocked @ 3000 - 4000/ha depending upon the
management capacity of the farmers like water change and feeding at later stages. Seabass
should be uniform in size to avoid differential growth in the culture pond. The seabass
stocked will feed upon the hatchlings fry and fingerlings of the fry initially and later on the
adult fishes also. Since, seabass has got higher growth potential than the forage fishes they
will dominate in the pond. After 3-4 months there will not be any forage fish in the pond. At
that stage, fresh stock of feed fishes should be introduced.

Production and Economics of Polyculture

Hatchery produced seabass seed supplied to farmers was followed for nursery and grow-out
practice. In Ganapavaram (A.P), seabass farmed with Tilapia stocking density @
10000nos/ha under polyculture with forage tilapia with an initial size of 70g. In 320 days of
culture a total production of 3740 kg was achieved with a productivity of 3120kg/ha under

35 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

polyculture system. Seabass production was highest with 1634kg, followed by Rohu 1458kg,
Tilapia 576 kg and Catla 70 kg. Seabass average body weight (ABW) was- 1.7 kg and the
maximum of 2.7 kg with the minimum of 0.750 kg was recorded in the harvested biomass.
20% of the seabass harvested biomass was more than 2 kg, 52% of the harvested biomass
was more than 1.5 kg and 28 % was less than 1 kg. Rohu, Catla and Tilapia ABW were
0.350kg, 3.5kg and 0.200kg respectively. Seabass survival of 90.25% was achieved. Gross
profit of Rs.234694/- was achieved.

Advantage and disadvantage of Polyculture method

Advantages

1. Polyculture method is more natural

2. Fish can take feed depending upon the requirement

3. Since, the live fish only serve as feed, the water quality maintenance is easier. No

contamination due to left over feed.

4. Cost effective.

Disadvantages

1. Not feasible at all places

2. The forage fishes like Tilapia itself are food fishes in some places and costlier.

Grow out Culture of Seabass in Cages

Fish culture in cages has been identified as one of the eco-friendly at the same time
intensive culture practice for increasing in fish production. Cages can be installed in open sea
or in coastal area. Cage culture system allows high stocking density, assures high
survival rate. . Feeding can be controlled and cages can be easily managed. Harvesting is not
expensive. Water depth and water current alone the criteria. Even in areas, where the
topography of the bottom is unsuitable for pond construction, cage can be installed. Diseases
can be easily monitored. Fishes in the cages can be harvested as per the requirement of the
consumers, which will fetch high unit price. Cages can be relocated whenever necessary to
avoid any unfavorable condition.

Training Manual on Seed Production and Farming of Brackishwater Finfishes I 36

Design of Cages

Grow out cages of 10 or 50 M2 are preferable for easy management and maintenance.
Cages are fabricated with polyethylene netting with mesh size ranging from 2 to 8 cm
depending upon juvenile fish propose to stocked in the cages. There are two types of cages

Floating net Cages

The net cages are attached to wooden frames kept afloat using plastic drums.
Anchors or Concrete weight blocks as anchors can be attached to the corners of the net cage
at the bottom. These types of cages can be installed in areas with water depth more than 4
meters with feeble water current.

Stationary net Cages

These are fixed enclosures, which can be installed, in shallow water areas in lagoons,
brackishwater lakes having water depth of 2-4 meters. The cage net is fastened to wooden
poles erected in the water system at the four corners.

Stocking Density

In the cages, fishes can be stocked @25-30nos/m2 initially when they are in the size
of 10-15 gm. As they grow, after 2-3 months culture, when they are around 100-150 gms
stocking density has to be reduced to 10-12 nos/m2 for space. Cage culture is normally done
in two phase – till they attain 100-150gms size in 2-3 months and afterwards till they attain
600-800 in 5 months.

Feeding in Cages

Fishes in the cage can be fed with either extruded pellets or with low cost fishes as per
the availability and cost. Floating pellets have advantages of procurement, storage and
feeding. Since, a lot of low cost fishes are landed in the commercial landings in the coastal
areas which are fetching around Rs.3-5/kg only used as feed for seabass culture. Low cost
fishes like Tilapia available in the freshwater and brackishwater also serve as feed for
seabass in ponds and in many cage culture operations. The rate of feeding can be maintained
around 20% initially and reduced 10% and 5% gradually in the case of trash fish feeding and
in the pellet feeding, the feeding rate can be around 5% initially and gradually reduced to 2-
3% at later stage. In the feeding

37 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

of low cost fish FCR works out around 6 or 7 (i.e. 7 kgs of cheaper fishes has to be given for
one kg of seabass). In the case pelleted feeding FCR is claimed to be around 1 to 1.2.
However, the cost effectiveness of the pellet feeding for seabass in grow out culture has to be
tested.

Cage Management

Since cages are inside the water and exposed to water current, the debris materials
drifted may adhere to the cages and clog the mesh restricting the water exchange. The
fouling organism will also attach and clog the meshes. Other animals like Crab may damage
the nets. The cages should be regularly checked for clogs and leaks. Damaged nets should
be repaired or replaced. The clogging will reduce water exchange, and lead to accumulation
of waste products depleting the oxygen causing stress to the fishes, affecting feeding and
growth. If the damage is not repaired immediately, the fishes will escape from the cages.

Production

Under cage culture, since seabass can be intensively stocked and properly managed,
the production will be high. Frequently culling and maintenance of uniform sized fishes in to
the cages will ensure uniform growth and high production. Production of 6-8 kg/m2 is
possible in the cages, under normal maintenance and production as high as 20-25 kg/m2 is
obtained in intensive cage management in the culture of seabass.

Training Manual on Seed Production and Farming of Brackishwater Finfishes I 38

 Broodstock development, Induce breeding and larval rearing of milkfish
(Chanos chanos)
Aritra Bera, M.Kailasam , Babita Mandal, Tanveer Hussain, Krishna Sukumaran,
M.Makesh, K.Ambasankar

Introduction

Milkfish (Chanos chanos) is traditionally cultured in Philippines. Indonesia also

through its small scale hatcheries is a major producer of milk fish seeds. Milkfish culture is
also practiced in some Pacific Islands viz. Kiribati, Nauru, Palau and the Cook Islands as well
as in countries like Taiwan. There is increasing production from intensive mariculture cages
now a days although most milkfish culture is undertaken in brackishwater ponds, low saline
areas with extensive and semi-intensive way. The milkfish, C. chanos, does not reach
gonadal maturity easily in captivity, although it achieves maturity under marine open sea
cages. Since 80’s artificial induction of milk fish spawning has been achieved in countries
like Philippines, Taiwan and Hawaii and other small Asia-pacific islands in marine earthen /
cemented ponds and sea cages.

Sources of broodstock and Holding facilities

Milkfish juveniles (weight, 250-350 g) can be obtained from brackishwater ponds and fish
pens. Milkfish juveniles are transported to the broodstock cages or tanks in a floating fish
cage or in a 1.5 m dia. Land-based concrete tanks measuring 12 m x 6 m x 2 m deep are
sufficient for milkfish broodstock. A double pipe drainage is installed to allow water to flow
out from the bottom. Water inlets and aeration lines are located at the top. Daily water inflow
should be adjusted to change at least 50% of the water volume. The sides and bottom of the
tank should be brushed monthly. The tank can be drained to at least a foot in depth for
brushing. During the spawning season, water inflow should be increased so that brushing and
draining can be minimized. In both tanks and cages, optimum stocking density should not be
more than 1 kg/m3.

39 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

 Fig : Milkfish Broodstock Tank (144t capacity)

Brood Nutrition

Two to four year old milkfish can be fed twice daily with commercial fish pellets (24%
protein) at 3% of their total body weight. Upon nearing maturation by the fourth year, fish are
fed twice daily with pellet feed with 36-42 % protein and 6-8 % lipid. Daily feeding ration is
increased to 4% of total body weight.

Maturation, Induced breeding and larval rearing

Major constrains to achieve mass artificial propagation age generally are limited
resource of mature broodstock, proper environmental manipulation and stress from frequent
handling, brood nutrition management and administration of required dose of synthetic
hormones are few. Natural maturation and spawning have taken place in floating sea cages
and more notably, in earthen marine ponds in recent past. In a path breaking experimental
trial Lee et al. (1986) tested five chronic hormone therapies and found that a combination of
LHRH-a cholesterol pellets and 17α- Methyl Testosterone (17α-MT) capsules effectively
stimulated maturation of milkfish and induced them to spawn under captivity. The lack of
commercial scale availability of hatchery produced seed is the major bottleneck for any large
scale venture of marine finfish farming. The availability of seeds from wild is often
unpredictable, risky as they comes with predator fish, un-uniform size group and hence
farming based on wild collected seeds may not be a sustainable venture. Hence the
development and standardization of seed production techniques milkfish should receive
research priority. (Lee et al., 1986; Tamau, 1988).

Training Manual on Seed Production and Farming of Brackishwater Finfishes I 40

Sexual Dimorphism

Milkfish being a bisexual fish mature males have, 2 openings in the anal region which
are externally visible in, and 3 in mature females. Female Milkfish attain sexual maturity at
around 5 years of age whereas males maturity earlier at around 4 years of age.

Source: The Proceedings of the Indian Academy of Science (Tampi, 1957)

Maturity Determination

Fishes fail to achieve their normal reproductive cycle under captivity. Culture
conditions much time do not provide an environment conducive to completing maturation of
the gonads and ultimately spawning. In some cases, changing the environment like
maintaining of salinity, temperature, water quality parameters has proven sufficient alteration
for fish to resume their normal reproductive activities. Milkfish do not exhibit any sexual
dimorphism and thus adult milkfish of 6-8 years old should randomly stock into cement/FRP
tanks with an assumed sex ratio of 2:1 (M:F). Sometime salinity affects ovulation and may
exert some influence on gonadal development when it is extremely high. Temperature (240 C
is minimum) and photoperiod (11L: 13D; 14L: 10D) is very important parameters but less
studied. It has been seen that gonadal maturation is synchronised with temperature rising
from 25 – 32ο C and 11-14 hour light.

Problems and solutions related to milkfish maturation

 Stress due to frequent handling seemed to be negative for the stimulatory effect of
hormones and initiates resorption of gonads in both sexes.

41 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

  Pellet implantation and hormone medicated feeding exerts lesser stress. But Protein
(Gonadotrophins : LHRH, GnRH, FSH) based hormones cannot be fed orally but sex-
steroids can be administered in that way.
 It was seen that spermatogenesis can be obtained in 4-year old milkfish through
feeding of 17 alpha-methyl testosterone.
 Milkfish <4 or 5 years old may not have developed receptors to respond to hormone
treatments.
 In wild collection ripe males are less in numbers. If not hormone induced, males
reabsorb milt after 2-3 days. To induce spermiation HCG + androgens can also be
used.
 It has been seen, milkfish sperms can be stored for 10 days in 12.5 % DMSO at near
zero temperature. As male fishes mature early than female cryopreservation of milt
may be of great help. (Lam, 1984)

Brood Nutrition

A diet formulated by SEAFDEC/AQD containing 36% protein & 6% lipid and

vitamin mixture to support sexual maturation and production of high quality eggs of milkfish
is used to feed the broodstock. . With adult brine shrimp A. salina feeding milkfish showed
reproductive readiness throughout the year in Isles Lagoon. (Lam, 1984)

Induction of Gonadal Maturation and Induced Breeding

Induced maturation of milkfish commonly starts at oocyte dia of 0.66 - 0.82 mm at 35

ppt salinity. Sex ratio of 2:1 (M:F) is better to maintain. Fertilized eggs are represented by
1.1-1.23 mm oocyte dia. Fish handling stress can be minimized by 2- phenoxy ethanol or
diazepam as tranquilizers during handling. The hormones used can be Salmon or Carp
pituitary homogenate in combination with HCG, or HCG alone. Hormone dosage can be
variable: (Lam, 1984)

(a) SPH 6-10 mg/kg

(b) CPH 5-25 mg/kg
(c) HCG 180-2500 IU/kg

Numbers of hormone injections are generally 1-5 (mostly 2) with injection interval of
6-24 h (mostly 8-12 h). Time to stripping is commonly 6-17 h (12 h appears best) in case of
Training Manual on Seed Production and Farming of Brackishwater Finfishes I 42
oozing female and male. There are certain behavioral markers which may help to determine
both whether the injection given is effective and also the time of stripping. These include the
following (Lam, 1984):

 Color change, due to melanophore stimulating hormone (MSH) in the pituitary

homogenate
 Increased drinking activity, probably to facilitate oocyte hydration
 Release of calcium deposits, probably with increased drinking; calcium is retained in
the gut and then released
 Distension of abdomen, indicating oocyte hydration
 Dribbling of some eggs, indicating that ovulation may be close, consequently
 a good reference point to determine the time of stripping.
 Milkfish less than 4 or 5 years old may not have developed the receptors to respond to
hormone treatments.

Similarly, spent fish may lack hormone receptors. It is not known how long it takes
spent fish to undergo recrudescence (rematuration) or whether they remature at all in
captivity. It is also not certain whether milkfish are total or intermittent spawners. ICAR-
CIBA has achieved induced breeding of Milkfish first time in India during June, 2015. Brood
stocks of milkfishes were maintained in 100 t capacity cement tank for more than 8-10 years
at Muttukadu Experimental Station of CIBA. After several number of LHRH-a hormone
pellet implantations final gonadal maturity and first successful spawning of milkfish were
happened. Fertilized egg diameter was 1.23 mm and length of newly hatched larvae was 3.4
mm. Successful maintenance of brood stock is one of the key factors in hatchery production
of milkfish seed.

Spawning

Natural spawning using pond-reared milkfish was achieved in Taiwan. The first
instance was in August 1980, when the milkfish broodstock in floating cages at
SEAFDEC/AQD spawned spontaneously. When mature, the ovary is usually around 10% of
body weight, but could be nearly 25% .A 5-13 kg female can produce 300000 eggs/kg body
weight. In wild, milkfish spawns more than once a year. Spontaneous spawning without
hormone treatment has also been achieved with captive broodstock maintained in floating
net-cages in the Philippines.

43 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

Larval Rearing

Feeding should initiate after 3rd day post hatching (dph). Critical period is between the
4th and 6th days. Chlorella salina added @ 3 – 4 X 105 cells/ml as feed for rotifer and to
condition the water. Application of mix algae like Isochrysis galbana and Tetraselmis chuii
shows significant improvement in larval survival. Larvae at day 0, 14 and 21 day are highly
euryhaline (0-70 ppt), those at day 7 are markedly stenohaline (27-28 ppt). This suggests that
rearing milkfish larvae at a constant salinity of 27-28 ppt may improve their survival rate.
(Lam, 1984). The hatchlings are stocked in the indoor larval rearing tanks (LRT) @ 30 no/l
and first feeding initiated with rotifer Brachionus plicatilis from 2 day post hatch (dph).
Rotifer density of 15 – 20 no/ml is ideal till 15 dph. Artemia nauplii was introduced along
with the rotifer on 15 day post hatch (dph) @ 0.5 – 1.0 number/ml. The milkfish larvae
reached to the early fry stage ( 20 mm) on 21 dph and accepts artificial feed .

Live Transport

Live transport of milkfish combines utilizing pre-transport starvation, anesthetic (2-

phenoxy ethanol) at capture, chilled water during transport of brood stock.,. Live transport
can be carried out in 4-13 year old broodstock placed in oxygenated transport plastic bags
(2m long x 0.5 m wide) containing 40-L chilled (20-22°C) seawater and 5ml 2-
phenoxyethanol (anesthetic). The transport bags were then placed in styrofoam/thermocol
boxes. Travel time should not exceed 6-7 hours to have fast recovery and no mortality. Eggs
(embryonic stage) and newly-hatched larvae were also transported in oxygenated plastic bags
containing 12-l seawater and supported by straw bags. Optimum density, temperature, and
salinity should maintain are: 100,000- 120,000 eggs or larvae per bag, 28-30°C and 32-34 ppt
of salinity.

Conclusion

Milkfish being herbivorous euryhaline species is slowly getting position as an

important cultivable species tin brackish water sector in India. As nursery rearing, pre grow
out and grow out culture of milkfish is easy , less risky, less capital intensive and good
economical return, it can lead to a sustainable culture option in vast underutilized saline areas
apart from shrimp farming areas in India. Successful induced breeding and commercial level
seed production of milkfish can open a new horizon to include more species for
brackishwater aquaculture. Milkfish can be cultured at coastal waters, estuaries and
brackishwater water bodies such as Chlika lake in Odisha, Pulicat lake in Andhra Pradesh,

Training Manual on Seed Production and Farming of Brackishwater Finfishes I 44

Bheries in West Bengal, backwater in Kerala, Goa and Karnataka. Milkfish having similar
look and spiny nature as Hilsa can have a ready market with a selling price of Rs.150-200/Kg
in West Bengal, Odisha and other North Eastern states of India and can be recognized as
Decan Hilsa in domestic market. Milkfish can be used as live bait for tuna industry also.

45 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

 Present status of milkfish Chanos chanos farming
Kailasm M, , Pankaj Patil , Aritra Bera, Tanveer Hussain, Babita Mandal

Introduction

The milkfish (Chanos chanos) is one of the most ideal finfishes for farming in coastal
areas. They are fast growing, tolerates a wide range of temperature, oxygen and salinity.
Milkfish are cultured in large scales in countries like Indonesia, Philippines and Taiwan in
ponds called “Tambak”. Milkfish farming in Indonesia, Taiwan Province of China and the
Philippines started about 4-6 centuries ago. In India too the popularity of its farming is
growing especially in Tamil Nadu and Kerala. The fish is either monoculture or polyculture
with compatible species of fish & shrimps.

Nursery operations in milkfish producing countries vary according to established

cultural practices. In Taiwan Province of China, where commercial hatchery and nursery
productions are integrated enterprises, milkfish fry are generally grown in either earthen
ponds or elevated canvas or concrete tanks at intensive stocking densities of >2 000/litre. In
Indonesia, a well-established backyard-type nursery is used. This consists of a series of
elevated canvas or concrete 1-2 tonnes tanks and similar stocking densities to those used in
Taiwan Province of China are employed. In the Philippines, milkfish nurseries are integrated
with grow-out facilities, where wild-caught or hatchery-reared fry are first acclimated into
nursery compartments which comprise one third to one quarter of the total area of the
brackish water pond. Fry are stocked at a density of up to 1 000/litre and are fed with a
naturally-grown micro-benthic food known as 'lab-lab' which grows on the fertilized pond
bottom. When natural food is becoming depleted, artificial feeds such as rice bran, corn bran,
and stale bread or formulated feeds are provided.

During the past decade, much progress has been made, particularly in regard to
milkfish propagation and the mass production of fry by private hatcheries, research
institutions and government agencies. Instead of relying on wild-caught fry, milkfish farms in
the Philippines, Taiwan Province of China and Indonesia now obtain the majority of their fry
from hatcheries, mainly due to the significant shortage of wild-caught fry.

Shallow water culture is practiced mainly in Indonesia and the Philippines. Milkfish
are traditionally cultured in shallow Brackish water ponds in which the growth of benthic

Training Manual on Seed Production and Farming of Brackishwater Finfishes I 46

algae is encouraged through inorganic or organic fertilization. Milkfish will survive on
benthic algae alone only if the productivity of the algae exceeds the grazing rate of the fish;
otherwise, supplemental commercial feeds are applied. The 'lab-lab' culture system in the
Philippines is equivalent to shallow water culture in Taiwan Province of China. 'Lab-lab' is
the term used in this country for the algal mat (and all micro-organisms associated with it) in
the ongrowing ponds. Brackish water ponds in the Philippines were mostly excavated from
'nipa' and mangrove areas. Shallow water pond design generally consists of several nursery
and production ponds with a typical area of 2 000 m² for nursery ponds and 4 ha for
production (ongrowing) ponds. Typically, ponds have a depth of 30-40 cm and are provided
with independent water supplies. The average yield of a typical integrated nursery, transition
and shallow grow-out system that produces 3 crops a year is 800 kg/ha. Modified modular
pond designs consisting of a series of grow-out compartments with a maximum of eight crops
a year have been shown to increase yield to a high as 2 000 kg/ha.

Deep water culture was developed in the mid 1970s in response to the decline of
profitability of shallow water culture, and the limited and increasing value of land and
manpower resources. Deep-water ponds provide a more stable environment and extend the
grow-out period into the winter season. Most deep-water milkfish ponds have been created by
converting either shallow water ponds or freshwater ponds, with a depth of 2-3 m. Production
from these systems has sharply increased in Taiwan Province of China.

Most milkfish ponds in the Philippines and Indonesia are of the extensive and semi-
intensive type, with large shallow pond units, tidal water exchange, natural food, minimal use
of fertilizer alternating with commercial feeds and other inputs, and low to medium stocking
rates (50 000-100 000/ha). The Taiwanese method of production, on the other hand, employs
intensive stocking densities (150 000-200 000/ha).

Indian scenario

Milkfish (Chanos chanos), is naturally present in Indian and Pacific Ocean. It is a

national fish of Philippines where it is known as ‘Bangus’. Milkfish is a tasty fish. It can
tolerate wide range of salinity. Culture of milkfish in brackish water ponds and pens is an
age-old and traditional practice in many tropical countries such as Philippines, Taiwan and
Indonesia and Pacific island countries. It is also farmed in freshwater ponds, lakes, reservoirs
and in marine cages in these countries. Global production is about 9 lakh metric ton during
2012. In 2014 Philippines, the leading producer of milkfish has produced 391,983 MT. In

47 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

India, occurrence and traditional culture of milkfish has been reported in coastal waters,
estuaries and brackishwater water bodies such as Chilika lake in Odisha, Pulicat lake in
Andhra Pradesh etc since long back. It was a favorite fish of Mysore king Tipu Sultan in the
19th century and he used to procure it from brackishwater ponds from Kundapur in
Karnataka. It is called as Paal Meen in Tamil, Pala Bontha and Tulli Chepa in Telugu,
Poomeen in Malayalam, Hoomeenu in Kannada, Golsi in Goa and Seba khainga in Oriya. Its
important characters such as herbivorous feeding habit, low disease occurrence, rapid growth,
wide range of salinity tolerance, attractive appearance, longer shelf life in ice, good taste and
texture and suitability as live bait in tuna industry, makes it a potential candidate for culture
to revolutionize aquaculture growth.

Status of seed sources

Worldwide, Milkfish seeds were collected from coastal areas since beginning for
brackishwater farming. This has led to decline in fry availability in nature presently. In
1970’s Philippines developed hatchery technology for seed production of milkfish which has
given an impetus for the milkfish farming. In India, wild caught seeds are collected in the
months of March to June and September to December from coastal states of Tamil Nadu,
Andhra Pradesh and Kerala etc. by traditional methods. Fry are more abundant during the
new and full moon periods. In India, increasing trend of the milkfish farming still depends on
the availability of seeds from wild resources which lacks good quality and sometime comes
along with predatory fishes. Therefore, technology development of breeding, seed production
and culture practices in brackishwater area is important for the development of milkfish
production in India.
Pen Culture

Pen culture of milkfish was first introduced in the Philippines in 1979. Fish pen size
ranges from 1 ha up to 50 ha. In this system, the milkfish feed mainly on plankton and also
forage for food at the bottom. However, there are times that supplementary feeding may be
required especially when stocked at higher densities or natural food becomes depleted.
Stocking density is about 30,000 to 50,000 fingerlings per hectare which equals 1 fish per
m3. The fish grow to market size (250-300 g) in 4 to 8 months with survival of 60-80% and
yield from 4,000 kg/ha to as high as 10,000 kg/ha. The fish reach harvest size of 250-275 g in
4-5 months with a survival rate of 80-90% and production of 1.5-5 kg/m2.

Training Manual on Seed Production and Farming of Brackishwater Finfishes I 48

Milkfish Culture in Cages

Milkfish cages may be installed in freshwater lakes, estuarine areas, and coastal
marine waters. Cages may be square or rectangular using bamboo frames or G.I. frames with
drum floats. More advanced design consists of high-impact polypropylene pipe frame which
serve also as float. Feeding of complete formulated diet (27-31% protein) is essential from
stocking of the fish to harvest. Small-sized fingerlings (5-10 grams) are initially stocked at
higher density in cages with nets having small mesh size for 1-2 months before being
transferred at desired density to grow-out cages. Stocking density depends on the carrying
capacity of the cage and the environment. Typical stocking densities in floating and
stationary cages are 10-40 pcs/m3 with a survival rate ranging from 70-90% and yield of 3-20
kg/m3. Offshore cages can be stocked with 40-100 pcs/m3 with a yield from 20-35 kg/m3
.Sizes at final harvest typically range from 350 to 500 grams. .

Polyculture

Milkfish is reared with shrimps, mud crab, rabbitfish, seabass, tilapia, seaweeds,
mollusks, and many other fish species either as primary or secondary crop. The polyculture of
milkfish with shrimps or with crabs however, are the most popular and profitable. They
compliment each other in terms of habitat and food requirements. Annual yield of milkfish as
the primary stock when grown together with shrimp ranges from 1,200 to 1,800 kg/ha while
annual shrimps production is from 100 to 200 kg/ha. On the other hand, about 550 kg per ha
of milkfish and 1,500 kg/ha of crabs per crop can be attained using the polyculture method.
Generally a minimum of 2 crops per year can be undertaken when milkfish is polycultured
with either shrimps or crabs.

Extensive traditional farming

In India, traditional milkfish farming can be done in different brackishwater areas like
Bheries in West Bengal, Chilika Lake in Odisha, Pokkali in Kerala and Ghazni in Karnataka
& Goa. These traditional ponds are having water depth of 40-60 cm which is suitable to stock
milkfish fingerlings of 7–10 cm body size with stocking density of 1000-1500
fingerling/acre/crop depending on the cropping pattern. 2 crops/year can be harvested in
batch stocking cropping pattern. After every 15 days, partial harvesting can be done by using
gill net in continuous stocking cropping pattern. Every partial harvest is followed by re-
stocking with milkfish fingerlings. In extensive traditional farming, milkfish fingerlings feed
on only natural food like Lab-lab (benthic algae) and Lumut (filamentous algae). No artificial

49 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

feed is provided. Final harvest of 1.5 to 2.5 tons/hectare/year is achieved with lablab feeding
whereas lumut feeding yields only 500- 600 kg/ha/year.
Intensive culture

The intensive milkfish culture requires smaller (0.1-1 hectare) but deeper (1-2 m)
grow-out pond, enormous capital investments, large working capital, and technical
proficiency. Paddle wheel aerators, feeding devices and pump for water exchange assist to
increase the natural primary productivity of pond. Milkfish fingerling of 7-15 cm body size
with stocking density of 8,000–12,000 fingerlings/ha to highest density of 30,000 fingerlings/
ha can be stocked in ponds. Feeding with floating pellet (CP 24-28%, CF 3-4%) improve
FCR. Daily feed ration should not exceed 1.5% of total biomass in a pond. After 3-4 months
of culture, Milkfish (200-300 g) can be harvested with the help of dragnet or gill net.
Production of 4–6 tons/ha/year to 12–15 tons/ha/year can be achieved after a culture period of
3 - 4 months. Mass mortality is a constant threat due to accumulations of toxic metabolites
such as ammonia and sulfides, oxygen depletion, and diseases. Procedures in pond
preparation, maintaining

Semi-intensive Culture

This method is characterized by smaller pond size of 1 to 5 hectares, at least 1 meter

depth of water, and an increased stocking rate of 8,000 to 12,000 fingerlings per hectare in
the rearing pond. Water exchange is enhanced by widening the gate, provision of separate
drain gate and using water pump. Oxygen supply is improved by providing paddlewheel
aerators and maintaining good phytoplankton growth later in the growing period. Natural
food, mainly lab-lab, is grown and used as food in the first 45 to 60 days of culture in the
grow-out ponds and commercial formulated diet with at least 27% protein is supplied
thereafter. This method allows 2 to 3 crops and yields of up to 7.5 tons per ha per year.

