Molecular Identification of Dermatophytes Among Clinical Isolates

See discussions, stats, and author profiles for this publication at: https://www.researchgate.
net/publication/327424931
MOLECULAR IDENTIFICATION OF DERMATOPHYTES AMONG CLINICAL

ISOLATES
Article · June 2018
CITATION READS
1 132
2 authors, including:
Hana K Maikhan
Independent Researcher
7 PUBLICATIONS 2 CITATIONS
SEE PROFILE
Some of the authors of this publication are also working on these related projects:
Isolation and Molecular Identification of Trichophyton rubrum var. raubitschekii from the Infant Groin View project
All content following this page was uploaded by Hana K Maikhan on 04 September 2018.
The user has requested enhancement of the downloaded file.

Asian Journal of Natural & Applied Sciences Vol. 5(2) June 2016
MOLECULAR IDENTIFICATION OF DERMATOPHYTES AMONG

CLINICAL ISOLATES
Khalid A. Habeb1, Hana K. Maikhan2, Shwan K. Rachid3
1
Professor, Microbiology, Department of Biology, College of Science for Women, University of
Baghdad, 2 Department of Biology, College of Education, University of Garmian, Kurdistan Region,
3
Professor, Molecular Biology, Assistant Chief of Charmo University, Kurdistan Region, IRAQ.
hana.baghdad@gmail.com
ABSTRACT
Aim: This study aimed to identification of dermatophyte species in clinical isolates by
both classical and molecular methods, using universal primers for amplification of
ITS gene. In addition, identification at a species level was carried out by using PCR-
restriction fragment length polymorphism (RFLP) technique.
Methodology and Results:In the present study, out of 220 suspected patients (157
females and 63 males) with dermatophytosis were identified by the supervision of
specialized dermatologist in the derma unit of the General hospital in Kalar
district/Sulaimania province\Iraq during the period from middle of November 2014 to
the end of June 2015. Samples were collected from patients included hair fragments,
skin scraping and nail clipping which transferred to the research laboratory of
Biology Department at Faculty of Education \ University of Garmian, where they
immediately examined. Based on the conventional laboratory methods, 80 clinical
isolates of dermatophytes showed positive culture which belonging to three genera
(Trichophyton, Microsporum and Epidermophyton) and 13 species, Trichophyton
rubrum (downy and granular types) was the most common species 35% followed by
T.mentagrophytes 17.5%, M.canis 10% and T.tonsurans 7.5%. Other species T.
soudanese; T.interdigitale; T. terrestre represented 5% of dermatophytes while
T.concentricum; T.schoenleinii; T.verucosum; Microsporum audouinii; M.gypsium
and Epidermophyton flocosum conistitute 2.5%. Molecular identification of
dermatophytes was carried by using the primers ITS1 and ITS4 to amplify the internal
transcript spacer (ITS) region of ITS1 5.8S ITS2 in rDNA gene, later the species
identification was made by digestion of amplified ITS regions by BstN1 restriction
enzyme in restriction fragment length polymorphism (PCR-RFLP) to produce distinct
band patterns.
Conclusions: Survey of dermatophytes showed that PCR-RFLP is a rapid and easy
method for dermatophytes identification; therefore we suggest using the PCR-RFLP
corresponding to the conventional laboratory identification including macroscopic
and microscopic examination.
Keywords: PCR, RFLP, Molecular
INTRODUCTION
Dermatophytes referred as a unique group of superficial keratinophilic and filamentous fungi
which have the ability to invade keratinized tissue of the skin, hair and nail in human and
animals leading to dermatophytosis or tinea (ring worm) (Alkahafajii, 2014). They are based
on the keratin as a source of nutrient and cause its hydrolysis by releasing number of enzymes
such keratinase as a proteolytic enzyme of keratin protein which is very hard and strong
(Sharma and Swati, 2012). Dermatophytes mainly classified into three anamorphic genera
Trichophyton, Microsporum and Epidermophytonin as well as teleomorphic genus
ISSN: 2186-8476, ISSN: 2186-8468 Print Leena and Luna International, Chikusei, Japan. Copyright © 2016
www.ajsc.leena-luna.co.jp (株) リナアンドルナインターナショナル, 筑西市,日本 P a g e | 108
Arthroderma, and they include more than 30 species. Among the most common etiological
agent of dermatophytosis in human include T.rubrum, Tmentagrophytes, T.tonsurans as well
as M.canis, M.gypseumand E.flocosum (Weitzman and summerbell, 1995).The conventional
laboratory methods for identification still based on the results of macroscopic and
microscopic examination. An important characters in the microscopic examination based on
the characteristic features of macro and microconidia, hyphal properties and colony properties
which seen on SDA (Kaneet al., 1997). Based on Davison et al., (1980), numerous molecular
approaches have been developed for identification of dermatophytes at the level of species
and strains. Consequently, some biotypes were identified preciously as species, at the present
cannot be distinguished molecularly (Nenoffet al., 2013). At the same time, these methods
cause confusion of taxonomical classification of dermatophytes and lead to novel
classification (Gräser and Summerbell, 2008). Development of molecular technique provides
better understanding in classification, epidemiology and ecology. At present, methodologist
depends on DNA amplification and sequencing analysis for identification of dermatophytes
(Cafarchia et al., 2013) and the most abundant approach for identification include gene-
specific PCR, PCR-RFLP, PCR fingerprinting, DNA hybridization and sequencing of ITS
region in rDNA gene which provide useful method for identification and phylogenetic
analysis of dermatophytes (Li et al., 2008).
MATERIALS AND METHODS:

Sample Collection
Two hundred twenty clinical specimens (63 Male and 157 Female) including hair, skin, and
nail were collected from patients between the periods from middle of November 2014 to the
end of June 2015, where suspected by dermatologist at Kalar General Hospital, Kalar,
Kurdistan region, Iraq. Each sample was divided into two parts one for direct microscopic
examination by KOH and the other for culturing on PDA and SDA with cyclohexamide and
chloramephinicol(SDACC), then incubated at 25-30ᵒC for 7-21 days and examined at regular
intervals. Macroscopical, microscopical and physiological features studied on the developing
colonies and the positive diagnosed dermatophytes preserved for molecular diagnosis.
Extraction of Fungal DNA
Fungal genomic DNA was extracted by using OMEGA bio-tek E.Z.N.A. ® SP (USA) fungal
DNA Mini Kit according to the instructions recommended by the manufacturer. About 100
mg of fungal tissue was collected in a 1.5 or 2 mL micro centrifuge tube and freeze by
dipping in liquid nitrogen then grinded to powder, 4μLRNase A and 600μL SFG1 Buffer was
added, vortexes vigorously to mix thoroughly. The mixture was incubated at 65°C for 10 min.
The sample was mixed twice during incubation by inverting tube then 140 μL SFG2 Buffer
was added and vortexes to mix thoroughly and Centrifuged at 10,000 × g for 10 minutes. The
Homogenizer Mini Column was inserted into 2ml collection tube then the supernatant was
transferred to a new microcentrifuge tube and centrifuged at 10,000 × g for 2 min. and the
supernatant was removed. The lysate was transferred to a new 1.5ml microcentrifuge tube
then 1.5 volume of SFG3 was added and vortexes to obtain a homogenous mixture. The
HiBind DNA Mini Column was inserted into 2ml collection tube then 650 μL of the sample
was transferred to HiBind DNA Mini Column and centrifuged at 10,000 × g for1 min., the
filtrate was discarded and the collection tube reused This step was repeated until the
remaining sample has been transferred to the HiBind DNA Mini Column. The column was
transferred into a new 2ml Collection tube then 650 μL of SPW Wash Buffer was added and
centrifuged at 10,000 × g for1 min. The filtrate was discarded and the Collection tube was
reused. This step was repeated. The empty HiBind DNA Mini Column
Copyright © 2016 Leena and Luna International, Chikusei, Japan. ISSN: 2186-8476, ISSN: 2186-8468 Print
109 | P a g e (株) リナアンドルナインターナショナル, 筑西市,日本 www.ajsc. leena-luna.co.jp
was centrifuged at maximum speed for 2min. to dry the membrane then the HiBindDNA
Mini Column was transferred into a nuclease-free 1.5 or 2ml microcentrifuge tube. 100 μL of
Elution Buffer ( heated into 65 ᵒC) was added then left for 3-5 min. at room temperature and
centrifuged at10,000 × g for 1 min.( This step was repeated) and the DNA sample was stored
at -20.
Primers and PCR Analysis
The ITS region in rDNA gene of different dermatophyte species was amplified by using one
pair of primers, symbolized as ITS1 forward primer (5’ TCC GTA GGT GAA CCT GCGC-
3’( TCC GTA GGT GAA CCT GCGC) and ITS4 revers primer ITS4 5’( TCC TCC GCT
TAT TGA TAT GC -3’). PCR was performed in 25 μL of PCR reaction mixture containing
was prepared by mixing12.5 μL of Prime Taq DNA Polymerase (2X master mix) containing:
Prime Taq DNA polymerase, 10 X reaction buffer containing MgCl2, Tris HCL, (NH4)
2SO4 and PCR enhancer. dNTPmixture, protein, stabilizer and sediment and 2 X loading dye
(GeNet Bio\Korea) which mixed with1.5 μL of each primer, 3.5µL of DNA template, 1.5 μL
of each primer and 6 μL of deionized water was added to made up the total volume of 25 μL
PCR reaction. The PCR reaction were spun down shortly for 5sec. and placed in thermal
cycle (TCY, Crealcon, and NL). Amplification conditions of each isolate were: initial
denaturation (1 cycle 95ᵒC for 5 minutes) followed by 35 cycle: denaturation (95ᵒC for 30
seconds), annealing (58-61ᵒC for 1 minute), extension (72ᵒC for 2 minute) followed by final
extension (72ᵒC for 5 minutes).Then the PCR products were loaded visualized on 1.5%
agarose gel and stained with ethidium bromide and visualized by UV transilluminator. A
100bp DNA marker was used as a reference to determine the size of fragments.
PCR-RFLP
In order to identification at a species level by specific PCR, all PCR products were subjected
to digestion with restriction enzyme BstN1 (BioLabs, England). The mixture contained 10 μL
of PCR product, 1.5 μL of 10 X NEB buffer, 0.5 μL of BstN1 enzyme,which recognizes the
sequence 5’ CC (T/A) GG 3’ to a final volume 12 μL. Subsequentlyreactions were incubated
at 60 ᵒC in dry oven for one hour. All PCR digested products were electrophoresed in a 2 %
agarose gel, after placing 5 μL of the ladder in the first well and 10 μL of the PCR products in
the other wells of the gel which supplied to the power at 90 volt for 90 min. The gel was
observed under a UV trans-illuminator and identification of the isolates was carried out
through comparing the electrophoretic RFLP patterns with those profiles (Jackson et al., 1999;
Matehkolaei, 2012; Elavarashi et al, 2013).
RESULTS
A total of two hundred twenty clinical specimens (63 Male and 157 Female) including hair
fragments, skin scraping and nail clipping, 200 (90.9%) of specimens showed positive result
by 10% KOH examination and 80 (36.4%) of clinical specimens revealed positive cultures of
dermatophytes(Table 1).
Table 1. Positive and negative results of the direct examination and laboratory culture of
dermatophytes
Total Number Direct Examination by KOH Laboratory Culture