Training Manual on Seed Production and Farming of Brackishwater Finfishes I 50

 Biology of Grey Mullet (Mugil cephalus)
Rekha M.U., Krishna Sukumaran, Dani Thomas, Tanveer hussain

Introduction

Mugil cephalus L. is cosmopolitan and contribute significantly to the economy of

countries of Southeast Asia, Mediteranean region, Taiwan, Japan and Hawaii. This species is
euryhaline and capable of surviving in wide variety of marine, estuarine and freshwater
environments of varying turbidity, salinity and dissolved oxygen levels (Thomson 1955,
Ibanez and Guitierrez-Benitez 2004).

Distribution in India

M. cephalus comes under family Mugilidae, which comprises a total of 20 genera and
70 valid species (11 of which belong to the genus Mugil) (Eschmeyer and Fricke 2011). In
India 13 species of mullets are well recognised. Of these, 8 species contribute to the
commercial catches. They are Mugil cephalus, M. cunnesius, Liza macrolepis, L. parsia, L.
fade, Ellochelon vaigiensis, Valamugil seheli and Rhinomugil covsula. The other known
species are L. carinatus, V. buchanani, Sicamugil cascasia, Plicomugil labiosus and
Crenimugil crenilabis. Mullets are caught along the sea coast, in the lagoons and the
adjoining brackish-water lakes, and in the estuaries. As they are caught almost throughout the
year, they are a valuable source of food-fish during the offseason of the other commercial
fisheries. Since mullets in general are hardy fish they are best suited for fish farming through
which could be obtained better increments in growth and a ready source of fish. Mullets are
usually distinguished by the presence of two separate dorsal fins, small triangular mouths,
and the absence of a lateral line organ. They feed on detritus and most species have unusually
muscular stomachs and a complex pharynx to help in digestion. Mullets are caught in cast
nets, dip nets and seins almost throughout the year and contribute to the fishery in estuaries,
backwaters and sea. (G. Luther, 1973)

However, this species appears to be rare in the Hooghly-Matlah estuarine system. In

the Chilka Lake this is the most common mullet and fishing season extends almost
throughout the year. It is mostly caught by Jano fishing. Fish with roe is common during
September-January forming a peak in October-November. This species undertakes seaward
breeding migration from September/October to December, when sizes between 35 and 53 cm
are common in the catches. In the Mahanadi estuary, the species is available throughout the
year, with the peak fishing season during September-November/December. In the Pulicat
Lake, this is very common in the mullet catch, and is abundant during April, June-July,
December and March with the dominant size between 20 and 43 cm. Fish with roe occurs for
a few months from November onwards. At Mandapam sizes between 7 and 31 cm are
commonly caught from the Palk Bay and the Gulf of Mannar. In the Kayamkulam and the
Vembanad Lakes the species is very common, occurring in the size range of 23-61 cm. and
with roe from October to January in the former. Fry of about 25 mm are abundant from
November to February, fingerlings 40-70 mm being common during January-February in the

51 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

Chilka Lake. In the Mahanadi estuary, post-larvae of 12-13 mm occur from January to April.
In the Pulicat Lake and at Mandapam fingerlings of about 70 mm are common in January. (G.
Luther, 1973)

Food and Feeding

Grey mullet is a diurnal feeder, consuming mainly zooplankton, dead plant matter,
and detritus. Larval M. cephalus are planktonic feeders in the offshore marine environment
and when they reach the size of about 20 mm SL, they undergo change in diet and started
feeding on benthic organisms. Mullet have thick-walled gizzard-like segments in their
stomach along with a long gastrointestinal tract with multiple pyloric caeca that enables them
to feed on detritus. They are an ecologically important link in the energy flow within
estuarine communities. Feeding by sucking up the top layer of sediments, flathead grey
mullet remove detritus and microalgae. They also pick up some sediment which functions to
grind food in the gizzard-like portion of the stomach. Mullet also graze on epiphytes and
epifauna from seagrasses as well as ingest surface scum containing microalgae at the air-
water interface. The amount of sand and detritus in the stomach contents increases with
length, indicating that more food is ingested from the bottom substrate as the fish matures.
(FAO, Fisheries and aquaculture)

Growth

It has been reported that the larvae hatch at approximately 2.6 mm in length and attain
17.7 mm by 42 days. Overall growth is rapid in the first year, with fish attaining 140-180 mm
SL in tropical and sub-tropical waters (Thomson, 1963) and 130-160 mm SL in more
temperate regions. The maximum size recorded for the grey mullet is 68-72 cm TL from sub-
tropical waters of Lake St. Lucia (Wallace, 1975). Usually Grey mullet attains approximately
300g in the first year and 1.2 kg in the second year.

Migration and Life history traits

Grey mullet is catadromous, frequently found coastally in estuaries and freshwater

environments. Mullets are generally schooling fish and the migration by the adult grey mullet
occurs during the breeding season in different parts of the world. Along the West African
coasts, adult grey mullet can take reproductive migrations of over 400 km (Bernardon and
Wall, 2004). Often the shoals of the ripe mullet congregate in the mouth regions of the
estuaries before moving out to the sea (Wallace, 1975). After spawning, the spent fishes came
back to the estuaries. Usually the larvae are seen near the sea surface between the coast and
the continental slope over the shelf (Ditty and Shaw, 1996).

Young grey mullet first enter the estuaries when the individuals are between 15 and
25 mm SL size and recruitment is only for short periods. The timing of M. cephalus
recruitment in to the estuaries coincides with onset of favourable conditions within the
nursery area usually after the rainy season which ensures productive marine larval habitats
(Payne, 1976; De silva and silva, 1979). In most parts of the world, M. cephalus spawns in
the near shore marine environment, the egg and early larval stages are spent drifting in ocean
Training Manual on Seed Production and Farming of Brackishwater Finfishes I 52
currents. Schools of these fry enter estuaries after a month at the sea (Hsu et al., 2009) and
colonise the entire length of these systems. The juvenile and sub-adult life stages are spent
mainly in estuarine waters and the adults then emigrate to the sea to spawn.

Reproductive biology aspects of Mugil cephalus

Oviparous teleost fishes can be separated into two groups according to their spawning
strategy: the semelparous fishes, which have the a single spawning event during their lifetime
such as some species of salmon (Crespi and Teo 2002) and the iteroparouus fishes, which
have several breeding events during their lifetime. Iteroparous species can be divided into
two sub categories (1) the annual spawner i.e., reproduce only once during the breeding
season each year. (2) the annual multiple spawners i.e., reproduce several times during the
breeding season each year. M. cephalus is annual single spawner and also considered as an
isochronal spawner (Greeley et al., 1987; Render et al., 1995).i.e., all the developing oocytes
in the ovary are at the same stage.

Gonadal development up until an advanced stage occurs in the estuarine waters but
ripe running stage is generally only attained in the marine environment (Bok, 1979; Wallace,
1975). If mature fishes are denied access to the sea during the spawning season, grey mullet
tend to resorb their gonads (Wallace, 1975). However, the success of fertilization and larval
survival of grey mullet depends on the environmental salinity and maximum larval survival is
reported in the salinity range of 30-40 ppt (Lee and Menu, 1981).

Size at first maturity and Fecundity

The size at maturity reported for the species ranges very widely from 22 cm SL from
High brackish lagoon, Southwest Nigeria (Soyinka, 2014) to 43 cm TL from Black sea
(Okumus and Bascinar et al, 1997). The fecundity ranges from 0.2 million eggs to 3.89
million eggs from grey mullets of size 33 cm TL to 58 cm TL respectively (McDonough et al,
2003).

Maturity stages of ovary of Mugil cephalus

In M. cephalus, the external sex distinguishing characters are absent. Generally the
males are slender and smaller than females. During the breeding season, oozing males and
females with large, flabby abdomen (Gopalakrishnan, 1991) could be found. The ovaries are
paired, elongated and covered by thin peritoneal membrane. The lobes are slightly
asymmetrical and the left lobe is smaller than the right lobe. Posteriorly the ovaries are fused
and open to the exterior through a common urogenital aperture just behind the anus. The both
lobes are separable and attached to the dorsal coelomic wall by a thin membranous
mesovarium.

The maturity stages can be classified based on shape, size and colour and a five point
scale is used to describe the maturity stages. The maturity stages of the ovary can be divided
into the following stages.

53 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

 1. Immature

The ovaries appear as reddish, translucent structures united at the posterior end. The
entire gonad occupies one fourth of the body cavity. The oocytes correspond to
previtellogenic oocytes and are characterized by a small size.

Immature ovary inside the body cavity Immature ova are seen under light microscopy.

2. Maturing

Gonads fill almost half of the abdominal cavity and arteries are clearly visible. Ovaries
are reddish yellow with a granular appearance. This stage indicates the onset of vitellogenesis
(endogenous and exogenous) and can be easily distinguished from stage 1 oocytes by the
presence of lipid droplets in the peripheral ooplasm (endogenous vitellogenesis). The average
diameter increases and this stage are generally identified at its beginning by the appearance of
small yellow spots visible to the naked eye that correspond to the stage 2 developing oocytes.
At the end of the stage 2, the ovaries are completely filled up with clearly visible yellowish
oocytes.

Maturing ovary inside the body cavity Maturing ova are seen under light microscopy.

3. Advanced maturation / Mature

A pair of distinctly orange yellowish cylindrical ovarian lobes is visible. The entire gonad
occupies three fourths of the body cavity and is fully packed with yolky oocytes. The ovarian
wall appears as very thin, distended and almost transparent. Ova are yellowish, granular,
round, yolk laden and appear as dark bodies under microscope. These ovaries characterize
fully vitellogenic females, close to the spawning period.

Training Manual on Seed Production and Farming of Brackishwater Finfishes I 54

 Matured ovary inside the body cavity Matured ova are seen under light microscopy.
Arrow indicates immature ova
4. Ripe

Ripe oocytes occur immediately before ovulation. The duration of this stage is short as
the female is undergoing the final rapid development of oocytes before ovulation. The
oocytes are characterized by the migration of the nucleus to the animal pole, and fusion of the
yolk globules and oil droplets. Finally, the yolk appears as a homogenous mass filling the
interior of the oocytes. Before spawning, the oocyte in the ovary are not transparent, but
simultaneously on gaining transparency, the eggs sharply increase in volume and their
specific density increases, allowing them to float in seawater of normal density i.e., an egg
become pelagic (Fulton, 1891). In fishes spawning in sea water, the egg volume increases by
several folds, sometimes by hundreds of times (Wallace and Selman, 1981). The increase in
water content i.e., hydration serves as primary cause for volume and weight gain in a follicle.
It is believed that hydration of oocytes in teleost fishes during maturation is a unique
phenomenon among vertebrates (Wallace an Selman, 1978).

Ripe ovary inside the body cavity Ripe ova are seen under light microscopy.

5. Spent

The ovaries are purple in colour, highly shrunken, collapsed and occupy approximately
half of the body cavity. They appear flaccid and loosely packed with primary oocytes.
Ovarian wall is thick. The immature stock of ova can be seen along with few large
disintegrated yolky oocytes which undergo the process of resorption. Blood shots are
common in spent ovaries.

55 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

 Spent ovary inside the body cavity Ova in the spent ovary are seen under light
microscopy.
Maturity stages of male Mugil cephalus

The testes are attached to the peritoneal cavity by means of mesorchium and protected
peripherally by thin connective tissue, the tunica albuginea. These are tubular, restricted type
and consisted of vasa differentia and a common sperm duct. The two lobes of the testis are
more or less of same length. The stages are divided in to five stages based on colour, shape
and size and classification is adapted from Elizabeth, 1987 with some modifications.

1. Immature

The testis appears as two threads like structures, united at the posterior end. They are
semi-transparent and measure about 5 to 7 cm in length. They occupy almost half of the body
cavity.

2. Maturing

The testes becomes more flattened, ribbon like, appears opaque with smooth surface. The
entire gonad occupies about two thirds of the body cavity.

3. Mature

This stage is characterised by the white, turgid and opaque testes with smooth surface. The
size of the testis is almost similar to that of the previous stage. A small amount of milt oozes
out when pressure is applied to the abdomen. Difference in lobe length is common in this
stage.

M. cephalus testes at different maturity stages a) Immature b) Maturing and c) Mature

Training Manual on Seed Production and Farming of Brackishwater Finfishes I 56

 4. Ripe/Oozing

The size, shape and colour are similar to the previous stage. But the testes appear more
turgid and the milt oozes out freely with slight application of pressure to the abdomen

5. Spent

The testes appear flaccid with surface thrown in to folds. Only a little quantity of the milt
oozes out on application of considerable amount of pressure to the abdomen.

Conclusion

M. cephalus is a detritivore feeder with well-developed digestive system and occupy

the lower trophic level in the food chain contributing significantly to the energy flow in the
estuarine systems. The capacity of fish to survive in aquatic habitats of varying salinity,
turbidity and dissolved oxygen levels makes it suitable candidate for freshwater,
brackishwater and marine farming. It is a high fecund fish with group synchronous ovary;
usually spawns once in the breeding season. The normal development of gonads in the
captivity is influenced by many factors like salinity, temperature, day length, nutrition,
handling stress and hormone dose; sometimes the development of the gonads is stopped due
to the unfavourable conditions, thus leading to atresia (resorption of the oocytes and
spermatocytes).

57 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

 Procurement, transportation and acclimatization of fishes for
broodstock development
Tanveer Hussain, Aritra Bera, Krishna Sukumaran, M.Makesh, Dani Thomas, M.Kailasam

1. Introduction

Successful seed production in the hatchery depends upon the availability of healthy matured
fishes. For selecting potential breeders, viable broodstock under captive conditions has to be
developed. Since seabass attains maturity after 2 years of age to develop broodstock from farm
grown fish, one has to wait more than 2 years. To save time, adult fishes could be procured from
the commercial catches, transported carefully to the hatchery holding facilities and maintained.
Whether from the farm or from the wild catches, fishes have to be procured with care,
transported and follow some protocols for healthy broodstock development.

2. Procurement

Adult Sbrood fishes can be procured from wild catch or from the farm harvest. The selection
of a suitable gear or method of capture is very important in order to avoid injury to the fish.
Mullet is usually caught using drag net, dip net, seine net andcast net Dip net is the best gear
for brood stock fish collection because, dip nets cause less injury to the fish. But operation of
this gear is not possible in many places.

2.1. Criteria for selection of fishes for broodstock development

1. The fish should be devoid of external injuries or internal haemorrhage.

2. Fins should be complete and there should not be any loss of scales

3. The jaw, snout, opercular region, eyes and gills should have not damaged,
since these parts are vulnerable to injuries during capture by different gears
and would lead further infections after released into the holding tanks.

4. The fish should be healthy and free from any parasite infection.

Training Manual on Seed Production and Farming of Brackishwater Finfishes I 58

3. Transportation

Fishes procured have to be carefully transported. If the procurement site is nearby,

transportation can be done manually. Fishes from distant places have to be transported using
transporting vehicles. Transporting vehicles with water holding facility and inlet and outlet
provision are to be selected. Inner lining with soft materials like foam is preferable so that the
fish will not get injured while moving. There should be provision for oxygen cylinder for
providing oxygen while on transportation. While transportation, anesthetic agents like
Aqueous or Chilaldine or MS 222 or Phenoxy ethanol may be used at lower concentration to
keep the fishes on rest.

Manual transportation from nearby areas is done by simply transporting the fishes
using buckets or troughs with water covering with perforated lids in the case of small fishes.
If the fishes are large, (more than 2.0 kgs in size), they can be transported using specially
designed transport tubes. Large rubber tubes specially designed with perforation for water
exchange can be used. Fishes can be allowed into the tube and can be manually toed along
the water-line to reach the hatchery. This would avoid injury to the fish.

4. Acclimatization

After the fishes are brought to hatchery/ holding facility, they should be released in
the acclimatizing tanks with quality filtered seawater of the same salinity, temperature etc., of
the transported medium and mild aeration should be given. Since the fish would be under
stress due to transportation, they can be kept in this tank for 1 – 2 hrs with flow through.
After normalization, the fish can be treated with acriflavin (1 ppm) for 10 minutes and or
with (5 ppm) KMNO4 for one hr as prophylactic treatment to avoid infection on minor
injuries, in any. The fish can be kept under hatchery condition 3-5 days for close observation
and later on can be shifted to brood stock holding facility for further maintenance.

4.1. Quarantine

Fishes collected may have some parasites or other pathogenic infections. To make
sure they are devoid of pathogens fishes have to be kept under quarantine condition in
separate tanks. If any fish develop symptoms of any disease, they should not be used for
broodstock.

59 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

4.2. Conditioning of broodstock

The fishes brought for broodstock development would have lived in different
conditions feeding with various types of feed. In the confined broodstock holding tanks or
cages or ponds, they may not be getting the feed they were used to feed. So, the fishes have
to be slowly conditioned to feed upon the feed which will be provided in the hatchery
conditions by slow weaning, which may take few days for accepting the new feed.

5. Broodstock holding facilities

Healthy broodstock fishes after observing as protocols can be transferred to

broodstock holding facilities like RCC tanks (preferably large tanks of 50 – 100 tonne
capacity) or cages or ponds for further maintenance and development providing required
feed, quality water and nutrition feed for maturation and spawning.

Training Manual on Seed Production and Farming of Brackishwater Finfishes I 60

 Broodstock Development and Induced Breeding in Grey mullet
(Mugil cephalus)

Krishna Sukumaran, M. Kailasam, Prem Kumar, Rekha M.U., Dani Thomas, M. Makesh

Introduction

Taxonomy
Class Actinopterygii
Order Mugiliformes
Family Mugilidae
Genus Mugil
Species Mugil cephalus

The flathead grey mullet Mugil cephalus Linnaeus is the most widespread species of
the family Mugilidae which comprises of 20 genera and 70 species. The species is known by
the vernacular names of “Madavai” in Tamil, “Thirutha” in Malayalam, “Kathiparega” in
Telugu, “Boi or Mangan” in Marathi, “Mala” in Telugu, “Gandhia or Boi” in Gujrati,
“Bhangor” in Bengali. Grey mullet is a cosmopolitan species occurring in all the major
oceans of the world. The species is discontinuously distributed mainly between the latitudes
420N and 420S in the freshwater, estuarine and marine habitats of the world. The species is
recognised economically as an important food fish. The roe of the species is used to prepare
“Bortaga cavier” a delicacy in Taiwan and Japan and hence referred to as “Grey gold” by the
fishermen of the region. In India grey mullet has good market in all the coastal states fetching
between Rs 300-400 per kg. Grey mullet is situated at the base of the food chain and feeds
on detritus and benthic micro-algae thus playing its significant ecological role as a converter
of primary productivity, particulate organic matter and detritus into quality fish protein. The
significant market demand, tolerance to wide salinity ranges and ability to utilise the
herbivorous and detrital food chain qualifies it as an excellent candidate species for
aquaculture.
Globally in the year 2014, the total production of grey mullets was recorded at
1,51,794 t of which 12,360 t was contributed by aquaculture production. Thus aquaculture
made 8.14 % contribution to the total global production of the species, this is significantly
low in comparison to the species like Asian seabass where 41.2 % contribution was made by
aquaculture towards its total production. The reasons which make a fish an ideal candidate
for aquaculture production are related to its seed availability, market value, growth rates,
feeding niche and acceptance of artificial feed, adaptability to aquaculture systems, rearing
environment and resistance to disease. Given its position in food chain and its good market
value, grey mullets have enormous potential for contributing to a sustainable aquaculture
model which is the need of the coming decades. It remain to be debated which of the factors

61 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

whether the seed, feed or the market has to be strengthened to enable the species to realise its
true aquaculture potential.
Currently, the largest contributors to the global aquaculture production of grey mullets
is Egypt, followed by Republic of Korea, Italy, Taiwan province of China and Israel. Most of
the global aquaculture production of grey mullet is reliant on wild seeds. In India, wild seed
availability of grey mullets is during the July to August in the west coast around Puduvypu
region and around October-November in the east coast of India around Kakinada.
Aquaculture of grey mullets in our country is on a limited scale in few pockets. The culture is
reliant on wild seed using natural productivity and supplementary feeding based on agro-bye-
products. There is good scope of improving the fish production if consistent hatchery
produced source is made available. It is in this context that broodstock development and
standardisation of the induced breeding protocols assume significance.
Broodstock Maintenance

A quality broodstock forms the foundation stone of a breeding programme. Different

facets that have to be taken care for maintaining a healthy broodstock.
Broodstock holding system- Lined ponds, tanks and raceways have been used
conventionally for holding the broodstock. At MES, CIBA broodstock have been held in 100
t tons RCC tanks provided with continuous flow through of seawater pumped from a deep
bore. Broodfish are maintained at a stocking density of less than 1 kg m-3. Both these factors
ensures optimal water quality conditions. The tanks are cleaned on alternate days. Relatively
smaller sized fish are being maintained in pond system for developing future broodstock.
Procurement of brooder- Brooders of grey mullet can be procured from the wild or
raised in ponds from early stages however the latter involves an investment of tremendous
effort and time. A sizeable broodstock of grey mullets are desirable. Grey mullet seem to
exhibit a state of social hierarchy in which only a small fraction of dominant females mature
(less than 20%) supressing the maturity of the con-specifics. This makes only a small number
of mature females available for use during the induced breeding in the season.
Broodstock availability- The availability of grey mullet spawners and the peak
breeding season is associated with the north east monsoon around October to January in
Kovalam backwaters (Mohanraj, 1994). The spawning season of grey mullet from Chilka
Lake is from September to December (Jhingran and Natarajan, 1969), Mahanadi estuary –
September to December (Shetty et al., 1965), Goa around September to February (Das,
1978).
Environment and water quality
Photoperiod plays a key role in initiating gonadal development and stimulating oocyte
growth. Water temperature is important for initiating vitellogenesis and regulates oocytes to
functional maturity. Environmental cues especially falling temperatures triggers aggregation
and subsequent spawning migration. Most of the records of spawning’s are recorded in
waters close to 20 0C mostly in deep offshore waters. The best results for attaining functional
maturity for grey mullets are obtained at combination of temperatures and photoperiod of 21
Training Manual on Seed Production and Farming of Brackishwater Finfishes I 62
0
C and 6L/18D respectively. A salinity of 32 ppt is found desirable. Grey mullet females
undergo vitellogenesis irrespective of salinity, however the rate of oocyte growth is slower in
fresh waters as is the proportion of females successfully completing oogenesis. A salinity
ranging from 13-35 ppt has been suggested as adequate for ovarian maturation. Being
confined to freshwaters during the spawning season is considered a major obstruction to the
natural progression of results leading to spawning in the wild (Tamaru et al., 1994).
Natural feed and formulated feed

Grey mullets are predominantly benthic foragers feeding mainly on detritus including
particulate organic matter especially benthic microalgae as diatoms, foraminiferans,
filamentous algae, protists, meiofauna and small invertebrates. Diatoms form 20-30% of the
stomach contents of the fish indicative of its selective feeding habit. This is also indicated by
the relatively long intestine of grey mullets to effectively breakdown diatoms in the diet.
Hence in most of the pond based broodstock, maintenance a substantial quantity of
periphyton substrates is desirable to allow a good surface area for the development of
periphytic organisms.
Grey mullets are regarded as species having a relatively high fat content compared to
other species, 4.9%. Broodstock of grey mullets have been maintained on formulated
maturation feeds developed by CIBA. The world over broodstock of mullet have been
maintained on feeds with a crude protein content ranging from 35- 40 % and a crude lipid
content of 4-8 %. Being bottom feeders sinking pellets are used and the fish are fed at the rate
of 3-5% twice daily. A feed rich in poly-unsaturated fatty acids and arachidonic acid,
adequate vitamin e and carotenoids, astaxanthin are recommended for broodstock maturation
and good larval quality.
Size at maturity and broodstock selection

Males of grey mullets mature between 250- 300 mm standard length while females
mature at slightly larger size, 270- 350 mm. Males are reported to mature at approximately 3
years of size while females mature at 4 years. A minimum fork length of 310 mm or three
years of age is suggested best for selection of broodstock.
Reproductive biology of grey mullets

Spawning grounds of grey mullets are located in the sea. Grey mullets are generally
reported to spawn once a year and exhibit synchronous ovarian maturation. Fecundity is
reported in the range if 1.2- 2.8 million for the species (Thompson 1983) and 0.5-2 million
(Bester 2009) and 849 eggs per g body weight (Nash et al., 1974). The optimum temperature
reported for egg development of grey mullet is 24 0C. The most suitable ova size for a
successful induced spawning is 600 micro-metre i.e. when the oocyte is in the tertiary yolk
globule stage, stage III. A fertilised egg is approximately 870 micro metre in size with an oil
globule of approximately 350 micro metre. The reported incubation time is approximately 26
h at 25 0C and can range upto 48- 60 h based on the temperature conditions.

63 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

 Parasites- Grey mullets can act as hosts for a number of parasites. Visual observation
and periodic examination of the fish for parasites is conducted. Fish should be carefully
observed daily for reduction in feed intake and swimming activity as signs of parasitic
infection. Periodic chemical treatment on a monthly basis is done using 100 mg L-1 of
formalin for 45 minutes. Infections of external crustacean parasite Caligus spp. and
Lernanthropus spp. have been reported in the grey mullet broodstock maintained at MES,
CIBA.
Hatchery production- Global status

The history of induced breeding of grey mullet dates back to the 1960’s in the
pioneering work of Tang (1964). Full scale commercial hatchery production of grey mullet is
not yet common. Induced spawning is achieved on an experimental and semi-continuous
basis at Hawaii, United States of America and Taiwan province of China. Few of the steps
followed are outlined. Egypt, the largest producer of cultured grey mullet (over 90 % of the
global production) has one experimental mullet hatchery producing few lakh fry annually and
largely depends on wild collection of grey mullet fry for aquaculture. Italy is another major
producer of cultured grey mullet. Unlike Egypt, most of the cultured fry in Italy originates
from hatchery produced mullet fry.
Maturity assessment

Maturity assessment of the broodfish is conducted with the approach of the spawning
season. The fish after being caught carefully and anaesthetised in 30 ppm 2-phenoxyethanol.
These fish are cannulated to assess the ova diameter and the condition of the eggs. Reports
suggest an ova diameter of 600 micro-m to be optimum for successful induced spawning.
Induction of maturity

For advancing maturity, CIBA uses hormone based pellets of LHRHa. A 200 micro-g
LHRHa pellet is suggested for implantation in the dorsal musculature of the fish (Tamaru et
al., 1989) for advancing maturity of the female of grey mullets. Use of a combination of
LHRHa and testosterone pellets have been shown to result in accelerated oocyte growth. For
males silastic implants containing 17-α-methyl-testosterone containing 10 mg 17- α-methyl-
testosterone has been found effective for 10 months in inducing testicular maturity (Lee,
1992).
Induced breeding and larval rearing

On obtaining mature fish with ova diameter exceeding 600 micro-m, a priming dose
of 20 mg kg-1 of carp pituitary homogenate and a resolving dose of LHRHa, 200 micro-g per
kg-1 is considered as the best combination for induced breeding via intra-muscular injections.
The fish are kept at a ratio of 2-3 males: 1 female for breeding. The fish are reported to spawn
12- 14 h of receiving the resolving dose. The eggs are incubated at around 500 no L-1 in
aerated tanks. High salinities of above 35 ppt are recommended for incubation of eggs. At 26
0
C an incubation time of 28 h is reported, based on the temperature incubation period of 48-

Training Manual on Seed Production and Farming of Brackishwater Finfishes I 64

60 h is also reported. Larvae of grey mullet begin feeding on the third day when they are
provided algae and rotifers. Feeding of artemia nauplii is initiated on the seventh day.
Other inducing agents used successfully in different studies

Partially purified salmon gonadotropin, SG-G100 has been recommended at doses between
12-21 micro-g per kg body weight in inverse proportion to the egg diameter ranging above
600-700 micro-m. One third of the total dose is given initially followed by the remaining
two-thirds after 48 h.
HCG- Priming dose of 20000 IU per kg followed by a resolving dose of 40000 IU per g after
24 h in fish with oocyte dia of 600 mcro-m (Kuo et al., 1973)
LHRHa- 300- 400 micro-g per kg body weight, one third as priming dose and two third as
resolving dose after 24 h (Lee et al., 1987).
Many successful combinations were tried by Lee et al., (1988). A priming dose of HCG at
5000IU and a resolving dose of LHRHa at 200 micro-g per kg resulted in 100% spawning
rate, so also an LHRHa priming dose of 200 micro-g per and a resolving dose of 20 mg per
kg. However the best fertilisation rate of 66-86% was reported was using a combination of
CPE and LHRHa at 20 mg kg-1 and 200 micro-g per kg respectively.
Priming dose- 20-70 mg CPE or 10,000 IU HCG and a resolving dose of 200 micro-g
LHRHa, ova dia- 570-580 micro-g was reported for successful spawning and fertilisation by
Gharabawy and Assem (2006).
In grey mullets strong dopaminergic control is reported, hence more recently, GnRHa-
priming dose- 10 micro-g per kg+ a dopamine antagonist- metoclopramide, 15 micro-g per
kg and resolving dose- 20 micro g per kg+ a dopamine antagonist- metoclopramide, 15
micro-g per kg were given 22.5 h apart for successful spawning (Aizen et al., 2005).
Further reading

Aizen, J., Meiri, I., Sivan, B.,L.,Rosenfeld, H., 2005. Enhancing spawning n grey mullet
(Mugil cephalus) by removal of dopaminergic inhibition. General and Comparative
Endocrinology, 142, 212-221.

Kuo, C-M., Shehadeh, D.H., Nash,C. E., (1973) Induced spawning of captive grey mullet
(Mugil cephalus) females by injection of human chorionic gonadotropin (HCG). Aquaculture
1, 429-432.

Lee, C.S., Tamaru, C.S. (1988) Advances and future prospects of controlled maturation and
spawning of grey mullet (Mugil cephalus) in captivity. Aquaculture, 74, 63-73.

Lee, C.S., Tamaru, C.S., Miyamoto, G.T., Kelley., C.D., 1987. Induced spawning of grey
mullet (Mugil cephalus) by LHRHa. Aquaculture, 62, 327-336.

Meseda, M., El-Gharabawy, Samira, S., Assem, S., 2006., Spawning induction in
Mediterranean grey mullet Mugil cephalus and larval development stages., African Journal of
Biotechnolgy, 5(19) 1836-1845.

65 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

Mohanraj, G., Nammalwar, p., Kandaswamy, S., Sekar, A.C., 1994. Availability of grey
mullet spawners in Adayar estuary and Kovalam backwater around Madras, India. J. mar.
boil. Assn. India, 36 (i-2): 167-180.

Shehadeh, D.H., Nash,C. E., (1973) Establishing broodstock of grey mullets (Mugil
cephalus) in small ponds. Aquaculture 2, 379-384.

Tamaru, C.S., Fitzgerald, W.J., Sato., V., (1993) Hatchery manual for the artificial
propogation of striped mullet Mugil cephalus (Ed: C.C. Trick). Guam Aquaculture and
Training centre, Technical Report 15.

Tang, Y-A, 1964. Induced spawning of striped mullet by hormone injection. Japanese journal
of Ichthyology, 12: 23-30.

Training Manual on Seed Production and Farming of Brackishwater Finfishes I 66

 Nursery rearing and Grow-out Culture of Grey Mullet
(Mugil cephalus)

G. Biswas, M. Kailasam, Tanveer Hussain and R. Subburaj and G. Thiagarajan

Introduction

Among 70 valid species of mullets belonging to 20 genera under the family

Mugilidae, the species having highest growth is widely called as striped/ flathead grey mullet,
Mugil cephalus. This species has a cosmopolitan distribution between latitude 40°N and 40°S
covering all the oceans. It is an economically important euryhaline and eurythermal species
contributing to a sizable fisheries of estuarine and coastal regions of countries like China,
Egypt, India, Israel, Italy, New Zealand, Nigeria, Sri Lanka, Taiwan and Tunisia. Owing to
omnivorous feed habit of grazing on plant detritus and microflora, it is an ecologically
important species feeding at the lowest trophic level and suitable for mono or polyculture. To
the world mariculture production, a substantial contribution is shared by striped/ flathead
grey mullet as one of the species.

In India, M. cephalus is a much relished for its flesh quality and good flavour and
farmed scientifically in brackishwater ponds and impoundments. However, traditional and
semi-intensive pond framings mainly depend on availability of seeds. Fry of M. cephalus
migrate into estuaries of south-east and south-west coasts immediately after onset of south-
west monsoons in November-April and north-east coast in January-March and July-August.
The small fry (15-25 mm) are not suitable for direct stocking in grow-out ponds. Since the
growth of this fish is slow during early life stage, it is desirable to conduct a pre-stocking
seed rearing to obtain bigger size individuals suitable for grow-out culture. In culture ponds,
the grey mullet accepts artificial feed in presence of natural food organisms. Pond
fertilization using organic and inorganic manures affects the growth of grey mullet.
Therefore, utilizing the natural pond productivity and employing other interventions grey
mullet seeds are reared for production of stockable size fingerlings.

Seed rearing of grey mullet

Among different seed rearing methods, such as only fertilization or feeding, combined
fertilization-feeding, fertilization-compost application and fertilization-periphyton systems,
the best performances of fish can be obtained in the combined fertilization-feeding and
fertilization-periphyton rearing systems.

67 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

Low density fertilization-feeding system

After treatment of pond bottom with lime, water is taken and fertilized with cattle manure,
urea and single super phosphate at 500, 30 and 30 kg/ha, respectively. After 7 days of
fertilization, ponds are stocked with M. cephalus fry (0.55 g/ 36.0 mm) at 15000 nos./ha.
Formulated feed prepared from locally available ingredients (mustard cake, rice bran, wheat
flour, fishmeal etc.) is provided as supplementary feed @ 20 to 5% of body weight. Ponds are
fertilized fortnightly with the above mentioned fertilization materials at the same dose.
Liming is done at fortnightly intervals with lime stone powder at 250 kg/ha. After 150 days
of rearing, grey mullet attains average body weight (ABW) of 96 g.

High density fertilization-periphyton system

After bottom treatment followed by water taking according to the method mentioned earlier,
ponds are fertilized with mustard cake, urea and single super phosphate at 200, 20 and 20

Operational cost (OC) Low density FF High density FP

Grey mullet fry 15000 @6/- 90000 30000 @6/- 180000
Feed 2000 kg @35/- 70000 -
Other inputs 30700 55370
Manpower 12500 10000
Sub-total 203200 245370
Interest on OC @ 10% annually
For 5 months 8467 For 4 months 8179
Total OC 211667 253549
Return from sale of fingerlings
Grey mullet fingerlings 12630 nos. @30/- 378900 28290 nos. @20/- 565800
Net return 167233 312251
Benefit-cost ratio (BCR) 1.79 2.23

kg/ha, respectively. After 6 days, bamboo poles are erected vertically in the pond to cover
10% of pond surface area as substrate for periphyton growth. After 10 days of bamboo pole
fixing, pond is stocked with M. cephalus advanced fry (3.36 g/ 63.7 mm) @ 30000 nos./ha.
During rearing, all the ponds are fertilized fortnightly with mustard cake at 100 kg/ha.
Agricultural lime at 100 kg/ha is applied one day before fertilization throughout the rearing
period. Grey mullet fingerlings attain ABW of 28 g in 120 days of rearing.

Training Manual on Seed Production and Farming of Brackishwater Finfishes I 68

Table: Economic analysis of combined fertilization-feeding (FF) and fertilization-periphyton (FP)
systems for grey mullet seed rearing. Calculation is for 1 ha pond and currency mentioned is Indian
Rupee.

Grow-out culture of grey mullet

Monoculture

Monoculture of grey mullet depends on availability of suitable seeds as stocking

materials. Various on-station and out-station trials have proven that monoculture of grey
mullet can be an economically viable farming option provided that ponds are stocked with
seeds reared initially in nursery. The pond for monoculture is prepared first, following
eradication of unwanted organisms and application of manures and fertilizers. Advanced
fingerlings of >50 g size are stocked at 10000 nos./ha. Fish are fed with supplementary feed.
In an 8-month culture, fish become 500-800 g with total production of 3-4 ton/ ha.

Polyculture

Polyculture is a farming practice where two or more species of fishes are reared
together. The concept of polyculture is based on rearing of two or more compatible aquatic
species together resulting in higher production compared to monoculture. The underlying
goal of polyculture involves increasing productivity by more efficiently utilizing ecological
resources within an aquatic environment. Sometimes, one species enhances food availability
to other species and thus increases total fish production per unit area. It is commonly believed
that polyculture gives higher production than monoculture in extensive and semi-intensive
systems and is considered more ecologically sound than monoculture. Before stocking of
seeds, pond is prepared well following eradication of pest and predatory fishes, removal of
bottom mud and liming, fertilization etc. The ready ponds are stocked with seeds of fish
species at 8000-15,000 nos./ha along with tiger shrimp seeds of 15,000-30,000 nos./ha. The
stocking density varies with the quantum of seed availability. Natural pond productivity is
maintained by fertilization. In addition, supplementary feed prepared from locally available
ingredients can be used at 2-5% body weight daily. This kind of system can yield a total
production of 1.5-3.0 ton/ha in 6-10 months. The preferred species among fishes are: mullets-
M. cephalus, Liza tade (tade grey mullet), L. parsia (goldspot mullet), milkfish- Chanos
chanos, pearlspot- Etroplus suratensis and tiger shrimp- Penaeus monodon. In an out-station
trial, 3 ponds (1 ha each) were stocked with M. cephalus (40-50 g size) as the major species
at 10000 nos./ha, L. tade (10 g size) at 2000 nos./ha, pearlspot (10 g size) at 1000 nos./ha and

69 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

P. monodon at 4500 nos./ha. Supplementary feeding was provided with floating feed. After
150 days, shrimp with 45 g ABW was harvested and fishes were harvested after 300 days. It
resulted in 2.8 ton/ ha production with a net return of Rs. 2.9 lakh/ ha. In another out-station
trial, ponds were stocked with M. cephalus at 5000 nos./ha, L. tade at 5000 nos./ha, L. parsia
at 10000 nos./ha and P. monodon at 40000 nos./ha. After 300 days, total production of 3.6
ton/ ha with a net return of Rs. 5.6 lakh/ ha was achieved.

Conclusion

Major constraint in mullet culture is inadequate availability of seeds. The natural seed
availability has become uncertain and sporadic now-a-days. Breeding and seed production of
mullets are yet to be achieved in India. Concerted effort on breeding and seed production has
been made by ICAR-CIBA since almost two decades and success in breeding will meet up
the seed requirement soon.

Further reading

1. G. Biswas et al., 2012. Effects of stocking density, feeding, fertilization and combined
fertilization- feeding on the performances of striped grey mullet (Mugil cephalus L.)
fingerlings in brackishwater pond rearing systems. Aquaculture 338-341, 284-292.
2. G. Biswas et al., 2012. Evaluation of productions and economic returns from two
brackishwater polyculture systems in tide-fed ponds. Journal of Applied Ichthyology 28,
116-122.
3. G. Biswas et al. 2016. Effect of feeding, periphyton substrate and compost application on
the performances of striped grey mullet (Mugil cephalus L.) in fertilized brackishwater
nursery ponds. In: Book of Abstracts, International Conference on Aquatic Resources and
Sustainable Management, 17-19 February 2016, Kolkata, pp. 162.

Training Manual on Seed Production and Farming of Brackishwater Finfishes I 70

 Breeding and seed production of pearlspot, Etroplus suratensis (Bloch)

Krishna Sukumaran, Rekha M.U., K.P. Kumaraguru Vasagam

Introduction

Class- Actinopterygii

Order- Perciformes

Family- Cichlidae

Genus- Etroplus

Species- suratensis

Pearlspot, Etroplus suratensis, is distributed in peninsular India and Sri Lanka. Its tolerance to
wide range of salinities makes aquaculture of the species possible in both freshwaters and
brackishwater bodies. Being omnivorous in nature, aquaculture of pearlspot is relatively simple,
economical and especially suitable for small scale aquaculture for supporting livelihood of fish-
farmers. Pearlspot is extensively farmed in brackishwaters of Kerala has shown productions upto
1t/ha when cultured with milkfish and mullets (George, 1971). Traditionally pearlspot has been
cultured in pokkali fields of Kerala along with other brackishwater fishes. Pearlspot has chiefly been
cultured by farmers as a component of polyculture in brackishwater systems. Small scale cage based
aquaculture experiments showed that stocking pearlspot @ 200 nos m-3 in 2 m3 net cages can give a
production of 26 kg m-3 in 200-260 days using commercial feed (crude protein-20%) (Padmakumar,
2009). More recently with the support of the state fisheries department many farmers and Self-Help
Groups (SHG’s) in Kerala are involved in culture of pearlspot in small cage (2-3 m3) and pond
systems. However, one of the major limiting factors for expansion of pearlspot aquaculture is
inadequate availability of seed for stocking in different culture systems.

Pearlspot exhibits a high degree of parental care and has very low fecundity as compared to
other brackishwater fishes. Successful induced breeding of pearlspot has not been reported. These are
the main reasons which makes mass scale seed production of the fish challenging. Hence development
of technologies which allow seed production at multiple locations in the form of backyard hatcheries
or small scale seed production systems is important. However the fish is easier to breed compared to
many other brackishwater fish and today different models in a range of systems are available or being
tested, so that seed production can be conducted by entrepreneurs, Self- Help Groups or farmers
themselves depending on their local resources.

71 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

The different methods of pearlspot seed production are discussed in this chapter.

i) Seed production of pearlspot in ponds

In pond based experimental trials conducted in CIBA, 50 brooders were stocked in ponds of
area, 100 m2 and depth, 1.2 m after systematic pond preparation (draining, liming, weed fish
eradication, manuring for promoting phytoplankton bloom). Additional spawning surfaces as
palmyrah leaves tied in bunches to fixed poles, coconut leaf petioles, coconut husks, bricks, pieces of
asbestos sheets etc. were provided. Salinity ranged between 15- 30 ppt and transparency was higher
than 0.5 m. Brooders were fed with artificial feeds prepared using groundnut oil cake, rice bran and
fish meal fortified with vitamins and minerals @ 2.5%. On observing the presence of hatchings,
manuring was done with cow dung @500kg ha-1 for enhancing of plankton production. Artificial feed
(25-30 g) was also fed once early in the morning. A production of 3500 fry was observed in a year
from 5 sets of breeding (Abraham and Sultana, 1995)

ii) Breeding of pearlspot in RCC tanks

Breeding of pearlspot fish has been standardized using 20 t RCC tanks provided with
continuous water flow through. Half of the tank bottom is provided with a soil base (4 inches), earthen
tiles for egg attachment and hide outs are provided within the tank. The tanks are stocked using
mature pearlspot brooders. The brooders are selected based on the size, sex of the fish and
bright coloration. A stocking density of 20 fish per tank at a male female ratio of 2:3 was used. The
male and female fish are identified based on the appearance of the genital papillae. Pair formation
and breeding occurs naturally. Eggs are deposited on the tiles and also on the walls of the tank. Fry
are collected at regular monthly intervals by lowering the water level. A production of 1200-3500
seed per batch can be obtained regularly. Seeds produced from this tank breeding system are supplied
to farmers and self-help groups.

iii) Seed production of pearlspot in small net cages/ hapas

In CIBA seed production of pearlspot has also been tried in small net cages (hapas) in the
secondary discharge pond of fish hatchery. The pond has conditions of gentle water flow, salinity of
approximately 25-30 ppt. Brooders are maintained in small cages on commercial fish feed. Small net
cages/ hapas are being used (dimensions, 1x0.75x1 m). These are fixed by casurina poles. Clay soil in
small plastic tubs is suspended at 0.5 m depth from a cross-fixed casurinapole. Just above the soil
surface 1-2 ceramic tiles are suspended to facilitate egg attachment. Each cage was stocked with 3-4
brooders (TL> 150 mm) and preferably with one fish having reddish and enlarged genital papillae
(indicative of readiness for breeding). Efforts at pair formation are usually observed a few hours after
release of fish within cages. Gradually the dominant pair occupies the soil container and territorial
defense centered on the plastic tub is also observed. The aggressive behavior of the fish increases

Training Manual on Seed Production and Farming of Brackishwater Finfishes I 72

towards breeding and thereafter continues as a defense for protecting eggs and larvae. Nest formation
was observed in the course of the breeding behaviour by mouth siphoning of the soil which led to
formation of small pits within the soil containers.

In the initial trials hatchlings were collected from pit nets in cage and subsequent larval rearing
was practiced, following this method a seed production of 1000-1500 seed per cage could be
observed. However, it is not always possible to observe the correct day of spawning especially if the
water is not transparent. The majority of seed production trials were conducted by allowing seed to be
reared within cage system with parental care. A production between 200-300 numbers of seed (avg.
lenth: 28.11±1.49 mm; avg.wt: 0.66±0.04 g) were observed per cage within 2- 2.5 months. The
advantage of the system is that it is simple, and adoptable by farmers especially cage farmers. It
requires a minimum investment (Rs. 400-500/unit) and the labor involved is chiefly in the initial cage
setting and final seed collection. This model was tested at with a pearlspot cage farmer at Ashtamudi
lake, Kerala. Using a set of eight hapas the farmer produced approximately 3000 fry in six months.

vi) Seed production of pearlspot in tank based recirculation system

Breeding trials of pearlspot breeding were conducted in one ton rectangular plastic tanks
provided with a continuous water flow using a biofilter facility. Each tank was provided with a small
plastic tub filled with clay soil to facilitate breeding. Each breeding tank was stocked with 4 mature
brooders (total length>160 mm) and fed with pellet feed twice a day @ 2-3 % body weight. Pairing in
the tanks could be observed within 2-3 days of stocking and the paired fish were observed to occupy
the soil filled plastic container provided. The aggressive behaviour was observed to centre on this
container and the breeding pair was seen to actively chase other approaching fishes from it. The
aggressive behaviour increased towards the approach of breeding, even leading to the mortality of the
remaining fish. Towards breeding the fish were observed to clean a small patch at the sides of the soil
filled container. The first breeding was noticed after 24 days of stocking. The eggs were seen to be
attached to the sides of the plastic container and the brood fish were observed to take turns in
defending the eggs. The larval clutch observed at the bottom of the soil filled container were separated
for larval rearing. One of the most promising results obtained was the production of 8000 larvae by a
single pair stocked in one tank in six breedings at an average breeding interval of 17.6 ±1.12 and an
average larval number per spawning of 1333.3±143.0. Pearlspot larvae collected from the tanks have
been reared using alternate live feeds, rotifer Brachionus plicatilis, artemia nauplii and by co-feeding
with commercial larval diets. On an average, by this method, a seed production of up to 1000 fry (2.0
cm size) per tank and per month can be obtained in 30 days period and annual total production of up
to 12000 seed per tank per pair of pearlspot.

73 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

vii) Breeding of pearlspot as single pairs in outdoor green water tanks using formulated
maturation feed

Though pearlspot is naturally a low fecund fish, it has a great potential for farming in varying
salinities ranging from 0 to 30 ppt. Maximum reproductive potential any fish can be realized
maximum with an optimum nutritional back-up and suitable rearing system. CIBA formulated a
maturation diet to have balanced amino acids and fatty acids to meet the minimum requirements
optimized for cichlid fishes. This maturation diet in combination with a more economical out-door
green water rearing system was found to be optimum in exposing the maximum breeding performance
in pearlspot. We have achieved repeated spawning and higher fry yield with single pairs. While
average fry production per pair per spawning was around 2500, a highest fry yield recorded was 3480
per spawning. While average number of repeated spawning per year was around 4 times, a maximum
of 8 repeated spawning was recorded. Average inter-spawning days ranged between 35 to 40 days.
Highlight of this breeding model is, we can have a good control over the parental fish and their young
ones produced. Further it is more economical in terms of water management and feed management for
both parents as well as their young ones.

With a planned feeding strategies and rearing model we were able to close the life cycle of
pearlspot by the age of 11 – 12 months in small one ton tanks itself. This is a good indication that,
pearlspot could be a good candidate for genetic selection. Termination of parental care was found to
be a key driver in inducing the pearlspot for recurring spawning. Hence, larval rearing in the absence
of parental care would be a most crucial in successful seed production of pearlspot under captive
conditions.

Conclusion

Pearlspot is emerging as an important food and ornamental brackishwater fish. In the recent
years there has been an increased research focus on different aspects of pearlspot seed production.
This has resulted in the emergence of alternate technologies which can be utilized by the stakeholders.
We definitely see a promise that in the near future one of the main constraints of inadequate seed
availability will be overcome and we may witness substantial increase in aquaculture production of
pearlspot.

References

Annual report 2008-09- Central Institute of Brackishwater Aquaculture (ICAR), Chennai pp. 25.

Padmakumar, K.G.,Manu, P.S.,Bindu, L. (2009) Open water farming of pearlspotEtroplussuratensis

(Bloch) in low-volume cage.Asian Fisheries Science 22(2): 839-847.

George, K.C.(1971)Salt water fish farming. Seafood Export Journal, 7(11): 7-14.

Abraham, M and Sultana, M. (1995) Biology, fishery, culture and seed production of the
pearlspotEtroplussuratensis (Bloch).

Training Manual on Seed Production and Farming of Brackishwater Finfishes I 74

 Brackishwater ornamental fish breeding techniques

S.N.Sethi, Babita Mandal, Aritra Bera and G.Biswas

Aquatic ornamental industry is today a multi-billion dollar industry with an estimated

value of 15 billion US dollars in which 1,500- 1,600 species are traded globally (Moorhead
and Zeng, 2010; Oliver, 2001). The bulk of the ornamental fish traded constitute of the
freshwater fish (almost 90%), however in terms of value their marine counterparts contribute
significantly higher. A notable difference is that the freshwater species are mostly captive
bred (approximately 90%) and that the marine species are collected from wild, 90- 95%
(Oliver, 2001). USA, Europe and Japan are the largest international markets for ornamental
fish; however, Asia is home to more than 65% of the exports (Ghosh et al., 2003). Singapore
has been a consistent leader in ornamental fish exports, followed by Indonesia, Malaysia and
China. In India, Kolkatta has emerged as the major hub for ornamental trade accounting to
almost 90% of the exports followed by Mumbai and Chennai. India can categorise its
ornamental fish species being traded into two categories; the exotic ornamental and native
fish of India. The exotic with almost 288 varieties dominates the domestic market. One
hundred and eighty seven indigenous species are traded from India. The wild catches form
the bulk of the exports (85%) as compared to the cultured ones (Rani et al., 2013). The
ornamental fish market has been showing steady improvement in India with the export values
touching USD 3.8 million, a growth rate of 14.4 % has been recorded in the ornamental fish
export. India’s favourite export destination is Singapore (42.85%), followed by Japan
(13.88%) and Malaysia (9.97%). Freshwater fish dominate the scenario of cultured
ornamental fish species; Molly, Guppy, Platy, Swordtail, Barbs, Cichlids, Angels, Siamese
fighter, Tetras, Gold fish, Manila carps, and Sharks (Ghosh et al., 2000). The existing
scenario of export market based on wild collection is not a healthy one. Efforts in developing
and propagating seed production of untapped indigenous species will go a long way for
developing a robust ornamental fish industry in India. In this regards, CIBA has placed a
major thrust on developing seed production technologies of many commercially important
species. Along with developing ornamental fish culture as large scale production models,
CIBA places a major emphasis on developing ornamental fish as a livelihood option.

Presently the important candidate species on which CIBA places a major thrust are
Spotted Scat, Scatophagus argus, Silver Moony fish, Monodactylus argenteus, Green
Chromide Etroplus suratensis, Orange chromide, Etroplus maculatus, Banded Chromide,

75 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

Etroplus canrensis and Crescent perch, Terapon jarbua are some of them, however, there
also lies great opportunity to do so in a sustainable manner by investing in research for
development of captive seed production technology for these brackishwater species.

Breeding aquarium fish is one of the steps to becoming a skilled aquarist. In order to
breed a species, the aquarist usually needs to be able to distinguish between the sexes and to
be able to recreate natural conditions to stimulate spawning process. There are many steps
(techniques) to be followed to breed an aquarium fishes such as follows:

I. Fish Sex Determination

Determining the sex of a fish is an important step in knowing whether one has a pair.
Most fishes can be categorized as sexually dimorphic or sexually isomorphic. In sexually
dimorphic species, the sexes can be easily distinguished by primary (shape of sex organs) and
secondary differences (size, shape, color). Male’s fishes are frequently more colorful, larger,
and have more elaborate finnage. Among the more brilliant outstanding of sexual
dimorphism can be found in Cichlids, Killifish, and Livebearers. In sexually isomorphic
species, there are minute, if any, apparent sexual differences. Often, the only way to
distinguish between the sexes is the shape of the genital papilla, which is only visible around
spawning times. In some isomorphic species, the males are slightly larger and the females are
slightly rounder in the belly. Some sexually isomorphic species have no known external
sexual differences.

II. Selection of Brood/Parent Fishes

Once males and females have been distinguished, a suitable pair or spawning group
should be chosen. There are several important traits to seek in choosing the parent fish.
Choosing fish that display good markings and color that should produce attractive young fries
and fingerlings. Use mature, healthy compatible fish for spawning because unhealthy fish, if
they spawn, may produce unhealthy or deformed young fishes.

III. Reproductive Strategies

A. Egg-layers Fishes

The majority of aquarium fish are egg-layers with external fertilization. Egg-layers
can be divided into five groups: egg-scatterers, egg-depositors, egg-burriers, mouth-brooders,
and nest-builders.

Training Manual on Seed Production and Farming of Brackishwater Finfishes I 76

Egg-scatterers: These species simply scatter their adhesive or non-adhesive eggs to fall to
the substrate, into plants, or float to the surface. These species do not look after their brood
and even eat their own eggs. These, often schooling, fish may spawn in groups or in pairs.
Example: Egg scatterers with non-adhesive eggs, Zebra Fish, Danio rerio and Egg scatterers
with adhesive eggs Gold fish, Carassius auratus submerged Hydrilla plants are used as
substrate to attach adhesive eggs.

Egg-depositors: These species deposit their eggs on a substrate (tank glass, wood, rocks,
plants). Egg depositors usually lay fewer eggs than egg-scatterers, although the eggs are
larger. Egg-depositors fall into two groups: those that care for their eggs, and those that do
not. Among eggs depositors that care for their eggs are Cichlids and some Catfish. Egg-
depositors that care for their young can be divided into two groups: Cavity spawners and
Open spawners. Cavity spawners lay their eggs in a cave, while open (shelter) spawners lay
their eggs on an open surface. These fish form pairs and have advanced brood care where the
eggs are defended and cleaned. The eggs take a few days to hatch, and the fry are often
guarded by the parents. Various Catfish, Cyprinds, and Killifish make up the majority of egg-
depositors that do not care for their young. These species lays their eggs against a surface,
where the eggs are abandoned. These species do not usually eat their eggs. Examples Barbs,
Rasbora sp.

Egg-burriers: These species usually inhabit waters that dry up at some time of the year. The
majority of egg burriers are annual Killifish,( Aplochelius sp.) which lay their eggs in mud.
The parents mature very quickly and lay their eggs before dying when the water dries up or in
drought condition. The eggs remain in a dormant stage until rains stimulate hatching. They
grow up to 3-4cm total length and are short lived.

Mouth-brooders: Mouth-brooding, also known as oral incubation and buccal incubation, is

the care given by some groups of animals to their offspring by holding them in the mouth of
the parent for extended periods of time. Paternal mouthbrooders include the arowana
(Osteoglossum bicirrhosum), Betta, Betta pugnax, and Sea catfish, Ariopsis felis, Arius sp.,
Tilapia, Sarotherodon melanotheron, and Snake head, Channa striatus. Fish species that
carry their eggs or larvae in their mouth. Mouth brooders can be broken up into ovophiles and
larvophiles. Ovophile or egg-loving mouth-brooders lay their eggs in a pit, which are sucked
up into the mouth of the female.

77 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

Nest-builders: Nest builders build some sort of nest for their eggs. The nest is usually in the
form of bubble-nest formed with plant debris and saliva-coated bubbles (Siamese –fighter
fish, Angel fish, Labyrinth fish, Catfish), or an excavated pit in the substrate (Cichlids). Nest
builders practice brood care.

B. Livebearers

Livebearers are fish that bear live young. There are two types of livebearers: ovoviviparous,
where the eggs form and hatch within the female before birth; and viviparous, where no eggs
are formed, and the young are nourished through an umbilical-like cord or from secretions by
the female. Livebearers are often prolific, easily bred species. One of the best ways to induce
fish to spawn, especially difficult-to-spawn species, is to simulate natural conditions.
Examples are Guppy, Platy, Mollies etc. Among factors that encourage fish to spawn are the
environment, the food, and the rainy season.

Environmental Parameters

The right water conditions are among the most basic requirements in spawning of
fishes. Thus the water conditions should be similar to those in the natural environment of the
species. By following the suggestions under "breeding" or "water" in the species descriptions,
approximate natural water conditions can be found. Another important environmental
condition is the right tank set-up including hiding places, spawning sites, and lighting, water
current and social conditions.

Food

The right foods are important to encouraging spawning. Without proper foods, natural
conditions cannot be entirely recreated. Some of the live foods that often can make a
difference in spawning success are mosquito larvae and fruit flies.

Monsoon Rainy Season

Many fish species spawn during the rainy season in nature. By simulating the rainy
season in aquaria, difficult-to-spawn species can be induced to spawn. Rains affect the water
chemistry, the water height, and the water temperature also.

Target Conditioned Fish:

Training Manual on Seed Production and Farming of Brackishwater Finfishes I 78

Paired Fish/Target fish can be used to help strengthen the bond between a fish pair. Target
fish can be another of the same species or a similar species that is placed in the tank with the
breeding pair. This third fish will serve as an object of the aggression of the pair. The pair
will work together to chase off the target fish and not fight between themselves. Only use the
target fish method in a large tank with plenty of hiding places, so that the target fish is not
harmed. PVC Pipes can be used as hide outs in the breeding tanks.

I) Spotted Scat, Scatophagus argus L. 1766

Taxonomy Position

Class- Actinopterygii

Order- Perciformes

Family- Scatophagidae

Genus- Scatophagus

Species- argus

The species is distributed in the Indo-Pacific, Southern India, Sri Lanka, Southern
Japan and Tahiti. It inhabits coastal muddy areas, lower courses of rivers and mangrove
areas. The fish is an omnivore feeding on detritus, filamentous algae, phytoplankton,
macrophytes and zooplankton. It attains a maximum total length of 380 mm. Females of the
species are reported to mature at 7-8 months at 150 g size while males mature at relatively
smaller size (Barry & Fast, 1992). Care has to be taken during handling of the species due to
its venomous spines on the dorsal, anal and pelvic fins which can cause pain for long hours.

CIBA has successfully developed protocols for captive maturation and induced
breeding of spotted scat (Kailasam and Arasu, 2011). Brood fish (up to 250- 300 g) size were
developed in ponds and tanks by providing optimal environmental conditions and feed for
accelerating maturation. Successful spawning of the captive broodstock was achieved by
hormonal manipulations. A female fish weighing 200 g with ova diameter of 426 μ was
selected and administered with Human Chorionic Gonadotropin (HCG) hormone as a prime
dose, followed by Luteinizing Hormone and Releasing Hormone (LHRHa), as a resolving
dose. Male fishes were also administered with the same hormones. Forty-eight hours after the
treatment, fishes responded and ovulation was observed. Ovulated eggs and milt were

79 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

stripped from fishes and fertilization was facilitated externally. Larvae hatched out after 19
hours of fertilization, and average size of the larva was 1.62 mm. Larvae were fed with
rotifers from day 3, up to day 10, and afterwards with brine shrimp, Artemia nauplii up to day
25, till they reached 7-9 mm size. Fry were weaned to formulated feed and reared further.
The hatchery produced juveniles were supplied to entrepreneurs for further propagation. The
fish fetches a domestic market price of Rs 30-40 per unit (2 inch size).

II) Silver Moony fish, Monodactylus argenteus

Taxonomy Position

Class- Actinopterygii

Order- Perciformes

Family- Monodactlylidae

Genus- Monodactylus

Species- argenteus

The species is distributed in the Indo-Pacific region. It inhabits bays, mangrove estuaries,
tidal creeks. The fish feeds on plankton and detritus. It attains a maximum total length of 270 mm.
Males and female fish, TL 50-55 g size were observed to be in mature stage (Prem et al., 2014). At
CIBA broodstock of the fish is being maintained in ponds with commercial feeds and captive
maturation has been attained in the fish also. Further trails are in progress for captive breeding of the
species (CIBA ANNUAL REPORT, 2013-14). Silver Moony fish fetches over Rs 100 per unit in the
domestic market.

III) Green Chromide/Pearl Spots, Etroplus suratensis

Taxonomy Position

Class- Actinopterygii

Order- Perciformes

Family- Cichlidae

Genus- Etroplus

Species- suratensis
Training Manual on Seed Production and Farming of Brackishwater Finfishes I 80
 The species is naturally distributed in the southern India and Sri Lanka. It inhabits
freshwater and estuarine water bodies. The fish is an omnivore feeding on detritus, aquatic
macrophytes and filamentous algae. It attains a maximum total length of 400 mm. Length at
first maturity has been reported as 195 mm in males and 200 mm in females (Bindu et al.,
2014). Fecundity of pearlspot varies from 500 to 7550 (Vijayaraghavan et al., 1981; Bindu,
2006). CIBA has developed seed production of pearlspot different systems; ponds, tanks,
cages.

Pond based system: In ponds, area, 100 m2 and depth, 1.2 m, salinity, 15- 30 ppt, 50
brooders was stocked after systematic pond preparation. Additional spawning surfaces were
introduced in the pond for egg attachment. Feeding was done using formulated feeds. On
observing the presence of hatchlings, manuring was done with cow dung @500kg ha-1 for
enhancing of plankton production, artificial feed (25-30 g) was also provided. A production
of 3500 fry was observed in a year from 5 sets of breeding (Abraham and Sultana, 1995).

RCC tank system (20 t): In RCC tank, continuous water flow through was provided. Half
of the tank bottom is provided with a soil base for egg attachment earthen tiles and hide outs
were provided. The tanks are stocked using mature pearlspot brooders at a stocking density of
20 fish per tank at a male female ratio of 2:3. Pair formation and breeding occurs naturally.
Fry were collected at regular monthly intervals by lowering the water level. A production of
1200-3500 seed per batch was obtained regularly (CIBA Annual Report, 2010-11).

RAS based tank system (1 t): Breeding trials of pearlspot breeding were conducted in one
ton rectangular plastic tanks provided with a continuous water flow using a biofilter facility.
Each tank was provided with a small plastic tub filled with clay soil to facilitate breeding.
The breeding tank was stocked with 3-4 mature brooders. After breeding the eggs were
attached to the sides of the plastic container and the brood fish were observed to take turns in
defending the eggs. Larvae were separated and reared using alternate live feeds, Rotifer,
Brachionus plicatilis, Artemia nauplii and by co-feeding with commercial larval diets. One
of the most promising results obtained was the production of 8000 larvae by a single pair
stocked in six breeding at an average breeding interval of 17.6 ±1.12 days and an average
larval number per spawning of 1333.3±143.0.

Seed production of pearlspot in hapas: Seed production of pearlspot were conducted in

hapas set in ponds having gentle water flow, salinity- 25-30 ppt. Brooders were maintained in
small cages on commercial fish feed. Hapas (1x0.75x1 m) were fixed by casurina poles and

81 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

clay soil in small plastic tubs were suspended at 0.5 m depth to facilitate nest building. Just
above the soil surface 1-2 ceramic tiles are suspended to facilitate egg attachment. Each cage
was stocked with 3-4 brooders. Efforts at pair formation are usually observed a few hours
after release of fish in the cages. In the initial trials hatchlings were collected from pits in the
cage and subsequent larval rearing was practiced, following this method a seed production of
1000-1500 seed per hapa could be observed. However, it is not always possible to observe
larvae due to turbidity in the pond. In majority of seed production trials, seed were reared
within hapa with parental care. A production between 200-300 numbers of seed (avg. TL,
28.11±1.49 mm; Average Weight; 0.66±0.04 g) was observed per cage within 2- 2.5 months.

IV) Orange chromide, Etroplus maculatus

Taxonomy Position

Class- Actinopterygii

Order- Perciformes

Family- Cichlidae

Genus- Etroplus

Species- maculatus

The Orange chromide is a species of fish endemic to freshwater and brackish streams, lagoons
and estuaries in southern India (Maharashtra, Goa, Karnataka, Kerala and Tamil Nadu,) and Sri
Lanka. It is also known as pallathi in Malayalam and Pitalanga kachi in Tamil. This species is well
known for its parenting behaviour. The fish is an omnivore feeding on zooplankton and filamentous
algae. It attains a maximum total length of 80 mm. Fecundity of the fish is 1378 (Jayaprakash et al.,
1979) approximately and reported to be between 140- 231 eggs per spawning (Bindu and
Padmakumar, 2012). Size of the fishes varied from 6-10g in weight and 6-9 cm length. In nature the
fish form a breeding pairs and attach eggs on the substrate. The species exhibits parental care and the
offsprings are taken care of by the parents.

Training Manual on Seed Production and Farming of Brackishwater Finfishes I 82

 Attached Eggs and two days old larvae of Orange Chromide, E. maculatus

Two months old larvae of Orange

Chromide, E. maculatus

V) Crescent perch /Target Fish, Terapon jarbua

Taxonomy Position

Class- Actinopterygii

Order- Perciformes

Family- Teraponidae

Genus- Terapon

Species- jarbua

A new candidate species for brackishwater ornamental/food fish Terapon jarbua commonly
called as Target Fish, Crescent Bass, Crescent Perch or Tiger Bass. This species is euryhaline in
nature and can tolerate fresh to seawater salinity and could be a potential candidate species in brackish
and freshwater aquaculture for food and also ornamental purposes. Breeding and seed production is
one of the major constrain in aquaculture of this fish, hence a breeding trail was attempted for seed
production of this fish under captive condition.

Mature male and female was collected from brackishwater pond and acclimatized to captive
condition for a week under flow through system (Temp: 25-260C; Salinity: 27 ppt ). To assess the
maturity, female fishes were cannulated and male were gently pressed near the vent. Female having
the oocyte diameter above 460µ were selected along with the oozing male. Four breeding set was

83 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

arranged, and in each set 2-male and one female was introduced. From the four set two set were
administered with HCG@300 IU/Kg and other two set were with LHRHa @ 75µg/Kg, half the dose
was given to male. After 36 h of post injection all set was spawned (fertilized embryo size: 750 µ) and
embryo was semi-buoyant in nature, and after 16-18 h of incubation hatching was observed. Sizes of
hatchlings were 2 mm with the yolk sac length of 75µ and a single oil globule. After 12 h of post
hatching concentrated algae (Chlorella sp.) were introduced in the larval rearing tanks @ 20 lit/400 lit
and after 48 h rotifer feeding was started. A total of more than 3 lakhs spawns (2 days old) were
collected and transferred to nursery rearing protocol. CIBA has successfully initiated induced
breeding trials with hormonal manipulation using HCG and LHRH (CIBA ANNUAL REPORT,
2014-15).

1-Month Old Fries 2-Month Old Fries of 3-Month Old Fingerlings

of T.jarbua T.jarbua of T. jarbua

VI) Banded Chromide , Etroplus canarensis

Taxonomy Position

Class- Actinopterygii

Order- Perciformes

Family- Cichlidae

Genus- Etroplus

Species- canarensis

Canara pearlspot or Banded chromide, Etroplus canarensis, is a species of cichlid endemic to South
Karnataka in India (only from the Kumaradhara-Netravati river system). Its habitats are highly-
seasonal in nature with annual monsoons bringing about severe increases in water depth, flow-rate

Training Manual on Seed Production and Farming of Brackishwater Finfishes I 84

and turbidity. The substrate was mainly composed of small rocks and leaf litter with some tree roots
projecting into the water along one margin. Temperature: 22 – 32 °C; pH: 6.0 – 7.5, Hardness: 18 –
179 ppm. The Maximum standard length is 100 – 110 mm. Omnivorous in feeding habits, Sexual
maturity appears to be reached at around 2 years of age. In Rivers the fish are thought to breed during
the months of December and January, when temperatures are cooler and high monsoon waters have
receded, and simulation of the change between these seasons can sometimes induce captive spawning
process. This is an endangered species under IUCN lists.

Conclusion:

India's share in ornamental fish trade is estimated to be less than 1 % of the global trade. The
major part of the export trade is based on wild collection. There is very good domestic market too,
which is mainly based on domestically bred exotic species. The overall domestic trade in this field
cross 1000 lakh and is reportedly growing at the rate of 20 per cent annum. The earning potential of
this sector has hardly been understood and the same is not being exploited in a technology driven
manner. Considering the relatively simple techniques involved, this activity has the potential to create
substantial employment opportunities, besides earning foreign exchange also.

Reference

Abalika Ghosh, B. K. Mahapatra and N.C. Datta (2000) Ornamental fish farming- an additional
income generating programme for women with a note on its constraints and prospects. The 5th IFF,
Asian Fisheries Society, January 17- 20, CIFA (ICAR), Bhubaneswar.

Abalika Ghosh, B. K. Mahapatra and N.C. Datta (2003) Ornamental fish farming– successful small
scale aqua business in India. Aquaculture Asia, 8, (3): 14.

Alex Ploeg (2013) The status of ornamental aquatic industry. OFI journal 72:48-61.

Alex Ploeg; The Volume of the Ornamental Fish Trade OFI journal: 48-61.

Barry, T. P. and Fast, A.W. 1992. Biology of the Spotted scat (Scatophagus argus) in the Philippines.
Asian Fish. Sci., 5: 163-179.

Bindu L. and Padmakumar., K. G. (2008) Food of the Pearlspot, Etroplus suratensis (Bloch) in the
Vembanad Lake J. Mar. Biol. Ass. India, 50 (2): 156 - 160

Bindu L. and Padmakumar., K. G. (2014) Reproductive biology of Etroplus suratensis (Bloch) from
Vembanad wetland system of Kerala. IJMS; 43, 4: 646-654.

CIBA Annual Report, 2010-11. Central Institute of Brackishwater Aquaculture, Chennai. pp: 21-22.

CIBA Annual Report, 2013-14. Central Institute of Brackishwater Aquaculture, Chennai. p: 38.

85 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

CIBA Annual Report, 2014-15. Central Institute of Brackishwater Aquaculture, Chennai. pp: 76- 77.

Jayaprakas, V., K. G. Padmanabhan and N. K. Balasubramanian. 1979. Food, feeding habits and
breeding biology of the Orange chromide Etroplus maculatus (Bloch). Bulletin of the Department of
Aquatic Biology, University of Kerala, India, 4: 9-21.

Kailasam, M., Thirunavukkarasu, A.R. (2011) Mass-scale propagation of spotted scat. ICAR NEWS
17, 3, p: 4.

Moorhead, Jonathan A., and Zeng, Chaoshu (2010) Development of captive breeding techniques for
marine ornamental fish: a review. Reviews in Fisheries Science, 18(4). pp. 315-343.

Olivier, K. (2001). The ornamental fish market. FAO/Globefish Research Programme, Vol. 67.
United Nations Food and Agriculture Organisation, Rome, Italy.

Prathvi Rani, Sheela Immanuel, P. S. Ananthan, S. N. Ojha, Nalini Ranjan Kumar, Mohan Krishnan
(2013) Export performance of Indian ornamental fish - an analysis of growth, destination and
diversity. Indian J. Fish., 60(3): 81-86.

Prem Kumar, A. R. Thirunavukkarasu, M. Kailasam, J. K. Sundaray , G. Biswas , R. Subburaj, G.

Thiagarajan and S. Elangeswaran (2014) Captive maturation of the silver moony fish Monodactylus
argenteus (Linnaeus, 1758) under laboratory conditions. Indian J. Fish., 61(1): 113-117.

Sultana, M.; Krishnamurthy, K. N. and Pillai, S. N. 1995. Biology, fishery, culture and seed
production of the pearlspot Etroplus suratensis (Bloch). CIBA Bulletin No. 7. Central Institute of
Brackish Water Aquaculture, Madras. 43 pp.

Vijayaraghavan L., Krishnakumari, L., Gopinath, V.J., and Dhavan, R.M. (1981) Aquaculture of
pearlspot Etroplus suratensis in estuarine pond: environmental characteristics, primary production,
growth and cost benefit-ratio. Indian Journal of Marine Science, 10- 82-89.

Training Manual on Seed Production and Farming of Brackishwater Finfishes I 86

 Biology of Spotted Scat (Scatophagus argus) –
A potential Brackishwater ornamental fish
Babita, M. Kailasam, Aritra Bera, Tanveer Hussain

Spotted scat Scatophagus argus is distributed along indo pacific coastal waters and occurred
commonly in mangrove and backwaters regions. It is also known as Green Scat, Argus Fish,
Spade Fish, Butter Fish, and Tiger Scat etc. popularly. It is a deep and compressed body fish
with greenish – brown / silvery - golden on the sides with brown to reddish-brown spots.
Juveniles are yellow in colour with darker spots on the body. They are considered attractive
and calm fish for aquarium. Spotted scat is a long known fish for its beautiful appearance in
ornamental trade and considered a food fish in countries like Philippines, Thailand, and India
etc. It is a euryhaline fish and can easily inhabit in salinities of freshwater to marine water. It
is an herbivore fish and prefers to eat macroalgae in nature. It is a candidate fish to be
polyculture with other Brackishwater fin and shell fishes. Spotted Scat belongs to family of
Scatophagidae hence it possesses paired poison glands associated with each fin spine which
can inflict pain for several hours on careless handling.

Taxonomy Position

Class- Actinopterygii

Order- Perciformes

Family- Scatophagidae

Genus- Scatophagus

Species- argus

An adult Spotted Scat

Worldwide Distribution

Adult Spotted Scat is found in tropical and warm temperate waters of the Indo-west and
Central Pacific regions. Occurrence of adult fishes has been reported from Philippines,
Taiwan, Thailand, Indonesia, Australia, Japan, China, India, Bangladesh, Cambodia, Fiji,
French Polynesia, Hong Kong, Iran, Kuwait, Malaysia, Micronesia, Myanmar, New
Caledonia, Palau, Papua New Guinea, Samoa, Singapore, Sri Lanka, and Viet Nam etc.
Juveniles are usually collected from mangroves and coastal waters. Most fry are caught
during high tides.

87 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

Habitat and feeding habit

The Spotted scat is usually found in estuaries, harbours, mangrove, and the lower reaches of
fresh water streams. It feeds on benthic algae, plant matter, and small benthic invertebrates in
nature. In captivity they readily accept formulated feed also. Adults form small schools of 5 –
6 fishes. Individual grows to 20–30 cm.

Life history and Reproductive Biology

Adult fishes can grow upto 1 kg in nature. Male and female become sexually mature at 11.5
cm and 14 cm body size respectively. Maturity is attained by 1 + year age group fishes. Sexes
can be differentiated in spawning season easily with bulged abdomen of female and running
milt of male. The females are larger and heavier than the male during spawning season. Some
researchers have reported female dominance in population in wild. Adult fishes are spawning
after onset of first monsoon. It has been reported to spawn April to October with 2 peak
periodicity; one during March – May and second during August – November. Adult fishes
are believed to spawn in coastal waters. Juveniles are generally caught from river mouths and
mangrove regions.

Fecundity of female Spotted scat is 0.1 – 0.2 million which depends on body weight of fish.
Generally, 800 – 900 eggs/g ovary weight is obtained from a mature fish. Oocyte size > 450
µm indicates complete vitellogenesis and final maturation in female fishes. Stages of oocyte
development in female fish are as follows:

Primary oocytes: Diameter 0.04 to 0.10 mm, white - transparent in colour.

Immature oocytes: Diameter 0.10 to 0.35 mm, white - light yellow in colour, visible
germinal vesicle.

Maturing oocytes: Diameter 0.35 to 0.60 mm, yellow in colour, visible follicle layer (0.01
µm thickness)

Final maturation: Diameter 0.60 to 0.75 mm, colourless cytoplasm, Complete GVBD

Atresia: presence of broken cell membranes; freed, membrane-bound oil droplets.

Fertilized eggs are 0.65 – 0.7 mm in diameter with single yellow oil droplet in centre. Larval
development completes in 18 – 20 hours. Larvae pass through a tholichthys stage which is
deep-bodied and laterally compressed. They are dark, have rough, scale less skin, and a well-
developed lateral line. They range in size from 0.60 to 1.2 cm and lasts upto 30 – 35 dph.
Presences of bony plates are characteristics of this stage which later disappears and larvae
transform in juvenile form.

Spotted Scat as an ornamental fish:

Spotted Scat is a peaceful companion in aquarium. Its bold dark spots attract attention.
Adult fishes are shiny silver coloured with a light greenish cast and is covered with black
spots all across the body extending onto the fins. In aquaria they readily accept formulated
Training Manual on Seed Production and Farming of Brackishwater Finfishes I 88
feed. They are hardy in nature and can tolerate varying levels of salinity, pH, Dissolved
Oxygen. Demands for juveniles in domestic and international market are high. Juveniles can
be sold for ₹20 – 35/pc. Adult fishes >150g fetch premium price as food fishes. Its flesh is
soft and tastes good. In aquaria adults and juveniles can be kept in varying salinities due to
their euryhaline nature.

89 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

Culture of live food organisms: microalgae, rotifers, artemia and copepods
T. Sivaramakrishnan, K. P. Sandeep, Babita Mandal, R.Subburaj, G. Thiagarajan,
Babita Mandal, S.N.Sethi, M. Kailasam

Introduction
One of the major bottlenecks in the hatchery production of finfish is the larval rearing which
includes the transition from an endogenous to an exogenous feeding by the larvae. There are two
types of finfish larvae: precoicial and altricial. In precoicial larvae, when yolk sac is exhausted, they
look as mini adults, having fully developed fins and mature digestive system including functional
stomach. These fishes can ingest and digest formulated feeds as a first feed. eg: salmon and trout. In
case of altricial larvae, when yolk sac is exhausted(3-4 days in most of the tropical fishes), they
remain in relatively undeveloped state. The digestive system is rudimentary, lacking a stomach and
much of the digestion takes place in the hind gut epithelial cells. Such a digestive system seems to be
incapable of processing formulated diets. Moreover, the larvae are inefficient to catch and chase their
food due to underdeveloped vision and other sensory functions.Fishes at the time of their first feeding
are quite fragile and delicate creatures. It is the most critical phase of their life when they need right
type of nourishment for their survival and growth. If this requirement is not met, they perish. So it’s
necessary to give live feeds at this stage.
Mainly live feeds consist of phytoplankton and zooplankton grazed upon by economically
important fishes.They include different group of organisms like, microalgae, rotifers, artemia and
copepods. Importance of live feed is due to several factors as small size, rich essential nutrients, broad
spectrum composition of food, better intake due to the movement, auto-digestion characteristics,
facilitate better nutrient assimilation in larvae, stimulate feeding behaviour due to soft texture and
attractability and ample scope for enrichment. This is why live feed organisms are called living
capsules of nutrition. Adequate quantity of these live feed should be present during larval rearing
phase for the successful larval rearing. Hence, the mass cultures of these live feeds are required to
meet the need of the hatchery operation.

 Microalgae as live feed

Marine microalgae are the floating microscopic unicellular organisms which are generally free
living, pelagic in the size range of 2 to 20μ. They play pivotal role in aquaculture as a means of
enriching zooplankton for feeding fish and other larvae. In addition to protein and energy supply, they
provide other key nutrients such as vitamins, essential polyunsaturated fatty acids (PUFA), pigments
and sterols, which are transferred through the food chain. Therefore, microalgae are inevitable source
of nutrition for fish larvae. Microalgae varied in their proportions of protein (6.6-52%), carbohydrate
(5.5-23%) and lipid (7-23%).

Training Manual on Seed Production and Farming of Brackishwater Finfishes I 90

  Culture techniques
Microalgae can be produced using a wide variety of methods, ranging from closely-controlled
laboratory methods to less predictable methods in outdoor tanks. Various chemical media are used for
indoor and outdoor cultivation. (Guillard’s F/2 medium, Walne’s medium, Johnson’s mediumetc).
There are five different stages in the growth of every microalga (Fig 1). The best quality microalgae
can be harvested/utilized during the late exponential growth phase.

Fig: 1. Different phases in life cycle of microalgae

Indoor culture allows control over illumination (3000-500 lux), temperature (24 ± 10C),
nutrient level, contamination with predators and competing algae, whereas outdoor algal systems
make it difficult to grow specific algal cultures for extended periods. Open cultures such as uncovered
ponds and tanks (indoors or outdoors) are more readily contaminated than closed culture vessels such
as tubes, flasks, carboys, bags, etc. Axenic cultures are free of any foreign organisms such as bacteria
and require a strict sterilization of all glassware, culture media and vessels to avoid contamination.
Even then, using strict management measures and continuous monitoring it is possible produce mass
cultures of microalgae in open outdoor tanks / ponds for hatchery operations. Different types of
microalgae cultures are described in the following section.

 Batch culture
The batch culture consists of a single inoculation of cells into a container of fertilized seawater
followed by a growing period of several days and finally harvesting when the algal population reaches
its maximum or near-maximum density. In practice, algae are transferred to larger culture volumes
prior to reaching the stationary phase and the larger culture volumes are then brought to a maximum
density and harvested. The following consecutive stages might be utilized: test tubes, 2 liter flasks, 5
and 20 liter carboys, 160 liter cylinders, 500 liter indoor tanks, 5,000 liter to 25,000 liter outdoor tanks
(Fig 2).

91 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

 Fig: 2. Diagrammatic representation of batch culture of microalgae
(Source: FAO fisheries technical paper: 361)

 Continuous culture
The continuous culture method, i.e. a culture in which a supply of fertilized seawater is
continuously pumped into a growth chamber and the excess culture is simultaneously washed out,
permits the maintenance of cultures very close to the maximum growth rate. Two categories of
continuous cultures can be distinguished: Turbidostat culture, in which the algal concentration is kept
at a preset level by diluting the culture with fresh medium by means of an automatic system.
Chemostat culture, in which a flow of fresh medium is introduced into the culture at a steady,
predetermined rate. The latter adds a limiting vital nutrient (e.g. nitrate) at a fixed rate and in this way
the growth rate and not the cell density is kept constant.

 Semi-continuous culture
The semi-continuous technique prolongs the use of large tank cultures by partial periodic
harvesting followed immediately by topping up to the original volume and supplementing with
nutrients to achieve the original level of enrichment. Semi-continuous cultures may be indoors or
outdoors, but usually their duration is unpredictable. Competitors, predators and/or contaminants and
metabolites eventually build up, rendering the culture unsuitable for further use. Since the culture is
not harvested completely, the semi-continuous method yields more algae than the batch method for a
given tank size. Large outdoor ponds either with a natural bottom or lined with cement, polyethylene
or PVC sheets have been used successfully for algal production. The nutrient medium for outdoor
cultures is based on that used indoors, but agricultural-grade fertilizers are used instead of laboratory-
grade reagents

Training Manual on Seed Production and Farming of Brackishwater Finfishes I 92

 Microalgae production chain in finfish hatchery
A finfish hatchery must be equipped with a well maintained indoor microalgae laboratory to
keep pure cultures different strains used. The temperature, light intensity and aeration/CO2 supply
should be controlled inside the lab. Sterilized media are used in the lab. The culture volume ranges
from 10 ml to 20 litres in the lab. The inoculum for intermediate culture is provided from the indoor
laboratory. Generally, 20 litre carboy cultures are used as inoculum for intermediate culture. The
intermediate culture contains 500-1000 litre FRP tanks in a room with transparent roof and adequate
ventilation. The cultures from intermediate room are used in large outdoor tanks (5-20 tons).
Commonly used microalgae species in fish hatchery are: Chlorella salina, Chlorella vulgaris,
Nannochloropsis occulata, Isochrysis galbana, Tetraselmis sp, Thalassiosira pseudonana etc.

 Nutritional perspective of microalgae

The nutritional value of any algal species depends on its cell size, digestibility, production of
toxic compounds, and biochemical composition. Although there are marked differences in the
compositions of the micro-algal species, protein is always the major organic constituent, followed
usually by lipid and then by carbohydrate. Expressed as percentage of dry weight, the range for the
level of protein, lipid, and carbohydrate are 12-35%, 7.2-23%, and 4.6-23%, respectively. The content
of highly unsaturated fatty acids (HUFA), in particular eicosapentaenoic acid (20:5n-3, EPA),
arachidonic acid (20:4n-6, ARA), and docosahexaenoic acid (22:6n-3, DHA), is of major importance
in the evaluation of the nutritional composition of an algal species to be used as food for marine
organisms. Isochrysis galbana is rich in DHA, whereas Nannochloropsis occulata is rich in EPA.

Fig 3. Concentrations of protein, lipid and carbohydrate in some species of micro-algae commonly
used in aquaculture

93 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

 Commonly used media for indoor microalgae culture

 Fertilizer used for outdoor microalgae culture

(Ammonium sulphate: Urea: Super phosphate 10:1:1; grams per 100 litres of seawater)
Training Manual on Seed Production and Farming of Brackishwater Finfishes I 94
 Rotifers as live feeds
Rotifers are also known as wheel animalcules due to its appearance. They are microscopic
organism abundantly found in all the natural aquatic systems. It feeds on unicellular microalgae like
chlorella, Nannochlopsis, Tetraselmis, Isochrysis etc. by movements of corona. In commercial finfish
hatcheries rotifers are cultured on mass scale since they are the first feed of many of the finfish larvae
due to their small and preferred feeding size (100 – 280 µ). Due to its high reproductive ability, mass
culture can be achieved in few days. Rotifers are having ability to tolerate wide range of
environmental conditions without affecting its growth and reproduction. Different kind of rotifers are
being identified and classified based on their size, shapes of lorica and spines. Reproduction and
growth of rotifers can vary due to rearing conditions like temperature and salinity. Brachionus
plicatilis and Brachionus rotundiformis are the two species cultured in marine and brackishwater
finfish hatcheries.

 Types of rotifers:
Generally, three types of rotifers are cultured around the world in finfish commercial hatcheries
depending on the size requirements:

1. SS (Super Small) type: 100 – 140 µm

2. S (small) type: 141 – 220 µm
3. L (large) type : >220 µm
 Specific requirements to start the mass culture:

For starting rotifer mass culture, algal cell density should be >10×106 cells/ml for
Nannochloropsis sp or Chlorella sp. Algae culture should be 2 -5 times higher than the volume of
rotifer culture. Temperature should be in the range of 27 – 28° C. FRP tanks/ Concrete cement tanks
are suitable to start the culture.

 Pure culture of rotifer:

Rotifers are easily available in coastal areas where the water is abundant with nutrients. To start a
pure culture, 50 – 60 liter water can be sieved through 50 – 80 µ mesh size net. This filtered water
contains different species and strains of rotifers. Preferred species can be selected and isolated
individually under microscope. These isolated single rotifers can be put in culture tubes with algae
water for further reproduction under diffused light. After every 12 hour fresh algae should be supplied
to maintain the algal cell density. Gradually increase the volume to 25 ml, in 50 ml beakers. Change
the culture daily once. Use 50 - 80 µ mesh to separate the rotifers. Continue this procedure, till the
density reaches 50 individual / ml and the volume up to 500 ml. Increase algal cells density to 3-4

95 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

million cells/ml. When the density exceeds the above, remove half of the quantity and mix clean sea
water to make up the quantity.

 Mass culture of rotifer

Start the microalgae (Chlorella/ Nannochloropsis) culture in the rotifer culture tanks and when
the culture reaches the density of 20 × 106 cells/ml, inoculate pure culture of rotifer to achieve an
initial density of 10 individuals/ml. Allow the culture for 7-8 days to increase the rotifer density.
Harvest and concentrate the rotifers using 50 µ mesh plankton net. After each harvest, thoroughly
clean the tanks with fresh water. Culture of live feed should be scheduled to ensure daily harvest for
uninterrupted production. Better reproduction and nutritional quality can be achieved by regulating
feed, water, temperature, salinity and aeration during the culture process.

 Rotifer enrichment

The nutritional quality of rotifers depends on their food source. Highly unsaturated fatty acids
(HUFA) are essential for the survival and growth of the larvae. Rotifer feeds containing DHA and
EPA can be valuable for marine and brackishwater fish larvae. Depending upon their food source,
rotifers are composed of about 52-59% protein, up to 13% fat and 3.1% n3 HUFA. The high content
of the essential fatty acid eicosapentaenoic acid (EPA 20:5n-3) and docosahexaenoic acid (DHA
22:6n-3) in some microalgae (e.g. 20:5n-3 in Nannochloropsis occulata and 22:6n-3 in Isochrysis
galbana) have made them excellent live food diets for boosting the fatty acid content of the rotifers.
The harvested / concentrated rotifers can be kept in these microalgae cultures for few hours for
enrichment. The ratio of EPA:DHA can be manipulated by using different proportions of algae for
enrichment. There are plenty of commercial enrichment media are available in the market for rotifers.

 Artemia as live feed

Among the live diets used in the larviculture of finfish, nauplii of the brine shrimp Artemia
constitute the most widely used food item. Annually, over 2000 metric tons of dry Artemia cysts are
marketed worldwide for on-site hatching into 0.4 mm nauplii. Indeed, the unique property of the small
branchiopod crustacean Artemia to form dormant embryos, so called 'cysts', may account to a great
extent to the designation of a convenient, suitable, or excellent larval food source that it has been
credited with. Those cysts are available year round in large quantities along the shorelines of
hypersaline lakes, coastal lagoons and solar saltpans scattered over the five continents. After
harvesting and processing, cysts are made available in cans as storable 'on demand' live feed. Upon 24
hour incubation in seawater, these cysts release free-swimming nauplii that can directly be fed as a
nutritious live food source to the larvae of a variety of aquatic organisms, which makes them the most
convenient, least labour-intensive live food available for aquaculture. Approximately 90 % of the

Training Manual on Seed Production and Farming of Brackishwater Finfishes I 96

world’s commercial harvest of brine shrimp cyst comes from the Great Salt Lake in Utah. All the life
stages of Artemia, the decapsulated cyst, nauplii, juvenile, and sub adults are used as feed.

High densities of hatching from cysts can be achieved with transparent funnel shaped
containers (20 -30 liter, cylindro-conical FRP tanks) that are aerated from the bottom, which keeps all
cysts in suspension condition. Illumination in hatching tanks is provided by 60 watt fluorescent lamp
from a distance of 20 cm. Complete hatching takes place within 24 -36 hours. After complete
hatching, nauplii can be collected by attracting them near light source.

Salinity 30 - 35 ppt
pH 7.5 – 8.5
Temperature 27 – 30° C
Oxygen >2 ml/l
Illumination >1000 lux
Cyst density 1 gm/liter
Table 1: Conditions required for artemia cyst hatching

The hard shell and chorion can be removed by a technique, decapsulation, to achieve higher
hatching percentage. Here, the cysts are hydrated in water for 30 minute and then dipped in sodium
hypochlorite (NaOCl) solution of 200 ppm for a while. During this time care should be taken so that
temperature should not exceed beyond 30° C. When cyst becomes orange in colour, aeration should
be stopped and cysts are washed with Sodium thiosulphate solution for less than a minute to remove
the residual chlorine. Cysts are to be rinsed with water and can be kept for incubation in hatching tank
with strong aeration in filtered seawater with minimum illumination of 2000 lux for 18-24 hours.
After complete hatching, the aeration can be stopped and all other light sources are to be turned off
and a light should be provided near bottom outlet for nauplii collection. After 10-15 minutes, nauplii
can be collected with the help of fine mesh net.

 Copepods as live feed

Most of the commercial finfish species are reared using rotifers and Artemia nauplii since
they can be cultured in large quantities at high densities. Unfortunately, using rotifers and Artemia
during this early period in life history does not always promote optimal larval growth since these live
preys may contain an inadequate fatty acid profile and, in some cases, be of an inappropriate size.
Obtaining enough copepods of desired stages at a specific time has been one of the barriers for
extensive use in aquaculture and experimental work with fish larvae or other copepod-feeding
organisms. Therefore, establishment of reliable production methods for copepods that can meet the
quantitative requirements of larval fishes is essential.

97 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

 Copepods are nutritionally superior to all other live feeds commonly used in marine and
Brackishwater aquaculture. This is the one of the main reasons by which copepod culture has got
immense importance in finfish larval nutrition. A number of beneficial effects have been linked to
copepod nutrient composition in relation to early larval nutrition. In particular, emphasis has been put
on lipid composition, and the content and ratio of the polyunsaturated fatty acids (PUFA)
docosahexaenoic acid (DHA), eicosapentaenoic acid (EPA), and arachidonic acid (ARA)

Fig 4. Fatty acid composition of various live feeds (Source: Meeren et al., 2008)

In contrast to rotifers, copepods are more difficult to culture on a commercial basis. Only a
few species of copepods have been mass cultured successfully. Many temperate copepods produce
resting eggs as a common life-cycle strategy to survive adverse environmental conditions, which is
analogous to Artemia and Brachionus sp. These characteristics can be made useful in aquaculture.

 Conclusion

Since the live feed are inevitable for larval rearing of many species of finfish, the research on
new vistas in live feed nutrition is the need of the hour. Apart from commonly used live feeds like
microalgae, artemia and rotifers there are many more live feed organisms with high nutritional profile
are yet to explore. The availability of on-grown live food would not only offer farmers and exporters a
better alternative option for feeding their fish, but more importantly, the possibility of enhancing the
fish performance and quality through bio-encapsulation. There are several potential live feed
organisms which are to be addressed for mass culture techniques. The researches in the area of live
feed nutrition will enhance the successful and effective larval rearing of many marine and
brackishwater finfishes.

 For further reading

 Algal culturing techniques, 2005.Edited by Andersen R A., Elsevier Academic Press.
 Manual on the Production and Use of Live Food for Aquaculture, FAO FISHERIES TECHNICAL
PAPER361.
 Live Feeds in Marine Aquaculture, 2003.Edited byJosianne G. Støttrup and Lesley A. McEvoy.
Blackwell Science Ltd.
 Handbook of Microalgal Culture, 2004. Edited by Amos Richmond. Blackwell Science Ltd.

Training Manual on Seed Production and Farming of Brackishwater Finfishes I 98

 Health management in brackishwater finfishes
M. Makesh, Krishna Sukumaran, Tanveer Hussain, M.Kailasam

Health management in fish hatcheries and grow-out culture is essential for the successful production
of fish seeds and table size fish. The quality and quantity of seeds produced is a yardstick to judge the
success of a hatchery and the quality of seeds produced depends on health management measures
adopted by the hatchery. In a hatchery, larvae are often stocked in high densities and hence poor
management will result in weak seeds with low survivability. Prevention is considered as the first step
in controlling infectious diseases in hatcheries. Simple measures like sanitizing hands before handling
fish, having foot dips with disinfectants at all entry points in the hatchery, disinfecting the source
water etc. can help a lot in controlling infections. The following health management techniques need
to be adhered to in hatcheries to maintain and produce healthy disease free seeds.

1. Quarantine: All brooders brought to the hatchery need to be quarantined till it is tested and
found to be free of any diseases or parasites. The quarantine tanks should ideally be placed
away from the hatchery operations. Brooders brought to the hatchery should be acclimatized
in the quarantine tanks and clinically examined for the presence of ectoparasites, lesions,
fouling of gills etc. Samples should also be collected and tested for the presence of subclinical
infections. Once the stock is found to be healthy and free of infections they may be
transferred to broodstock tanks.

2. Biosecurity: Biosecurity is critical for the successful operation of the hatchery. Strict
biosecurity measures need to be adopted to prevent the possible entry of pathogens into the
hatchery. The following measures need to be adopted in a fish hatchery.
a. Ideally the hatchery should be located away from general public and the entry to the
hatchery should be restricted to authorised personnel only.
b. Visitors should be asked to follow the guidelines strictly while visiting the hatchery.
c. All vehicles entering the hatchery should pass through a disinfectant pit so that the
entire tire of the vehicle comes in contact with the disinfectant before the vehicle
enters the hatchery.
d. The source water should be filtered with sand filters to prevent entry of weed fishes
which may carry pathogens. The source water should also be disinfected to get rid of
pathogens by UV treatment or ozonisation.
e. Each building /culture area must be provided with foot dips containing potassium
permanganate.
f. The hatchery personnel need to sanitize their hands before any hatchery operation.

99 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

 g. Bird proofing, rodent and stray animal control measures should be adopted.

3. Use separate tools like hand nets, beakers and siphoning tubes for different batches of larvae
and ideally use separate tools for each tank. Disinfect the tools periodically and at least once a
day.

4. Avoid stocking broodstock and larvae of different species in the same premises. If
unavoidable, keep them separated as far as possible.

5. Disinfect the entire hatchery premises between seasons.

6. Clean and disinfect the tanks with bleach between batches and allow them to dry completely.

7. Use pathogen free live feed for the larvae. The live feed sample may be tested periodically to
know the presence of pathogens.

8. Aeration to the tanks should be stopped briefly in the morning allowing debris and dead
larvae to settle down. Siphon out the tank bottom to remove debris and dead larvae. Count the
dead larvae as this gives an indication of the problem if there is any.

9. Check water quality parameters like dissolve oxygen, ammonia, nitrate, nitrite, temperature
etc. daily. Maintain optimum water quality.

10. Excess feeding of the larvae should be avoided as it will deteriorate the water quality resulting
in reduced survival.

11. If more dead larvae are found than the usual numbers, send some live larvae to the laboratory
to isolate and identify the pathogens, if any, present. Tanks suspected of having an infection
should be treated with disinfectants and drained immediately to prevent the spread of
infection. All tools used for the infected tank, including the tank should be thoroughly
scrubbed, disinfected and allowed to dry before using them again.

12. If case of high mortality, the cause of the mortality should be identified at the earliest. Larval
and water samples should be sent to laboratory for testing. Further the source of the infection
needs to be ascertained to prevent further infections. For this all input materials like feed and
water should be tested.

Training Manual on Seed Production and Farming of Brackishwater Finfishes I 100

 13. Use of probiotics in larval rearing water is reported to be beneficial and is said to increase
survivability and reduce mortality. Hence suitable probiotics may be used at the
recommended dose.

14. The total bacterial load and vibrio load in the inlet water should be monitored regularly. If any
increase in the load is observed, suitable measures should be taken to disinfect the water.

15. Higher stocking densities of larvae lead to stress and bacterial infections. Hence optimal
stocking density should be maintained.

16. All dead fish and larvae needs to be disinfected before disposal.

17. Use aseptic techniques while collecting biopsy, hormone injection, implantation in broodfish
to avoid any accidental introduction of pathogens. Disinfect the catheters, needles etc. used in
sampling and injecting before each use.

18. Mullets are very sensitive to handling. Hence handle broodfish carefully to avoid stress and
injury. Use anaesthetics like, phenoxy ethanol, clove oil, MS-222 to anaesthetize the fish
before handing and injecting.

19. Periodically examine the brooders for the presence of any ectoparasites. If any parasites are
observed treat the fish with appropriate chemicals like formalin, dichlorvas to remove the
parasites.

20. Vaccinate the brooders for diseases which are endemic, if vaccines are available.

21. Observe the fish regularly for symptoms of stress/disease.

22. Educate/train all hatchery personnel on the importance of biosecurity and the do’s and don’ts
of the hatchery.

23. Record keeping is very much essential in hatchery operations. All activities of the hatchery
should be recorded regularly.

The following are the important diseases reported in brackishwater finfish.

101 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

Bacterial diseases

Aeromonad septicemia

Aeromonad septicaemias are caused by Aeromonas hydrophila and to certain extend by A. caviae.
The bacteria are ubiquitous and are commonly found in soil and water samples. It is also found in the
gut and tissues of healthy freshwater and marine fishes. A. hydrophila are Gram negative, motile rods
measuring 0.3-1.0 x 1.0-3.5 µm in size. The bacteria are not fastidious and can be easily isolated in
nutrient agar. The colonies appear within 24 h and are white, circular and convex. Selective media
such as Rimler-Shotts agar containing novobiocin and aeromonas isolation media containing
ampicillin can be used to isolate the bacteria from samples containing other bacteria. The disease
occurs when the fishes are stressed due to over-crowding, poor water quality, handling, high water
temperature etc. Affected fish exhibit darkening of the skin, haemorrhages on the skin and the base of
fins. Shallow ulcers may develop at haemorrhagic sites. Congestion and haemorrhages can also be
observed in the internal organs. The affected fishes may be treated with antibiotics like oxytetracyline
and sulphonamides. Improving the water quality with frequent water exchange and removal of dead
and moribund fish will reduce mortality.

Vibriosis

Vibriosis is caused by Vibrio anguillarum, V. alginolyticus and V. parahemolyticus. Members of the

family vibrionaceae are Gram negative, straight or curved rods. They are commonly found in marine
and brackishwater environment. The causative agents can be isolated in media containing seawater
salts and common nutrient media containing 1-2% sodium chloride. Thiosulphate citrate bile salt
sucrose (TCBS) agar is a selective medium for isolation of pathogenic vibrio species. Colonies appear
smooth, convex and white and develop within 48 hours. Vibrio species can be differentiated form
Aeromonas by their sensitivity to vibriostat 0/129. The bacteria are present in the alimentary tract of
normal healthy fishes. The incubation period varies with temperature and virulence of the strain.

The disease occurs in late summer when water temperature is high. Mortality may reach 50% in
young fish. The affected fish are anorectic with darkening of the skin. In acute infections deep
necrotic skin ulcerations with blood coloured exudate is observed. Spleenomegaly is a common
feature with petechial haemorrhage on most of the internal organs. In chronic cases skin ulcers may
become granulomatous. Gills become pale and corneal opacity is frequent.

Although commercial vaccines are available in developed countries, they are not available in India.
Antibiotic therapy with oxytetracycline and sulphonamides is the practical method of reducing
mortality during outbreaks.

Training Manual on Seed Production and Farming of Brackishwater Finfishes I 102

Viral diseases

Viral nervous necrosis

Viral nervous necrosis (VNN) is one of the important diseases of brackishwater fishes affecting a
wide range of fishes. Although mortality due to VNN is not high in mullets and milkfish, it can cause
considerable loss. The causative agent of the disease, viral nervous necrosis virus (VNNV), a
betanodavirus has four genotypes viz, barfin flounder nervous necrosis virus (BFNNV), redspotted
grouper necrosis virus (RGNNV), striped jack nervous necrosis virus (SJNNV) and tiger puffer
necrosis virus (TPNNV). The disease affects early larval and juvenile stages only. The virus is
transmitted both horizontally and vertically. The virus also produces persistent infection especially in
the adults resulting in asymptomatic carriers which act as a source of infection to larval and juvenile
stages. Vaccination of juveniles and young adults appear promising in protecting the fish. Vaccination
of brooders provides protection to larval stages through maternal transfer of immunity.

Red Sea bream iridoviral disease

Red sea bream iridoviral disease caused by red sea bream iridovirus (RSIV) has been reported in more
than 30 cultured species including grey mullets. All age groups are susceptible to the virus although
juveniles are more susceptible than adults. The virus affects spleen, kidney, heart, intestine and gills.
The transmission of the virus is through horizontal routes. The disease is more prevalent in summer
when water temperature is above 25°C. The control measures include introducing disease free fish,
implementing strict biosecurity measures, and maintaining good water quality, reducing activities
that induce stress, such as overcrowding and overfeeding.

Lymphocystis virus disease

Lymhocystis virus disease (LCD) is caused by Lymphocystis virus disease virus (LCDV) belonging
to the genus Lymphocystivirus of the family Iridoviridae. The disease produces nodular skin lesions
in a variety of fresh, brackish and marine water fishes. The nodules appear small, cream-coloured on
the skin and fins externally and on the mesentries and peritoneum internally. The disease is
transmitted horizontally when the lymphocysts break, through the skin abrasion caused by handling or
parasites. The disease is not lethal and do not cause mortalities unless vital organs are affected.
However it results in economic loss due to the poor marketability of the affected fish. The disease is
diagnosed by the presence of wart like growth on the skin, gills and fin. No specific treatment is
available. However the virus is sensitive to potassium permanganate, formalin and sodium
hypochlorite. The warts slough off or regress if the fish are maintained well with good water quality
and free of stressors like parasites. Affected fish must be isolated to prevent spread of infection.

103 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

Parasitic diseases

Argulosis

Argulosis is caused by a large ectoparasite, Argulus, commonly called as fish lice. It is the most
important branchiurans parasite belonging to the family Argulidae. These parasites are dorsoventrally
flattended measuring up to 1 cm in length. They are commonly found in the skin and fins of
freshwater fishes and to a lesser extent in brackish water fishes. The trauma induced by the parasite
due to the attachment and feeding method causes haemorrhagic ulcers and leads to secondary
bacterial infection. Affected fish show lethargy, irritation and loss of appetite. It is practically difficult
to eradicate argulus in culture waters as the adults and larval stages are active swimmers. Infested fish
can be treated with formalin or organophosphorus insecticides. Drying the ponds and tanks between
cycles will reduce argulus infestation.

Marine Ich

Marine Ich, caused by Cryptocaryon irritans is one of the common salt-water parasitic ciliate
infestations. Some of the common signs of marine Ich are rubbing on the pond side or bottom,
increased mucus secretion, breathing problems, loss of appetite, abnormal swimming behaviour,
frayed fins, cloudy eyes, and white spots especially on the dorsal side. The incidence is high with poor
water quality and over-crowding. The parasite infests almost all marine and brackishwater teleosts.
Fishes should be quarantined and only those free of any parasites have to be taken to the hatchery.
Marine Ich can be diagnosed by microscopic examination of skin and gill scrapings. C. irritans can be
observed as 0.3-0.5 mm structures with multi-lobed nucleus. The parasite can be kept away by
maintaining good water quality. Infected fish can be treated with formalin @ 100 ppm for 1 hr for 3
days or copper sulphate @ 0.5 ppm for 7 days or by immersing the fish in freshwater for one hour
daily for three days.

Amyloodiniosis

Amyloodiniosis caused by Amyloodinium ocellatum, a dinoflagellate is one of the most frequently

encountered pathogens affecting tropical marine ornamental fishes. Amyloodiniosis is also called as
‘marine velvet’. The symptoms of amyloodiniosis are difficulty in breathing, sluggishness, pale gills,
excess mucus secretion, rubbing its surface against objects in the aquarium and anaemia. The parasite
initially infects the gill and subsequently spreads throughout the body giving a velvety appearance and
thus the name marine velvet. The parasite may invade the tissues also. Affected fish appear dark in
colour and emaciated. Amyloodiniosis can be easily diagnosed by microscopic examination of gill
and skin scrapings. The condition can be treated with copper sulphate @ 0.5 ppm for 4-5 days or bath
treatment with formalin @ 200 ppm for 1 hour. Good water quality is also advocated.

Training Manual on Seed Production and Farming of Brackishwater Finfishes I 104

Monogenean infestation

Monogenans are ectoparasites that infest skin, gill and fins and are commonly known as gill or skin
flukes. The common monogenans encountered in brackishwater fishes are Dactylogyrus,
Gyrodactylus, Diplectanum and Benedenia. During heavy infections these parasites can cause high
mortality in fry and fingerlings. High stocking density combined with poor water quality will result in
high incidence of monogenean infestation. Clinical signs include lethargy, high mucus production,
rubbing of body against substrate, abnormal swimming behaviour and anorexia. Diagnosis is simple
by observing the parasites from gill and skin scarping under microscope. Treatment includes formalin
dip @ 100 ppm or dichlorvos @ 1 ppm for 1 hour.

Copepod infestation

Copepods are crustacean parasites having free living and parasitic stages. The important copepods
infecting cultured brackishwater fishes are caligus spp (sea lice), Ergasilus Spp. (gill maggots) and
Lernaea spp. (Anchor worm). Parasites are introduced into the culture system through water, live
feed, wild fish and contaminated tools and equipment. Poor water quality and overcrowding leads to
heavy infestation with copepods. Caligus can cause serious damage if present in large numbers. The
damage is caused by pre-adult and adult stages which abrade the skin surface and feed on cutaneous
and subcutaneous tissues. The parasite is introduced into farmed stock though introduction of wild
fish. Heavy infestation with copepods results in mechanical damage, impaired respiration, petechiae,
anaemia and emaciation. These parasites also act as act as mechanical vector for other bacterial and
viral pathogens. Diagnosis can be done by simple microscopic examination of the gills and skin.
Adult anchor worms are visible to the naked eye. Copepod infestation can be controlled using fresh
water bath for 15 min. or by using hydrogen peroxide @ 1000 ppm or formalin @ 100-200 ppm for
60 minutes. Complete draining, disinfecting and drying of the tank periodically help to break the life
cycle of the parasite.

References:

1. Jitthendran K P and Sujeet Kumar. 2013. Diseases in brackishwater aquaculture. The marine products
export development authority, Kochi.
2. Roberts R J (Ed). 2012. Fish Pathology, Fourth edition. Wiley-Blackwell
3. N. Pirarat, T. Katagiri, M. Maita, T. Nakai, M. Endo. Viral encephalopathy and retinopathy in
hatchery-reared juvenile thread-sail filefish. Aquaculture 288 (2009) 349–352.
4. E. Gomez-Casado, A. Estepa, J.M. Coll. Comparative review on European-farmed finfish RNA
viruses and their vaccines. Vaccine 29 (2011) 2657–2671.
5. S Gibson-Kueh, G H Ngoh-Lim, P Netto, J Kurita, K Nakajima and M L Ng. A systemic iridoviral
disease in mullet, Mugil cephalus L., and tiger grouper, Epinephelus fuscoguttatus Forsskal: a first
report and study. Journal of Fish Diseases 2004, 27, 693–699

105 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

 Development of formulated feed for cultivable brackishwater finfishes

K. Ambasankar, K. P. Kumaraguru Vasagam, J. Syama Dayal, and K.P. Sandeep

Introduction

Aquaculture farming has shown phenomenal growth in the last decade in India
producing protein rich health food and earning valuable foreign exchange. Feed is a major input in
fish farming. The development of nutritionally balanced feed involves understanding the dietary
requirements of candidate species, selection of feed ingredients, formulation of feeds and appropriate
processing technology for producing water stable pellet feeds. Depending upon the type of farming, a
wide range of feeds is used for feeding stocked shrimp and fish. While no feed is used in traditional
farming systems, supplementary and balanced feeds are used in extensive and semi-intensive
aquaculture.

All animals including fish requires food to supply the energy that they need for movement
and all the other activities that they engage in for growth. However, they are ‘cold-blooded’ and as
their body temperature is the same as the water they live in, they do not therefore need to consume
energy to maintain a steady body temperature and they tend to be more efficient users of food than
other farm animals. The nutrient requirement of different species of finfish vary in quantity and
quality according to the nature of the animal, its feeding habits, size, its environment and reproductive
state.

Fish diet should have adequate energy, not only to meet the needs of body maintenance called
basal metabolism, but also for growth. In nature shrimp and fish feeds on a variety of food items and
derive their balanced nutrition for healthy growth. When they are cultured in confined pond they
should be provided with a balanced diet as close to natural food as possible. This is the reason for
understanding the nutritional requirement of candidate species which assumes paramount importance
in developing the feeds for the candidate species.

Nutritional requirement of different brackishwater species

Protein

Protein is the most important nutrient in the diet of shrimp and fish. Protein requirement of
aquatic organism is higher than terrestrial animals. Fish require food protein in the form of essential
amino acids for maintenance of life, growth and reproduction and the requirement of protein depends
on animal characteristics i.e., species, physiological stages, size as well as dietary characteristics, i.e.,
protein quality (digestibility and biological value), energy level etc.. Scarcity of carbohydrate and

Training Manual on Seed Production and Farming of Brackishwater Finfishes I 106

abundance of protein and lipid in the natural aquatic food web is also probably responsible for the
common trend of aquatic organisms to use protein as an energy source.

Protein is required in the diet to provide indispensable amino acids and nitrogen for synthesis
of non-indispensable amino acids. A deficiency of indispensable amino acid creates poor utilization of
dietary protein and hence growth retardation, poor live weight gain and feed efficiency. In severe
cases, deficiency reduces the ability to resist diseases and lowers the effectiveness of the immune
response mechanism. Experiments have shown that tryptophan deficient fish become scoliotic,
showing curvature of the spine, and methionine deficiency produces lens cataracts.

Protein requirement vary with the age of the fish. Younger animal generally require higher
levels of protein (5-10% more protein) than older animals. Carnivores require high dietary protein
(40-50%) than omnivores (25-35%). Among the brackishwater finfishes, requirement of protein for
Asian seabass (Lates calcarifer), milkfish (Chanos chanos) and mullet (Mugil cephalus) is 40-45%,
40% and 27-35%, respectively.

Amino acids

The growth of fish is directly related to the quality of protein in terms of amino acids. After
digestion of protein, amino acids are metabolized at tissue level to form new proteins for growth,
maintenance and energy. Among 25 amino acids present in protein 10 amino acids must be supplied
in the diet since fish cannot synthesize them and termed as essential amino acids (EAA). These are
arginine, histidine, isoleucine, leucine, lysine, methionine, phenylalanine, threonine, tryptophan and
valine. A large proportion of the amino acid consumed by a fish are catabolized for energy and fish
are well adapted to using an excess energy in this way. It is found that if the amino acid composition
of the protein in the feed matches with the amino acid composition of shrimp body tissue, such feed
promotes good growth.

Table 1. Nutrient requirements of different brackishwater fishes

Nutrient L. calcarifer Mugil cephalus/ Etroplus suratensis

Liza tade

Energy (Kcal/kg) 4000-4500 4000-4500 4000-4500

Protein % 45-55 27-35 30-32

Lipid % 6-18 6-9 6-8

Carbohydrate % 10-20 30-40 30-40

107 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

Lipid

Lipid is a complex mixture of simple fat, phospholipids, steroids, fatty acids and other fat
soluble substances such as pigments, vitamins A, D, E and K. Apart from its major role to supply
energy lipid also act as precursors to many reactive substances. Phospholipids are responsible for the
structure of cell membranes (lipid bi-layer). Fatty acids are the main active components of dietary
lipids. Deficiency of essential fatty acid result in general reduction of growth and a number of
deficiency signs including depigmentation, fin erosion, cardiac myopathy, fatty infiltration of liver
and ‘shock syndrome’ (loss of consciousness for a few seconds following an acute stress. Fat levels of
6-8% are adequate in most of the fish diets. However, the quality of fat in terms of fatty acids is more
important. Carnivorous fish such as seabass can utilize lipids more effectively and lipid level as high
as 20 % can be used in their diet. However, lipid level should be adjusted in diet considering the
technological problems in feed manufacture and storage. Fish oil and soya oil are generally used as
lipid source during feed formulation.

Fatty acids

Fish and shrimps are unable to synthesize fatty acids of the n-3 and n-6 series and must be
provided in their diets. Aquatic animals require higher n-3 fatty acids than terrestrial animals. Among
aquatic animals, marine habitat requires more HUFA than freshwater counterparts. Among the long
chain fatty acids polyunsaturated fatty acids (PUFA) such as linoleic acid (18:2n6), linolenic acid
(18:3n3), eicosapentaenoic acid (20:5n3) (EPA) and docosahexaenoic acid (22:6n3) (DHA) are
essential for growth, survival and good feed conversion ratio. The n3 fatty acids are more essential
than the n6 acids. The fatty acids, EPA and DHA, which are known as highly unsaturated fatty acids
(HUFA) of n3 series, are particularly important. Quantitatively EPA and DHA are needed at 0.5% and
1.0% in the diet of larvae and fry of brackishwater fish. Fresh water fish show requirement for n6 and
n3 essential fatty acids (EFA), whereas marine fish show requirement of n3 and also HUFA.

Phospholipids

Fish require phospholipids for growth, metamorphosis and maturation. Lipids of squid, clam,
shrimp, fish and polychaetes are excellent natural source of phospholipids. The phospholipid
phosphatidylcholine (lecithin) is essentially required in the diet of larval and fry stages of fish for fast
growth and good survival. Soya lecithin is a good source of phospholipid. It is required at 1-2% level
in the diet. The development and survival of larvae is significantly improved when the diet contains
lecithin..

Training Manual on Seed Production and Farming of Brackishwater Finfishes I 108

Carbohydrate

Carbohydrate is an inexpensive source of energy in fish diet. Among the different types of
carbohydrates available, fish are found to utilize disaccharide and polysaccharide better than
monosaccharide. Omnivorous fishes have enzymes to digest carbohydrates while carnivorous fishes
have poor ability to digest carbohydrate. Polysaccharides are better utilized than monosaccharide.
Generally carbohydrate utilization by fish is found to be lower than that of terrestrial animals. Fish
can utilize dietary carbohydrate up to 40%. For carnivorous fish carbohydrate level in the diet may be
in the range of 10-20 %. Depending upon the total energy content required in the diet, carbohydrate
can be used from 10-40% level. Using starch as source of carbohydrate in diet has dual advantage.
Besides being energy source, it can act as binder if gelatinized by cooking with moisture and hence
improve water stability of diet. Corn flour, wheat flour, tapioca flour and other grain flours are good
source of starch in shrimp and fish diet. Another polysaccharide, cellulose is required in the diet as
roughage for improving the feed efficiency in fish.

Mineral requirement

Micronutrient such as vitamins and minerals significantly influence the growth and survival
of fish and these cannot be synthesized by these organisms.

Fish can absorb minerals directly from aquatic environment through gills and body surfaces
or by drinking. Hence, dietary requirement of minerals is largely dependent on the mineral
concentration of the aquatic environment. About 20 inorganic elements (macro and micro) are
required to meet the metabolic and structural functions in the body of animals. The aquatic organisms
regulate the mineral needs through dietary source and also through internal regulatory mechanisms in
the kidneys and gills. In saline water, calcium (Ca) is abundant, which is absorbed by most aquatic
animals. Since the availability of phosphorus (P) through water medium is poor, P should be made
available through diet. Usually the preferred Ca:P ratio is 1:1 in feeds of aquatic species. Mono and
dicalcium phosphate contain more available P than tricalcium phosphate. Incorporation of P should be
very discrete in fish feeds, as most of it gets excreted leading to eutrophication. The dietary
requirement of P ranges from 0.5-0.9% in fishes. The requirement of magnesium (Mg) in fish ranges
between 0.04-0.3%. The requirement of zinc (Zn) ranges from 15-30 mg/kg diet for fishes. The
requirement of iron (Fe) ranges from 150-200 mg/kg diet for fishes. Major

deficiency symptoms of manganese (Mn) in fishes are cataracts and abnormal curvature of the
backbone and malformation of tail. A dietary supplementation of 11-13 mg/kg restores normal growth
in fishes.

109 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

 Trace minerals like copper (Cu), cobalt (Co), selenium (Se), iodine (I) and chromium (Cr)
have some role in general upkeep of the organism. Their dietary incorporation enhances growth and
survival.

Vitamin requirement

Micronutrient such as vitamins and mineral significantly influence the growth and survival of
fish and this cannot be synthesized by these organisms. Even though, some vitamin such as niacin can
be synthesized by number of animals but are typically insufficient to meet physiological demand.
Hence, supplementation of vitamins in feed becomes necessary for most of the aquatic organisms.
Unlike domestic higher animals, the recommended doses of vitamin for aquatic animals are higher, as
many vitamins are lost during the process of feed manufacture and also due to leaching. Destruction
of vitamin-C due to oxidation is one of the biggest problem during feed manufacture. Many fishes
cannot synthesize Vitamin- C from glucose due to absence of enzyme L-gulonolactone oxidase.
Major role of vit C is in the formation and maintenance of intracellular material having collagen or
related basal constituents in bones and in soft tissues. Among the 11 water soluble vitamins, three (vit
C, inositol and choline) are required in large quantities. Sources of choline include cottonseed meal,
fish meal, shrimp meal, soyabean meal and yeast. Stable form of vit C is available commercially.

Table 3. Vitamin requirements of fish

Vitamin (mg/kg) Fish

Thiamin 10
Riboflavin 20
Pyridoxine 10
Pantothenic acid 40
Niacin 150
Folic acid 5
Vit B12 0.1
Choline 3000
Inositol 400
Vit C 100
Vit E 30
Vit A (IU) 2500
Vit D (IU) 2400
Vit K 10

Training Manual on Seed Production and Farming of Brackishwater Finfishes I 110

 Genetic factors in broodstock management in finfish hatchery

Rekha M.U., Krishna Sukumaran and M. Makesh

Introduction

With global population expansion, the demand for high-quality protein, especially
from aquatic sources, is rising dramatically. Increased aquaculture production is clearly
needed to meet this demand in the third millennium, because capture fisheries are at capacity
or showing precipitous declines due to overfishing, habitat destruction and pollution. Further
increases in capture fisheries are not anticipated under the current global conditions. In order
to meet the requirement, the aquaculture production has to be increased. Increased
aquaculture production mainly depends on the availability of quality fish seeds. The finfish
hatcheries produce larval and juvenile fish for transfer to aquaculture facilities where they are
grown to reach harvest size. Hatchery production confers three main benefits such as out of
season production, platform to carry out genetic improvement programmes and reduced
dependence on wild-caught juveniles. The production capacity of the hatchery depends on the
number and efficiency of the broodstock maintained. The characters of the broodstock like
growth rate, feed conversion efficiency are transmitted to their progeny and we expect the
progeny to be superior to their parents. To achieve that, efficient and sustainable genetic
management programmes should be followed by the hatcheries.

Quantitative genetic traits

Usually the finfish hatcheries aim at improving the quantitative traits such as growth
rate, feed conversion, oxygen tolerance, percentage of body fat, meat production, disease
resistance, and stress resistance. These traits are said to be quantitative because they typically
vary among individuals within a population. They have a range of expression and are
controlled by minor genes. These traits are measured using a continuous distribution system
and statistics. Such traits are described and reported around their central tendencies, such as
average (mean), variance, standard deviation and range. These traits are also influenced by
the environment. Gene expression levels, the environment, and the interaction of the two can
play a significant role in the variation of quantitative traits.

Genetics of small population

Hatcheries usually rear their own broodstock and do not recruit from natural water
bodies or exchange breeders between farms. Each hatchery therefore can be considered as an

111 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

isolated, self-sustaining and genetically closed unit (Eknath and Doyle, 1990). It is now
established that in genetically closed hatchery systems, potential selective pressures exerted
on finite and often small culture populations by various farm management practices such as
selection of founder stock, number of breeders maintained, method of replenishing
broodstock, stocking density, feeding regime etc. can result in indirect or negative selection,
inbreeding and genetic drift (Doyle, 1983). Retarded growth, reduction in reproductive
performance, morphological deformities, increased incidence of disease and mortality of
hatchery produced seeds of carps have been reported.
There has been an unconscious negative selection in some hatcheries as the relatively
bigger (hence fast growing) individuals from the grow-out ponds are sold and the remaining
smaller (hence the slow growing) fish are used for broodstock replacement. This practice is
also followed by nurseries in selling fast-growing; hence the good fingerlings and keeping the
smaller unsold fingerlings to raise as the broodstock. This will lead to negative selection and
then to poor growth and survival.

Inbreeding depression

There is every possibility of inbreeding in hatcheries as the male and female are
chosen from a finite (small) population for mating, with a greater chance of crossing sib
(brother-sister) or closely related fish. Moav and Wohlfarth (1976) stated that a single full sib
mating of a particular fish might result in 10-20% depression in growth and considerable
proportion of individuals might show physiological abnormalities. Because of generations of
inbreeding and accumulation of unfavourable alleles from closed mating, genetic
deterioration of existing stock might make them less suitable for culture. An inbred or
homozygous population normally loses its general vigour. This result from lack of knowledge
of broodstock management practices, especially about the need for recruitment of new
breeders in to the stock at regular intervals, maintenance of proper stocking density of the
brood fish of desirable size, injecting adequate hypophysation dosage, mating unrelated male
and female breeders, basic disease control, water quality maintenance, and record keeping of
broodstock and spawning. Inbreeding is not harmful always. Linebreeding is the form of
inbreeding that is used to increase an outstanding animal’s contribution to a population.

Genetic drift

Genetic drift is the random changes in the gene frequency that occur because of
sampling error. Sampling error can be natural or manmade. Natural sampling errors are those

Training Manual on Seed Production and Farming of Brackishwater Finfishes I 112

that occur when earthquakes, floods, landslides, or other natural disasters subdivide a
population and isolate small groups of organisms. Manmade sampling errors are inaccurate
collections, sampling only fish that possess a certain phenotype or that spawn on a day, etc.
When culturing a fish, the important change occurs in gene frequency as a result of genetic
drift occurs during the creation of the next generation or during the acquisition of the
population. This is when the genes are transferred from parents to offspring or when they are
transferred from one hatchery to another.
The loss of alleles via genetic drift has two effects, first it increases homozygosity;
consequently, it has an effect similar to that seen for inbreeding. The simultaneous effect of
increase in inbreeding and loss of alleles via genetic drift as result of a decrease in effective
breeding number (Ne) can cause severe genetic problems. Secondly, the loss of alleles
reduces the genetic variance. Genetic material is the raw material up on which the selection
works. If there is no genetic variance, there will be no heritable differences, which mean the
selection cannot improve a phenotype. Equally important, if a population is being cultured for
stocking lakes and rivers, the loss of genetic variance may doom the project to failure.
Natural populations need broad gene pools i.e., they need as much genetic variance as
possible. Populations with narrow genetic bases are likely to survive in the long term. Genetic
drift has been shown to have damaged the gene pool of several hatchery populations

Measures to preserve and enhance genetic variability

a. Maintaining the effective population size (Ne)

Managing effective population size (Ne) is the most important aspect in fish husbandry.
Genetic diversity of the broodstock is a major factor for the fitness of seed (Primack, 1993).
Genetic variability decreases rapidly if the Ne of the brood stock is reduced. The effective
population size is the number of individuals that would give rise to the rate of inbreeding
appropriate to the conditions under consideration, if they bred in the manner of the idealised
population. The genetic stability of a closed hatchery population depends on effective
breeding size or population size (Ne). Both the inbreeding and genetic drift are inversely
related to Ne.
The rate of inbreeding can be given by the formula

113 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

Where, F is the rate of inbreeding, Ne is the effective population size
The relationship between Ne and genetic drift is

Where,σ 2 Δq is the variance in the change of gene frequency, and p and q are the frequencies
of alleles p and q for a given gene.
The variance of the change in the gene frequency is the way genetic drift measured. There is
no universal Ne that can be used to manage every population. It must be customized for every
population. When managing a population’s Ne, the major goal is to maintain Ne at a constant
level for every generation. If Ne drops below the desired value for a single generation, the
genetic goals cannot be achieved. Maintaining Ne at the desired level generation after
generation may be the most difficult aspect of broodstock management, because the Ne can
decline for a variety of reasons. Sudden and drastic decreases in Ne are called bottlenecks,
and they can cause permanent and irreversible genetic damage. The ultimate effect of small
Ne is the loss of alleles via genetic drift. Rare alleles will be lost more easily, but common
alleles can also be lost. The loss of genetic variance can produce irreversible damage to the
population’s gene pool. Parameters of genetic variability such as allele frequency, average
number of alleles per locus and average heterozygosity reveal that genetic variation decreases
in the founder stock which represents the first generation of artificially propagated captive
broodstock collected from wild population. Use of large number of brooder fish is
recommended for minimizing genetic erosion.

b. Minimizing negative selection

A hatchery should have short term and long term plans to avoid the risk of negative
selection. The following practices are recommended to minimize indirect or negative
selection in hatcheries
 The base population should be collected from natural waters or from a known source.
 Record of locations of collection, date of collection/transfer, species, size and weight
of the stock, number of individuals at the time of stocking in nursery/rearing ponds,
etc. should be kept.
 The fast growing and best individuals from nursery/ grow out ponds should be
selected for raising as broodstock and few individuals from as many stocks as
possible for each species should be selected.
Training Manual on Seed Production and Farming of Brackishwater Finfishes I 114
  The brood fish should be marked or tagged for record keeping.
 Hatchery produced seed of different selected stocks should be stocked separately or in
a pool. All necessary records, like number of breeders used in each slot with their tag
numbers, date of spawning, date of stocking and number individuals stocked in each
nursery/ rearing pond should be maintained.
c. Avoiding stock deterioration due to inbreeding

Stock deterioration due to inbreeding can be avoided by

 Keeping an adequate number of brood fish in order to select the best performers in
terms of size, maturation and breeding efficiency.
 Maintaining pedigree records to reduce or avoid the chance of mating between closely
related breeders.
 Exchanging broodstock among the hatcheries to minimize inbreeding
 Adopting a well-planned selective breeding and line crossing program to improve
desirable traits in the founder stocks.

DNA- marker assisted broodstock management

Because genetic factors determine the fitness and adaptability of the organisms,
preservation of genetic capital of broodstock is important in the production of high quality
seed for aquaculture and sea ranching. Even if founder specimens of brood stock are
genetically intact, lack of a proper management strategy can lead to inbreeding and rapid
decline in diversity at gene level. Deletion of wild alleles and erosion of genetic variability
are known to adversely affect inherent strength and fitness of the stock. Seeds originating
from genetically degraded broodstock have poor chances of survival in the wild. They have a
reduced ability to respond to changing conditions of the environment (Waples et al., 1990).
Further, those that survive to maturity are likely to contaminate the gene pool of the
natural populations through reproductive interaction, resulting in propagation of genotypes
that are not as much adaptable as those of the wild populations. Production of seed fish for
stock enhancement programs should be based on well-organised broodstock management
strategies. Unknown and known genetic changes and possible loss of genetic variation in
broodstock and progeny should be monitored. DNA markers are good monitoring tool for
broodstock management. The information generated by these markers can be utilized in
maintaining genetic variability and effective population size in broodstock and their progeny
(Perez- Enriquez et al., 2001)

115 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

Conclusion

The inbreeding and genetic drift are inevitable in small hatchery populations and
together result in loss of heterozygosity which leads to loss or fixation of alleles and finally
leaves the population homozygous for a particular allele. We can get rid of these unwanted
effects by maintaining effective population size, avoiding mating of closely related
individuals and by proper record keeping.

Training Manual on Seed Production and Farming of Brackishwater Finfishes I 116

 Hormonal control of reproduction in fish

Dani Thomas, M.U.Rekha, Krishna Sukumaran, M. Kailasam, Sherly Tomy, Aritra Bera

Introduction

Reproduction in fishes is regulated by external environmental factors that trigger internal

mechanisms into action. The final event of the reproductive cycle, the release of eggs and sperm
resulting in spawning, can be controlled by either placing the fish in an appropriate environment or by
changing the fish’s internal regulating factors with injected hormones or other substances. The
internal mechanism that controls the process of reproduction in fish is the brain-hypothalamus-
pitutary-gonad axis. The external environmental factors that control reproduction, however, vary
considerably among species. For this reason, more is known about the internal regulatory mechanism
of fish reproduction than the specific environmental requirements for spawning each species.

Environmental factors

Environmental factors that regulate maturation and spawning in fishes are: photoperiod, water
temperature, water quality, water current, tides and cycles of moon, weather cycles, nutrition, diseases
and parasites.

Brain–Hypophyseal–Gonadal Axis

Environmental signals, in conjunction with social cues, are conveyed by sensory signals to
various brain centers culminating in the hypothalamus. In the absence of hypophyseal portal system,
hypothalamic nerve fibers branch throughout the adenohypophysis to regulate the synthesis and
release of the gonadotropic hormones, follicle-stimulating hormone (FSH) and luteinizing hormone
(LH). Dopamine secreted by the hypothalamus may inhibit LH release. FSH and LH, in turn, bind to
their respective receptors in the gonads (FSH-R and LH-R, respectively). The gonads respond by
secretion of sex steroid hormones; estradiol 17 β (E2) in females that promotes oogonial proliferation
and vitellogenesis, and progestogens such as 17 α, 20 β dihydroxy-4-pregnen-3-one (DHP) that
promotes initiation of germ cell meiosis and follicular maturation and ovulation (Figure 1(a)). In
males, these are androgens, mainly 11 ketotestosterone (11-KT), that regulate spermatogenesis and
spermiogenesis, as well as DHP that initiates the meiotic division of spermatogonia and controls the
spermatozoa maturation and spermiation. DHP in its free or conjugated forms serve as pheromones as
well. Generally, fertilization occurs in the water, either in specialized nests guarded by one or both
parents or in the open water.

117 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

 Figure 2: Schematic representation of the reproductive axis in fish

Endocrinology of fish reproduction

Gametogenesis (spermatogenesis and vitellogenesis) and final maturation (spermiation and

OM) are regulated by a cascade of hormones along the brain–pituitary–gonad (BPG) axis. In this axis,
the secretion of the pituitary gonadotropins FSH and LH is controlled by the brain via the stimulatory
action of the GnRHs, which are the primary neuropeptides regulating reproduction, acting as
integrators of external information (example, environment, temperature, water fall and social
interactions). Dopamine (DA) in some fishes exerts a negative effect on the functions of GnRH on the
pituitary gonadotrophs. The FSH and LH are released into the bloodstream to act on the gonad, where
they stimulate the synthesis of the sex steroid hormones (androgens, estrogens and progestogens),
which are the ultimate effectors of gonadal development. Testicular spermatogenesis, as well as
spermiation, is regulated by pituitary FSH and LH secretion through the action of the sex steroid

Training Manual on Seed Production and Farming of Brackishwater Finfishes I 118

hormones, as well as other growth factors. Before the onset of spermatogenesis, spermatogonial stem
cell renewal seems to be regulated by E2 acting on Sertoli cells. The androgen 11-keto testosterone
(11KT) is the major regulator of spermatogenesis, while the maturation inducing steroid (MIS)
regulates sperm capaciation and spermiation. Both steroids are synthesized by the somatic Leydig
cells of the testes, after GtH stimulation. The LH is mainly involved in the stimulation of androgen
production in Leydig cells, whereas FSH seems to exert more complex functions in the male testes,
stimulating androgen production from the Leydig cells, as well, but also regulating Sertoli cell activity
during spermatogenesis. The onset of spermatogenesis is a process controlled by the secretion of
pituitary GtHs (mainly FSH). The FSH acts on Sertoli cells and stimulates 11KT biosynthesis, which
in turn regulates the full process of spermatogenesis, mediated also by growth factors (example,
insulin-like growth factor I, IGF-I or activin B) secreted by the Sertoli cells. In males, FSH levels are
high at early spermatogenesis; on the other hand, LH is low during early spermatogenesis. In males,
androgen production (T and 11KT) remains high through the entire spawning period, even while MIS
levels are high, since spermatogenesis, spermiogenesis and spermiation occur concurrently. In
females, a predominant role has been suggested for FSH during vitellogenesis in fishes with
synchronous ovarian development. On the other hand, in fish with asynchronous ovarian development
the role of FSH in vitellogenesis is less clear and a possible function has been ascribed also to LH.
The control of reproduction with the use LH has been worked in bakishwater fishes.

Figure 3: Regulation of LH secretion. Schematic presentation of brain and gonadal regulation of LH secretion

At the conclusion of vitellogenesis, OM is triggered by the action of LH on the follicle cells,

which synthesize and secrete the maturation inducing hormone (MIH) or maturation inducing steroid
(MIS). In some other marine species, a derivative of 17,20bP the 17a, 20b, 21- trihydroxy-4-pregnen-
3-one (20bS) has been described to act as MIS. Both 17,20bP and 20bS are acting as MIH in
European sea bass, striped bass and red seabream. The MIS binds to specific receptors on the oocyte
plasma membrane and the signal received in the oocyte surface is transduced to the cytoplasm to

119 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

finally result in the formation and activation of the maturation-promoting factor (MPF), which is
responsible for the resumption of meiosis and completion of oocyte maturation

Reproductive dysfunction in cultured

Reproduction is regulated by the brain via the release of GnRH from the hypothalamus and it
stimulates the release of gonadotropin (GtH) from the pituitary. Dopamine provides a negative control
of pituitary GtH release, mostly freshwater species. Fish possess two or three different variants of
GnRH, and 14 variants have been so far identified from various vertebrates. Pituitary control of
reproduction is via a dual GtH system, with follicle stimulating hormone (FSH or GtH I) regulating
vitellogenesis and spermatogenesis, and luteinizing hormone (LH or GtH II) regulating FOM and
spermiation. In female, most cultured species exhibit some degree of reproductive dysfunction when
reared in captivity. Problems are more widespread in female broodstock and can vary from
inconsistent spawning only, to the complete failure of oogenesis. The failure of captive fish to
undergo FOM in captivity in some fish was found to be the absence of LH release during the
spawning season. Fish that exhibit this type of dysfunction undergo normal vitellogenesis, but with
the onset of the spawning season the developing oocytes fail to initiate FOM; instead they undergo
atresia. Treatment of such broodstock with exogenous GtH or GnRHa at the completion of
vitellogenesis stimulates gonadal steroidogenesis FOM and ovulation. The final and most severe form
of reproductive dysfunction of captive female broodstock is the failure to undergo vitellogenesis.
During vitellogenesis, FSH or LH stimulate the production of testosterone (T) by the theca cells and
its aromatization to 17β -estradiol (E2) in the granulose. In response to stimulation by E2 the liver
produces vitellogenin, which is sequestered by the oocytes in a receptor-mediated process enhanced
by FSH. At the completion of vitellogenesis a surge in plasma LH stimulates a drop in plasma E2, a
transient increase in plasma T during GV migration, and a dramatic elevation in the plasma levels of
the maturation inducing steroid (MIS), which acts at the level of the oocyte membrane to induce
FOM. In male gonadotropins regulate spermatogenesis via the production of androgens by the testes,
mainly 11- ketotestosterone, since T is the precursor of 11-KT, the levels of the two androgens covary
during most of the reproductive season. Plasma 11-KT levels peak during spermiogenesis and decline
just prior to, or during the spermiation period.

Hormones for induced spawning

Hormone-induced spawning techniques influence this sequential mechanism at several levels,

by either promoting or inhibiting the process. The primary substances used for hormone-induced
spawning have been: pituitary extracts and purified gonadotropins to stimulate the ovaries and testes;
LHRH analogs (LHRHa) alone or in combination with dopamine blockers which enhance the potency
of LHRHa to stimulate the pituitary; or steroids to stimulate the gametes directly. The appropriate

Training Manual on Seed Production and Farming of Brackishwater Finfishes I 120

hormone preparation should be selected on the basis of the species to be spawned and the availability
of the hormones. Many variables impact the ability of injected hormones to induce spawning,
including: 1) condition of the fish; 2) stage of sexual maturity; 3) size of the fish; 4) previous
spawning history, 5) water temperature; and 6) season of the year. Pituitary extract The pituitary gland
produces and stores gonadotropin hormones (GtH), which play a decisive role in ovulation and
spermiation. Injected pituitary material bypasses the brain-pituitary link, acting directly on the ovaries
and testes, providing the surge in blood GtH levels that normally precedes spawning just prior to
spawning. This is a problem when adult fish are scarce. Fresh pituitary glands should be used
immediately or preserved by either freezing or acetone-drying. Glands can simply be placed in a
sterile vial or plastic bag and stored in a freezer until needed. To acetone-dry, the glands are
immediately placed in a vial with acetone. After collecting the required number, the acetone in which
the glands were placed is drained off and replaced with fresh acetone. The acetone is again changed 8
to 12 hours later. After 24 hours in acetone, the glands are air dried on a paper towel. The dried
pituitaries are then stored in a sealed.

Commercial pituitary extracts: Common carp pituitary or salmon pituitary extracts, available
commercially, are widely used for induced spawning. These are crude acetone-dried powdered whole
pituitaries. As with fresh pituitaries, these preparations also contain the pituitary tissue and hormones
unrelated to reproduction, in addition to GtH. In general, the closer the donor species is related to the
recipient fish, the greater the chance of successful induced spawning. Therefore, carp, goldfish,
Chinese carps, catfish, etc., are more likely to spawn successfully when injected with pituitary
extracts from carp. Salmon, trout, etc., are more likely to spawn successfully when salmon pituitary is
used. However, both are effective on a wide variety of fish species. There is always uncertainty about
the hormone potency of pituitary material. Hormone content necessary for spawning is greatest in
sexually mature fish just prior to spawning and lowest in immature fish and mature fish after
spawning. The potency of pituitary material can also be destroyed by improper collection, processing,
or storage.

Purified gonadotropin: To better quantify the hormone injected, purified gonadotropin hormones are
frequently used. Human Chorionic Gonadotropin (HCG) is the most common purified gonadotropin
hormone used for induced spawning. In fish, the injected gonadotropin mimics the natural GtH
produced by the fish’s pituitary. Just as is the case with pituitary extracts, purified hormones such as
HCG bypass the brain-pituitary link, acting directly on the ovaries and testes. HCG has been used to
spawn fish such as striped bass, white bass, red drum, catfish, and mullet. HCG + pituitary extract
HCG, however, is not effective on all species. HCG has been used in combination with common carp
pituitary extract; for some species, the combination has shown to have improved potency than either
preparation used alone. The two hormones can be prepared and injected separately, or the HCG
solution can be used when mixing the pituitary extract.

121 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

Luteinizing hormone-releasing hormones: Injections of mammalian Luteinizing Hormone-
Releasing Hormone (LHRH) have been used experimentally to mimic the fish’s GnRH. However, a
comparatively large dose and frequent injections were required. Recently, synthetic LHRH analogs,
referred to as LHRHa or GnRHa, have been manufactured. These hormones last longer in the fish’s
system and have potent stimulator effects on ovulation and spermiation in fishes. Therefore, only one
or two small doses are needed to induce spawning. LHRHa stimulates the fish’s own pituitary to
produce and release the GtH necessary for spawning. LHRHa has been used to induce ovulation in a
wide range of fishes. One of the synthetic analogs that has been used successfully is Des-GLY10 ,
[DAla6 ]-LH-RH Ethylamide.

LHRHa + dopamine blockers: Although LHRHa has not been shown to be species specific, some
fish do not respond to injections of LHRHa alone (e.g., goldfish, redtailed black shark, rainbow
shark). Dopamine inhibits the release of hormones from the pituitary, effectively blocking the
pituitary’s positive response to injected LHRHa. There is a family of drugs that act as dopamine
blockers, either by preventing the release or by inhibiting the binding of dopamine. Experimental
results indicate that the use of dopamine blockers prevents this negative feedback, enhancing the
effectiveness of LHRHa for these species. Because of the tremendous variety of aquarium species and
their individual spawning requirements, as compared to food and sport fish, development of hatchery
spawning technology has been more difficult. Many ornamental species have had to be imported from
wild populations. The use of LHRHa with dopamine blockers has helped change this situation.
Haloperidol {4-[4-(4-chlorophenyl)- 4-hydroxy-piperidino] -4’- fluorobutyrophenone} has been used
recently as a dopamine blocker in ornamental fishes and tested experimentally for food and sport fish
production.

Steroids: Several steroids (e.g., progesterone and testosterone) have been tried experimentally for
inducing maturation, ovulation and spermiation in fishes. However, there appears to be little
indication of widespread importance of these substances for hormone-induced spawning.

Conclusions

Reproduction in fishes is regulated by both internal mechanisms within the fish and external
environmental factors. The environmental factors trigger the internal mechanisms into action. The
internal mechanism that controls the process of reproduction in fish is the brain-hypothalamus-
pituitarygonad chain. Hormone-induced spawning techniques influence this sequential mechanism at
several levels, by either promoting or inhibiting the process. The primary substances used for
hormone-induced spawning are: (1) pituitary extracts and purified gonadotropin to stimulate the
ovaries and testes; or (2) LHRH analogs (LHRHa) alone or in combination with dopamine blockers
which enhance the potency of LHRHa to stimulate the pituitary.

Training Manual on Seed Production and Farming of Brackishwater Finfishes I 122

References

Aizen J, Kasuto H, Golan M, et al. (2007) Tilapia follicle-stimulating hormone (FSH):

Immunochemistry, stimulation by gonadotropinreleasing hormone, and effect of biologically active
recombinant FSH on steroid secretion. Biology of Reproduction 76: 692–700.

Devlin RH and Nagahama Y (2002) Sex determination and sex differentiation in fish: An overview of
genetic, physiological, and environmental influences. Aquaculture 208: 191–364.

Ge E (2005) Intrafollicular paracrine communication in the zebrafish ovary: The state of the art of an
emerging model for the study of vertebrate folliculogenesis. Molecular and Cellular Endocrinology
237: 1–10.

Yaron Z and Levavi-Sivan B (2006) Reproduction. In: Evans DH and Claibourne JB (eds.) The
Physiology of Fishes, 3rd edn., pp.343–386. Boca Raton, FL: CRC Press/Taylor and Francis.

Zhou L-Y, Wang D-S, Kobayashi T, et al. (2007) A novel type of P450c17 lacking the lyase activity
responsible for C21-steroid biosynthesis in the fish ovary and head kidney. Endocrinology 148: 4282–
4291.

123 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

 Synthetic hormones and their role in brackishwater finfish breeding

Aritra Bera, Krishna Sukumaran, Babita Mandal, Dani Thomas, M.Kailasam

Introduction

Reproduction is the biological process resulting in the production of new individual.

With the introduction of several species and intensification of aquaculture, control of
reproduction process of fish in captivity to produce high quality seed is a major factor
determining the success of the aquaculture. Unfortunately majority of the cultured teleost
species exhibit some degree of reproductive dysfunction when reared in captivity. An
understanding of the reproductive process in teleost fish is important for the industry to
modify the timing of puberty and maturation which could aid in steady supply of fish off-
season and also aid in genetic manipulations. The fish reproductive cycle are controlled by
the reproductive hormones of the brain, pituitary and gonad. The nervous and the endocrine
system works together to control reproduction through the hypothalamo-pituitary-gonadal
(HPG) axis. The main hormones involved are the hypothalamic decapeptide gonadotropin-
releasing hormone (GnRH), gonadotropins (GTHs) from the pituitary and the sex steroids
from the gonads. In addition to steroids, several factors such as stress and nutrition are
involved in the control of the HPG axis to modulate the release of gonadotropin release.

Hormonal therapies

Although the growth phase of reproductive development is concluded in captivity in most

fishes-the major exemption being the freshwater eel (Anguilla spp.), oocyte maturation (OM)
and ovulation in females and spermiation in males may require exogenous hormonal
therapies. In some fishes, these hormonal manipulations are used only as a management tool
to enhance the efficiency of egg production and facilitate hatchery operations, but in others
exogenous hormones are the only way to produce fertilized eggs reliably. The findings that
heterologous pituitary homogenates could induce females to ovulate lead to the development
of the hypophysation technique which was used extensively in aquaculture species. The
exogenous LH preparations act directly at the level of the gonads to induce FOM and
ovulation. However, the technique was later replaced by methods employing either human
chorionic gonadotropin (hCG) extracted from the urine of pregnant women, or piscine
pituitary extracts and purified LH obtained through chromatographic separation. Of the

Training Manual on Seed Production and Farming of Brackishwater Finfishes I 124

gonadotropin preparations, hCG often in combination with GnRHa, has been tested with
variable success in many commercially important fish. The major drawbacks of all GTH
preparations is their high cost and the fact that fish may become refractory to similar
treatment in subsequent spawning seasons.

Dopamine, a catecholamine neurotransmitters, exerts inhibitory effect on some fish.

Secretion of dopamine from nerve terminals in the pituitary and its binding to D2 receptors
localized on gonadotrophs results in inhibition of basal and GnRH-stimulated release of LH.
Treatment with a DA antagonist causes an increase in the numbers of LH-like gonadotrophs
and is directly proportional to time and the dose of the antagonist. The inhibitory effect of
DA on LH secretion changes over the course of the reproductive cycle, with the maximum
DA inhibition occurring during the final stages of gametogenesis. This feature is utilised in
aquaculture of Cyprinidae by using dopamine antagonists in ovulation-inducing therapies,
e.g., domperidon, pimozide, reserpin, metoclopramide, haloperidol, isofloxythepin .

The use of synthetic GnRH and its hyperactive agonists for maturation induction
commonly employed, trigger the secretion of the fish’s own GTH, thus activating its
pituitary-gonad axis. GnRH being a small decapeptide apparently does not trigger an immune
response and acts at a higher level in the BPG axis, thus providing a more balanced
stimulation of reproductive events. GnRHa is synthesized chemically and does not carry the
risk of transmitting diseases to the broodstock, a danger always associated with the use of
pituitary extracts. The development and utilization of synthetic GnRHa in fish culture has
greatly increased the level of sophistication and control of hatchery production and
contributed substantially to the growth and diversification of the aquaculture industry.
However, injection of GnRHa does not always result in 100% ovulation and often multiple
injections are often necessary to induce ovulation. Thus there was a need for the development
of a hormonal formulation that does not require repeated applications in the development of
spawning induction therapies which lead to the development of controlled-release delivery
systems for GnRHa which has proven to be an important broodstock management tool, and
have contributed to the species diversification of the aquaculture industry in the last decade.
The hormones implants mixed with cholesterol, ethylene-vinyl in biodegradable
microspheres have been efficient in inducing maturation and spawning in many cultured fish.

Kisspeptins are involved in the onset of puberty by the activation of the HPA axis in
fish. Exogenous kisspeptin injection either centrally or peripherally causes stimulation of the

125 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

HPG axis in many mammalian as well as in teleosts. Cirrhinus mrigala was induced using
synthetic Kissppetin-10 hormone alone and in combination with domperidone. Central as
well as peripheral administration of KP robustly increases systemic levels of the LH and FSH
in sexually immature and mature rodents
A major concern with regard to hormonal therapies is their effect on gamete quality,
compared to naturally maturing or spawning broodfish. The main factors that may have
significant consequences on gamete quality-mainly on eggs-and should be considered when
choosing a spawning induction procedure include (a) the developmental stage of the gonads
at the time the hormonal therapy is applied, (b) the type of hormonal therapy, (c) the possible
stress induced by the manipulation necessary for the hormone administration and (d) in the
case of artificial insemination, the latency period between hormonal stimulation and stripping
for in vitro fertilization.

Commercially used synthetic compounds for induced breeding

1. Gonadotropins:

Three types of gonadotropins are generally used for fish breeding. Mammalian
luteinizing hormone (LH); Human chorionic gonadotropin (hCG); Pregnant mare serum
gonadotropin (PMSG). HCG has different brand names like Prolan, Antuitrin, Sumaach
and Synahorin which contains mammalian pituitary extract also.

2. Synthetic Gonadotropin relaeasing hormone:

Scientists were successful in synthesizing analogues of GnRH substituting amino acids at
position 6,7 or 10 to make GnRH or LHRH analogues (GnRH-a/ LHRH-a).
Commercially available forms are- Ovaprim containing Salmon GnRH-a (sGnRH-a) and
domperidone (Syndel Laboratory), Ovatide containing GnRH-a and pimozide
(Hemmopharma), Ovapel available in pellet form containing mGnRH-a and
metoclopramide (University of Gödöllo), WOVA-FH containing GnRH-a (Biostat
Agrisciences, Wockhardt).

3. Steroid Hormones:
11- deoxycorticosterone acetate (DOCA), 17 alpha, 20 beta di-hydroxy progesterone
(17α,20β-DHP), 17α Methyl Testosterone (17-MT) has been extensively used for fish
breeding.

Training Manual on Seed Production and Farming of Brackishwater Finfishes I 126

4. Other drugs:
Prostaglandin and Antiestrogens like Tamoxifen has been used to stimulate natural
spawning and inducing ovulation in fishes in captivity.

Induced breeding of captive reared brackishwater fishes at ICAR-CIBA

Understanding of sex-steroid pathway in wild fish and manipulating it in captivity

through hormonal manipulation is essential to achieve gonadal recrudescence. Carnivorous
and herbivorous fishes differ significantly in reproductive physiology particularly in the area
of final oocyte maturation (FOM) and the age of first maturity. Brood fishes at CIBA are
ideally maintained in 100 ton RCC tanks or earthen ponds with flow through system having
salinity > 29 ppt. Carnivorous finfishes like sea bass (Lates calcarifer), grouper (Epinephelus
tauvina) and cobia (Rachycentron canadum) achieves final oocyte maturity and spawning
only under hormonal manipulation (neuro-peptides and sex-steroids). Gonads of carnivore
fishes have characteristic feature of spontaneous maturity based on brood nutrition with trash
fishes (@ 5% body weight) and spawning takes place only after final LHRH-a (75 mg/kg)
/HCG (250 IU/kg) injection when oocyte reaches 450 µm and 650 µm in sea bass and cobia,
respectively. On the other hand, herbivorous fishes need sustained release of GnRH
throughout its reproductive cycle to mature and spawn. Milkfish (Chanos chanos) and grey
mullet (Mugil cephalus) were given intra muscular implantation of LHRH-a (50 mg/kg) and
17 α –Methyl Testosterone (50 mg/kg) at monthly interval to mature. Milkfish and mullet
spontaneously spawn when oocyte diameter reaches 850 µm and 600 µm respectively after
several intramuscular hormone implantations. Mystus gulio, a high valued fish locally known
as “Nuna Tengra” was induced to attain oocyte maturation under captive condition through
dietary manipulation and LHRHa hormone pellet implantation. Fish were observed to spawn
after 4 – 6h of injection. Breeding trials in the gold spot mullet, Liza parsia, a species with
high consumer demand in West Bengal and Orissa, showed that oozing males and females
with developing ovary following treatment with LHRHa. Broodstock feed containing high
protein and lipid along with other vital micronutrients plays a major role in achieving
maturity. Proper monitoring of brood-stock health, nutrition and water quality with controlled
manipulation of environment (salinity and temperature) is most important factors in marine
herbivores.

127 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

Conclusion

Among the significant advancements in the field of aquaculture is the development of

techniques to induce reproduction in fish. The multiplicity of neuroendocrine signalling
pathways in teleosts is probably due to the gene duplication event but several evidences have
suggested their unique roles and functional significance in a variety of reproductive strategies
in teleosts. Numerous hormones have been used to induce reproduction of a majority of
economically important fishes. These techniques have allowed farmers to profitably breed
and raise species that do not naturally reproduce in captivity, and to manipulate the timing of
reproduction to suit production cycles. Compared to multiple injections, sustained-release
GnRHa-delivery systems reduce the necessary handling to a minimum. Although major
advances have been done over the last years in our understanding of these central
mechanisms, there still are a number of unresolved issues that represent true bottlenecks for
the development of sustainable aquaculture. Further research aimed at understanding the
reasons for reproductive dysfunction should contribute to future progress in the area of
artificially stimulation of oocyte and maturation and ovulation of fish in captivity.

Training Manual on Seed Production and Farming of Brackishwater Finfishes I 128

 Soil and water quality management of finfish hatcheries and
culture practices

P.Kumararaja, R.Saraswathy, S.N.Sethi

Introduction
Aquaculture, as the aquatic equivalent of agriculture and terrestrial animal husbandry
business.It has numerous and often poorly defined risks, both biological–physical and social–
economics, and characterized by considerable variability as most biological processes,
including meteorological and other abiotic uncontrolled effects, which are interrelated
(Webber, 1973). The steadily growing importance of aquaculture has compelled
improvements in the technologies necessary for securing the initial and basic requirements
for productive aquaculture; namely the production of fish seed for stocking. A fish hatchery
is a place for artificial breeding, hatching and rearing through the early life stages of animals.
It consists of different units like egg collection, incubation, larval rearing for culture and
nursery rearing facilities. The efficient operation of a fish hatchery depends on a number of
factors such as suitable site selection, soil characteristics and water quality. The site selection
should be a broad, sound and careful process. In the majority of the cases and especially on
those for hatcheries, the most important aspect is water quality in relation with species, live
stages and culture systems. Site selection for aquaculture facilities should be based on culture
potentialities assessments of sites, according to needs on different aspects and criteria,
especially (a) species to be work with, its life-cycle requirements and tolerance limits of main
environmental parameters; (b) present and future working objectives that should be
accomplished; (c) purposes of the facility (educational, research, production, etc.); (d)
working scale (experimental, pilot, commercial, and extension-demonstration); (e) working
aspects (seed production, nursery, and grow-out); (f) type of system that would be used
(enclosures, fish pens, cages, ponds, and tanks); (g) system intensity (extensive, semi-
intensive, intensive, and super intensive).

Important Water Quality Parameters affecting hatchery production

Water quality determines to a great extent the success or failure of a fish cultural
operation. Water is the culture environment for fish and other aquatic organisms. It is the
physical support in which they carry out their life functions Water quality is one of the most
critical factors besides good feed/feeding in fish production. For a successful aquaculture
venture, the dynamics and management of water quality in culture media must be taken into

129 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

consideration. Water quality parameters can be divided into three main categories: physical
(density, temperature); chemical (pH, conductivity, nutrients) and biological (bacteria,
plankton and parasites. All living organisms have tolerable limits of water quality parameters
in which they perform optimally. A sharp drop or an increase within these limits has adverse
effects on their body functions.

1) Water Temperature

It is important to note that intrinsic differences exist in adaptation of fish to water

temperature. For each species, there exists an upper and lower limit, as well as an optimum
range for growth, which changes with development. The temperature for optimum growth of
fish is called the SET, standard environmental temperature. Most warm water fishes need
water temperatures ranging from 20 to 30ºC for their propagation. Fish are stressed and
disease outbreaks occur after a sudden temperature change or when temperatures are
chronically near their maximum tolerance. The metabolic rate of ectothermal animals is said
to double with each 10°C rise in temperature, a relationship called the Q10 factor.
Temperature controls the solubility of gases in water, and the reaction rate of chemicals, the
toxicity of ammonia, and of chemotherapeutics to fish.

Extreme cold or warm temperatures inhibit final gonad development. The optimum
temperature range for development of eggs and rearing of fry is 26 to 28°C. Cobia requires
23-27°C during spawning. If the temperature is too low, hatching and development are
prolonged. At higher water temperatures, embryos develop too fast and there may be a high
incidence of malformed or nonviable fry.

Considerable energy is required to heat or cool water and it is usually too costly to
attempt major changes in water temperature. Therefore, the temperature of the water supply
should be near 80°F before it enters the hatchery. An in-line water heater can be used to
ensure a minimum temperature in the inlet water. The opposite condition may be encountered
late in the spawning season when surface waters become too warm for use in hatcheries.

2) Dissolved gases in water

In addition to the atmospheric gases natural waters contain additional dissolved gases
that result from erosion of rock and decomposition of organic matter. Oxygen (O 2) and
carbon dioxide (CO2) are important molecules because they are involved in photosynthesis
and respiration processes. Nutrients like nitrogen are essential on biological metabolism. But
when is excreted by the fish as ammonia (NH3), in certain circumstances it can be toxic and
Training Manual on Seed Production and Farming of Brackishwater Finfishes I 130
even lethal in high doses. Besides the physical and chemical factors, the biotic component
also can change the water composition. For example, algae can consume of produce oxygen
and carbon dioxide, depending on light presence or absence.

a) Dissolved oxygen

Oxygen is the first limiting factor for growth and well-being of fish. Fish require
oxygen for respiration, which physiologists express as mg of oxygen consumed per kilogram
of fish per hour (mg O2 /kg/h). The respiratory rate increases with increasing temperature,
activity, and following feeding, but decreases with increasing mean weight. At a given
temperature, smaller fish consume more oxygen per unit of body weight than larger fish.
Therefore, for the same total weight of fish in a tank, smaller fish require more oxygen than
larger fish. Actively swimming fish consume more oxygen than resting fish. Oxygen
consumption of fish will increase after feeding; multiple feedings per day (3 or more) will
result in less variation in oxygen demand than 1 to 2 feedings per day. The oxygen
consumption rate of fish of different species ranges range from 200-500 (mg O2 /kg/h).
Oxygen concentration at saturation in a given elevation varies in relationship to water
temperature. Oxygen concentration should not be less than 70% of saturation.
In ponds, the major source of oxygen is from algal photosynthesis and from wind
mixing the air and water. In tanks oxygen is supplied by the inflowing water, which should be
near saturation for the temperature. A common generalization about oxygen requirements for
aquaculture is that the minimum DO should be greater than 5 mg/L for growth of warm water
fish. Thus for a circular tank, oxygen of the effluent water should be at least 5 mg/L. At
temperatures optimum for growth, fish are stressed at if the available oxygen concentrations
less than 5 mg/L. If the condition is chronic, fish stop feeding, growth slows down, stress-
related disease begins.

b) Carbon dioxide

The primary sources of carbon dioxide in fish ponds are derived from respiration by
fish and the microscopic plants and animals that comprise the fish pond biota. Decomposition
of organic matter derived from unfed feed and excreta matter is also a major source of carbon
dioxide in fish ponds. The problem with the potential toxicity of carbon dioxide can be
related to the daily fluctuating pattern of dissolved oxygen and carbon dioxide
concentrations. Carbon dioxide concentrations are highest when dissolved oxygen
concentrations are lowest. If environmental carbon dioxide concentrations are high, the fish

131 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

will have difficulty reducing internal carbon dioxide concentrations, resulting in
accumulation in fish blood. This accumulation inhibits the ability of haemoglobin, the oxygen
carrying molecule in fish blood, to bind oxygen, and may cause the fish to feel stress similar
to suffocation.

Carbon dioxide concentrations are maximum during winter and minimum during
summer. Warm water temperatures increase the metabolism of all pond organisms and
therefore respiration rates are high. It is also a time of year when feeding rates are high. The
decomposition of wastes generated by large quantities of organic matter added to fish ponds
in the summer requires large quantities of dissolved oxygen and produces large quantities of
carbon dioxide. High levels of dissolved carbon dioxide interfere with respiration by eggs and
fry concentrations up to at least 10 ppm seem to be well tolerated, provided that dissolved
oxygen concentrations are adequate.

3) Salinity

Salinity is the dissolved salt content of water and is often expressed as the parts of salt
by weight per thousand parts of water by weight (ppt). Salinity and dissolved solids are made
up mainly of carbonates, bicarbonates, chlorides, sulphates, phosphates, and possibly nitrates
of calcium, magnesium, sodium, and potassium, with traces of iron, manganese and other
substances. Spawning is one of the processes affected by the salinity of water. Some fish
migrate from marine to freshwater environment while others do vice verso for spawning and
complete their life cycle. Asian Seabass requires 28-32 ppt for reproduction. Eggs can hatch
and fry will develop in waters with salinities up to at least 32 parts per thousand.

4) Turbidity

Turbidity is the term associated with the presence of suspended solids. Analytically,
turbidity refers to the penetration of light through water (the lesser the penetration, the greater
the turbidity. Turbidity can be caused by many substances, including microscopic algae
(phytoplankton), bacteria, dissolved organic substances that stain water, suspended clay
particles, and colloidal solids. Turbidity caused by clay particles is generally undesirable
because it keeps light from penetrating the water, and light is required for algal growth. At
very high concentrations, clay particles can also clog fish gills or smother fish eggs. Turbidity
in excess of 100,000 parts per million do not affect fish directly and most natural waters have
far lower concentrations than this. In general, turbidity less than 2,000 parts per million is
acceptable for fish culture.

Training Manual on Seed Production and Farming of Brackishwater Finfishes I 132

 5) Alkalinity and Hardness

Alkalinity is the buffering (alkaline) capacity of the water. Alkalinity is a measure of

the capability of water to neutralize acids. In most natural waters, the predominant bases are
bicarbonate and carbonate. Alkalinity is expressed as ppm equivalent CaCO3.Fish eggs and
fry thrive in waters with a wide range of alkalinity, although waters of very low alkalinity
(<10 ppm as CaCO3) should be avoided as hatchery supplies if possible. These waters are
poorly buffered and pH can fluctuate drastically with small additions of acid or base. More
importantly, dissolved metals such as copper and zinc are very toxic to fry in waters of low
alkalinity. Waters with high alkalinity are undesirable because of the associated excessive
hardness or high concentrations of sodium salts. So water with alkalinities between 120-400
ppm is optimum.
Hardness refers to the amounts of calcium and magnesium in the water and is
expressed as ppm of equivalent CaCO3. Adequate concentrations of environmental calcium
are required for “hardening” of eggs and for normal bone and tissue development of fry. A
minimum of 5 ppm calcium hardness is required for adequate egg hatchability and for
development and vigour of sac fry. Higher calcium concentrations are desirable because
calcium also protects fry from ammonia and metal toxicities. All things considered, hatchery
water supplies should contain at least 20 ppm calcium hardness.

Potential Hydrogen (pH/Acidity)

Acidity refers to the ability of dissolved chemicals to "donate" hydrogen ions (H +).
The standard measure of acidity is pH, the negative logarithm of hydrogen-ion activity. The
pH of most productive natural waters that are unaffected by pollution is normally in the range
of 6.5 to 8.5 at sunrise, typically closer to 7 than 8. The controlling factor for pH in most
aquaculture facilities is the relationship between algal photosynthesis, carbon dioxide (CO2),
and the bicarbonate (HC03-) buffering system. At night, respiration by bacteria, plants, and
animals results in oxygen consumption and carbon dioxide production, producing carbonic
acid (H2C03), then bicarbonate HC03- and H+ions; the increase in H+causes the pH to drop.
During sunlight, respiration continues, but algae use CO2 for photosynthesis reduction in CO2
level consume H+ for HC03- reducing the abundance of H +
ions, and pH goes up. Fish can
die from pH shock, a consequence of a sudden change in pH (1.7 pH units) that may occur
when moving fish from pond to tank, or tank to pond. Toxicity of other compounds to fish,
especially ammonia and chlorine, are affected by pH.

133 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

 6) Presence of Metabolites
a) Ammonia

Of all water quality parameters, which affect fish, ammonia is the most important
after oxygen. Ammonia is the principal nitrogenous waste product of fishes that represents
60% to 80% of nitrogenous excretion of fish. It is also, the main nitrogenous waste material
excreted by gills beside urea and amines and an end product of the protein catabolism. In
water, total ammonia consists of non-toxic (ionized ammonia) referred to as ammonium
(NH4+) and toxic un-ionized ammonia (NH3). The equilibrium between these two forms is pH
and temperatures dependant. Toxicity from high TAN is more likely at high pH and high
temperatures, conditions that occur in mid-summer in ponds with high standing crop of fish,
which are also likely to have a heavy algal bloom, and mid-afternoon pH values close to
9.The NH3 molecule is soluble in lipids. It is 300 to 400 times more toxic than NH 4+. Un-
ionized ammonia (UIA-N) can readily diffuse across the gill membranes due to its lipid
solubility and lack of charge. Ammonia tends to block oxygen transfer from the gills to the
blood and can cause both immediate and long term gill damage. Also it can cause impairment
of cerebral energy metabolism, damage to gill, liver, kidney, spleen and thyroid tissue in fish.

b) Nitrite

Nitrite an intermediary product of bacterial nitrification is highly toxic to fish. A more

rapid growth rate of Nitrosomonas in the biological system can lead to accumulation of
nitrite. It oxidises the haemoglobin to methemoglobin which is incapable of transport of
oxygen. As the pH increases nitrite toxicity increases. Nitrite toxicity decreases slightly as the
hardness and chloride content of water increases.

Management of water quality in finfish seed production

In brood stock tank stocking density should be maintained at 1 kg/m3 in the. The
stocking density in cages can be doubled depending upon the water quality and feed
management. Bloodstock maintained in captive condition should be provided with the
environmental quality prevailing in the sea for maturation and spawning. Even if not all
conditions, the water quality should be maintained to maximum extent to that of sea water.
The source water should be passed through high pressure sand filters to remove the impurities
and toxicants. The problem of algal bloom is minimal in flow through system and tanks with
over cover. Tanks open to sky prone to aquatic weeds which inhibit the light penetration and
restrict the fish movement and feeding. To overcome the problem of aquatic feed, removal of

Training Manual on Seed Production and Farming of Brackishwater Finfishes I 134

unfed feed and metabolites the tank should be cleaned in the early morning hours by reducing
the water level to a minimum. After cleaning bottom and walls of tank replacing the water
results in exchange of 70 -80% water daily. In flow through system the water quality
parameters should be monitored regularly and necessary actions should be taken depending
upon water quality. Dissolved oxygen in the brood pond should remain above 5 mg/L at all
times for successful spawning. If a tank is stocked with fish, over several weeks of a growth
cycle, the fish will grow, reducing their consumption rate (inverse OC-fish size relationship),
but the density (kg/m3) will increase. Flow to the tank will have to be increased or the
population divided to handle the larger oxygen demand. Under emergency conditions,
application of dissolved oxygen enhancers improves the water quality. Water exchange is the
best solution to prevent low DO problem in brood stock tank. Aeration and mixing are the
most effective available mechanical methods for the management of carbon dioxide and
dissolved oxygen. Vigorous aeration accelerates the diffusion of carbon dioxide out of water
and mixing will prevent or minimize the establishment of a carbon dioxide-rich layer of water
near the pond bottom. Maintaining a moderate plankton density (Secchi disk visibility
between 6-12”) will maximize the biological uptake of carbon dioxide. The salinity of the
brood stock and spawning tank should be same. If there is any variation of salinity from the
optimum level it can be restored by mixing with brine solution or freshwater. After 2 – 3
days when fish got acclimatized to the spawning tank conditions, the salinity of the water is
reduced to 24 ppt. The fishes are maintained in this condition for about a week and then the
salinity is increased to 30-32 ppt by daily water exchange over a period of 10 days. Ammonia
and nitrite level can be maintained below the critical limit by providing aeration, removal of
debris and plankton and water exchange.

Rearing of hatchlings through the various developmental stages providing required

environmental parameters and feed is the most important phase in the seed production
technology. Water quality in the rearing tanks is very important for better survival and
growth of the larvae. Water provided to the larval rearing tanks should be free from
flagellates, ciliates and other unwanted pathogenic organisms. Water should be filtered
through biological filters; pressure sand filters and UV radiation filters for get rid of
pathogenic organisms. Residual chlorine should be removed if chlorine treated water is used
because fish larvae is highly sensitive to chlorine. The bottom debris in larval rearing tank
should be removed by siphoning with a filter net of 100-200µ upto 9 days and 200-400 µ
mesh size afterwards. The salinity of the water should be maintained to a level of 25- 30 ppt

135 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

and a temperature of 27-29oC. The temperature can be maintained by heaters during winter
season. Un-ionized ammonia is quite toxic to sac fry and early swim-up fry. Ideally, water in
rearing troughs should be free of ammonia for optimal health and growth of fry, and the
maximum concentration of un-ionized ammonia that should be allowed is about 0.05 ppm
NH3-N. Above this concentration, fry develop more slowly and are more susceptible to
infectious diseases.

Soil and water requirements for brackish water finfish grow out culture

Soils are a major factor in pond aquaculture and the condition of pond bottom
influences water quality and production. Concentrations of nutrients and phytoplankton
productivity in pond water are related to pH, and nutrient concentration in soils. Before
initiating aquaculture operation, one should be well acquainted with the nature of soil as it
affects the fish production.

Soil texture has direct effect on the productivity of ponds. Soils with moderately
heavy texture such as sandy clay, sandy clay loam and clay loam are highly suitable for
aquaculture. In general soil pH ranging between 6.5 and 7.5 are best suited for brackish water
environment. Under this pH range, the availability of nitrogen, phosphorus, potassium, sulfur,
calcium and magnesium concentration is maximum. Soil rich in CaCO 3 content promotes
biological productivity as it enhances the breakdown of organic substances by bacteria
creating more favourable oxygen and carbon reserves. The productive soil should have
calcium carbonate more than 5%. Organic matter is an important index of soil fertility. It
helps in prevention of seepage loss, increases arability of pond bottom and supplies nutrients.
It reduces turbidity of pond water and act as antioxidants. Organic matter influences
microbial activity and productivity of pond. Soil which has organic carbon content less than
0.5 % is low productive, 0.5-2% is medium productive and > 2% high productive. Optimum
value is 1.5-2%. In sediments, when organic matter exceeds the supply of oxygen, anaerobic
condition develops. This reducing condition can be measured as the redox potential and is
represented as Eh. The redox potential of mud should not exceed -200 mV.

Conclusion

Fish culture today is hardly possible without the artificial propagation of fish
seeds of preferred cultivable fish species. The need for the production of quality fish seed for
stocking the fish ponds and natural water bodies has indeed increased steadily. Water is the
physical environment where fish develop, growth and reproduces. Successful hatchery

Training Manual on Seed Production and Farming of Brackishwater Finfishes I 136

operations can be possible by only use of good quality water and regular monitoring of water
quality parameters and maintaining it by different measures such as water exchange, aeration,
etc. To ensure sustainable fish production, soil and water quality are two major parameters.
Daily monitoring of the pond conditions and fish behaviour along with accurate record
keeping helps the farmer to recognize and prevent deleterious environmental conditions in the
pond and there by maximize the production and profit.

Appendix

Table 1: Suggested water quality parameters for water resources

Parameter Value
pH 6.5 – 9.0
Dissolved oxygen 5 ppm – saturation
Carbon dioxide 0 – 10 ppm
Total alkalinity (as CaCO3) 50 – 400 ppm
Total Ammonia Nitrogen (TAN) 0 – 0.05 ppm
Nitrate 0 – 3.0 ppm
Nitrite 0 – 0.05 ppm
Phosphate 0.01 – 3.0 ppm
Manganese 0 – 0.01 ppm
Iron 0 – 0.5 ppm
Zinc 0 – 0.05 ppm
Lead 0 ppm
Hydrogen sulphide 0 ppm
(Saraswathy et al., 2012)

137 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

Table 2: Desirable range of some of the water quality parameters in a brood
stock tank
Parameter Value
pH 7.0 – 8.2
Dissolved oxygen More than 5 ppm
Carbon dioxide 0 – 10 ppm
Total alkalinity (as CaCO3) 50 – 400 ppm
Total Ammonia Nitrogen (TAN) Less than 0.1 ppm
Nitrite Less than 0.01ppm
Phosphate 10-20 ppm
Suspended solids 2 - 5 ppm
(Thirunavukkarasu et al., 2013)

Table 3: Desirable range of some of the water quality parameters in a larval

rearing tank
Parameter Value
Temperature 28-30oC.
Dissolved oxygen More than 5 ppm
pH 7.0 – 8.2
Total Ammonia Nitrogen (TAN) Less than 0.1 ppm
Nitrite Less than 0.01ppm
Phosphate 10-20 ppm
Suspended solids 2 - 5 ppm

(Kailasam et al., 2013)

Table 4: Soil requirements for brackish water finfish culture

Parameter Value
pH 6.5-7.5
Electrical conductivity (dS/m) >4
Clay content (%) 18-35
Organic carbon (%) 1.5-2.0

Training Manual on Seed Production and Farming of Brackishwater Finfishes I 138

 Calcium carbonate (%) >5
Available nitrogen (mg/100g) 50-70
Available phosphorus (mg/100g) 4-6

(Saraswathy et al., 2015)

Table 5: Optimum water quality parameters for fish culture ponds

Parameter Value
pH 6.5 – 9.5
Dissolved oxygen >5 ppm

139 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

 Transportation methodology of brackishwater finfishes
Prem Kumar, S. N. Sethi, Rekha M. U., Dani Thomas, R. Subburaj and G. Thiagarajan

1. Introduction

There are two types of transport systems for live fish - the closed system and the open
system. The closed system is a sealed container in which all the requirements for survival are
self-contained. For example sealed plastic bag partly filled with water and oxygen. The open
system consists of water-filled containers in which the requirements for survival are supplied
continuously from outside sources. The simplest of these is a small tank with an aerator
stone.

2. Factors associated with live fish transportation

Fish survival in a good state of health during transport is influenced by a number of

factors, or combination of factors

2.1 Health of fish

The fish to be transported must be healthy and in good condition. Weakened

individuals should be eliminated from the transportation. The fish to be transported, except
for the larval stages should be starve for at least a day; if the fish is not starved, the possible
time of transport is reduced to a half, though the conditions may be the same (Pecha, Berka
and Kouril, 1983; Orlov et al., 1974). Natural ice is used to cool the water; the ice of carbonic
acid should be avoided. As a guide ratio, 25 kg of ice will cool 1 000 litres of water by 2°C.
If the water contains fish during the cooling process, the temperature drop should not be
faster than 5°C per hour.

2.2 Dissolve oxygen

The most important single factor in transporting fish is providing sufficient level of
dissolved oxygen. The ability of fish to use oxygen depends on their tolerance to stress, water
temperature, pH, and concentrations of carbon dioxide and metabolic products such as
ammonia. The crucial factors underlying oxygen consumption by fish in relation with oxygen
metabolism during transport are fish weight and water temperature. Heavier fish and those
transported in warmer water need more oxygen. During fish transport in closed systems with
pressurized oxygen atmosphere, oxygen content in water usually is not a limiting factor
because there is enough pressurized oxygen in a closed bag. In closed systems, slight shaking

Training Manual on Seed Production and Farming of Brackishwater Finfishes I 140

of the bag supports the penetration of atmospheric oxygen to water. During long steps when
the bags with fish are left without movement, the fish may die though the oxygen reserve in
the bag is still high.

2.3 pH, carbon dioxide and ammonia

The water pH level is a control factor because the proportions of toxic ammonia and
CO2 contents are direct functions of Ph. With increasing transport time, CO2 production
through fish respiration reduces water pH towards acidity. Water pH levels about 7–8 are
considered as optimum. Rapid changes in pH stress fish, but buffers can be used to stabilize
the water pH during fish transport. The organic buffer trishydroxylmethylaminomethane is
quite effective in fresh and salt water. It is highly soluble, stable and easily applied. Dose of
1.3–2.6 g/litre are recommended for routine transport of fish (Piper et al., 1982). In general,
for each milliliter of oxygen a fish consumes, it produces approximately 0.9 milliliters of
CO2. Another important factor is chlorine concentration in water, although - like carbon
dioxide - chlorine is also removed from the water by aeration. The concentration of 0.2
mg/litre is considered as dangerous (Shevchenko, 1978).

Ammonia (NH3) builds up in transport water due to protein metabolism of the fish
and bacterial action on the waste. Decreasing metabolic rate of the fish by lowering the water
temperature, and thus lessening fish activity, reduces the production of NH3. The production
of NH3 by bacterial action can be decreased by starving the fish before transportation/
emptying the stomach and intestine. Temperature and time of last feeding are important
factors regulating ammonia excretion. For example, trout held in water at 1°C excrete 66%
less ammonia than those held in 11°C water. Fish larger than 10 cm should be starved at least
48 h; those 20 cm and larger should be starved 72 h (Piper et al., 1982). The amount of un-
ionized ammonia increases as water temperatures and pH increase

2.4 Temperature

Water temperature is an important factor. When water temperature is low, the pH

remains higher and fish metabolism decreases. The generally applicable zones of optimum
temperatures for transported fish are 6–8°C for cold-water fishes and 10–12°C for warm-
water fishes in summer. Naturally, these temperature ranges do not apply to the early stages
of fish fry. The early fry of cyprinids cannot be transported at temperatures below 15°C.

141 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

2.5 Density

As to fry, the ratio of the volume of the fish transported and the transport water should
not exceed 1:3. Heavier individuals, e.g., parent fish can be transported in a fish: water
weight ratio of 1:2 to 1:3, but with smaller organisms this ratio decreases to 1:100 to 1:200
(Pecha, Berka and Kouril, 1983).

2.6 Stress during transportation

When fish were transported at higher densities, the levels of corticoids and glucose in
the plasma increased and was retained when the transport was finished. During transportation
besides transportation stress, stress may also be caused by the deterioration of water quality
(Erikson et al., 1997), salinity and temperature fluctuations (Mires and Shak, 1974), which
might consequently alter the metabolism of the fish. Transportation of Grey mullet for
stocking is mainly carried out in the fry and fingerling stages. Biochemical parameters serve
as reliable indicators of physiological status of organism (Ferry-Grahm and Gibb, 2001). The
tertiary response is the final stage, which leads to disease or exhaustion, growth retardation
and finally death (Chatterjee et al., 2006). Catecholamine and cortisol induce glycogenolysis
and gluconeogenesis respectively. Both processes together cause a rise in blood glucose level.
Blood glucose and hepatic glycogen are therefore commonly measured parameters of stress
response (Manush et al. 2005).

3. System of live fish transportation

3.1 Closed system

Transport of fry in polyethylene bags with oxygen is common example of closed

system of transportation. It is generally used to transport fry.

3.2 Polythene bags

The bags used for fish transport in water with oxygen atmosphere are produced in a
number of modifications. They are manufactured from a thin (soft) or thicker (hard)
transparent polyethylene foil and usually have the shape of sack or sleeve. The bags of the
traditional shape (sacs) usually have the dimensions of 0.8– 1.1 . 0.35–0.45 m. The upper end
is usually fully open. The bottom either has a seam in the middle or consists of a rectangular
piece of foil; the latter variant is better because it helps avoid losses of the fish squeezed in
the corners. During transportthe bags with fry are placed in outer cases protecting the bags
against mechanical damage. These cases can be cardboard boxes, suitable plastic containers,

Training Manual on Seed Production and Farming of Brackishwater Finfishes I 142

wide polyethylene cans, and polystyrene boxes. If water with transported fry is to be cooled,
bags with ice should be placed under the fish-transport bags on the bottom of the polystyrene
box. It is not recommended to put the ice inside the transportbag. The volume of ice placed
under the bag with transport water is usually 10–20% of the transport water. Other sealed
container is generally made of cured plastics

3.3 Open system

In all cases of fish transport in open systems, it should be borne in mind that even a
short-time transport of 10–30 m in open plastic or metal tanks should be done under the
conditions of constant air or oxygen supply. In recent year most tanks are constructed are
insulated, usually with styrofoam, fiberglass or urethane.

Styrofoam and urethane are preferred materials because of their superior insulating
qualities and the minimal effect that moisture has on them. A well-insulated tank minimizes
the need for elaborate temperature-control systems and small amounts of ice can be used to
control the limited heat rises. Circulation is needed to maintain well-aerated water in all parts
of the tank. Although most tanks presently in use are rectangular, the trend in recent years has
been towards elliptical tanks, such as those used to transport milk. Self-priming pumps
powered by gasoline engines are used to circulate water in many transport units. Pumping or
aerating systems should be able to circulate at least 40 percent of the tank water per minute.

4. Mitigation of transportation stress

The chemical used during transportation to reduce the transportation and handling
stress and increase the survival of the fish. Some of the commonly used chemicals include
anaesthetics, water-hardening and oxygen-producing chemicals, bacteriostatics, buffering and
antifoam.

4.1 Use of tranquilizer

It is best to sedate the fish in the holding facility for 30 min before loading and then to
continue exposure to a lower concentration of sedative during transport. Anaesthesia usually
applies only to transport brood fish. In practice, the fish are first tranquilized with the normal
dose and put into the transport tank, where original concentration is diluted by 50 percent by
adding the same amount of fresh water. The brood fish will remain tranquillized well in that
diluted solution (Woynarowich and Horvath, 1980). As Woynarowich and Horvath (1980)
assert, fish transport in cold water of 5–10°C is the simplest and best method of anaesthesia.

143 I Training Manual on Seed Production and Farming of Brackishwater Finfishes

Among the broad spectrum of anaesthetics, tricaine methanesulfonate (MS-222) and
quinaldine (2–4 methylchinolin) appear to be used most frequently.

4.2 Use of sodium chloride and calcium chloride

Handling stress and delayed mortality of fish can be decreased by the addition of
sodium chloride (NaCl) and calcium chloride (CaCl2) to the transport water. The sodium ion
tends to “harden” the fish and reduce slime formation, and the calcium ion suppresses
osmoregulatory and metabolic dysfunction. Calcium chloride may not be needed in hard
water already containing high concentrations of calcium. Dupree and Huner (1984)
recommended the addition of 0.1 to 0.3 percent salt and 50 mgL-1 calcium chloride. Some of
the fishes that tolerate wide ranges of salt in the water, such as striped bass, tilapias, carp, can
benefit from as much as 0.5 percent salt.

4.3 Oxygen source

Huilgol and Patil (1975) tested the use of hydrogen peroxide on transported carp fry
and found that one drop (1 ml = 20 drops) of hydrogen peroxide (6 percentconcentration),
applied to 1 litre of water, increased the oxygen content by 1.5 mgL-1 when the temperature
was 24°C. CO2 content and water pH were not influenced by the addition of hydrogen
peroxide.

Training Manual on Seed Production and Farming of Brackishwater Finfishes I 144