Positive Negative Positive Negative
% % % %
220 result result result result
200 90.9 20 9.09 80 36.4 140 63.6
From 200 cases were showed positive KOH examination, only 80 (36.4%) revealed positive
culture for dermatophytes which belonging to three genera Trichophyton,Microsporum and
Epidermophyton and 13 species as seen in Figure 1&2. Conventional identification of
dermatophytes based on macroscopic features (colony surface, colony reverse and texture of
colony), microscopic examination (macroconidia, microconidia, vegetative hyphae and
arthroconidia) and some physiological characters.T.rubrum (downy and granular strains) was
the most common isolate (35%) followed by T.mentagrophytes (17.5%), M.canis (10%) and
T.tonsurans (7.5%) (Table 2).
Table 2. Frequency and ratio of fungal species were positive culture
N. Isolates Number of isolates %

1. Trichophyton rubrum 28 35%
2. Trichophyton mentagrophytes 14 17.5%
3. Trichophyton tonsurans 6 7.5%
4. Trichophyton soudanese 4 5%
5. Trichophyton interdigitalae 4 5%
6. Trichophyton concentricum 2 2.5%
7. Trichophyton schoenleinii 2 2.5%
8. Trichophyton terrestre 4 2.5%
9. Trichophyton verucosum 2 5%
10 Microsporum canis 8 10%
11 Microsporum audouinii 2 2.5%
12 Microsporum gypsium 2 2.5%
13 Epidermophyton floccosum 2 2.5%
14 Total 80
A B
C D
E F
Figure 1: colonial morphology of dermatophytes on SDACC at 25ᵒC after one week of incubation. A
& B) Colony surface and reverse of T.rubrum. C) Colony surface of T.mentagrophytes . D) Colony
surface of E.flocosum. E & F) Colony surface of M.canis
A B
C D
C D
Figure 2: microscopic morphology of dermatophytes showing micro and macroconidia (40X). A) 1)

Macroconidia 2) Microconidia in T.rubrum. B) 1)Macroconidia . 2) Microconidia in
T.mentagrophytes . C) Macroconidia in E.flocosum. D) Macroconidia in M.canis
Fungal genomic DNA was extracted from clinical isolates of dermatophytes using liquid
nitrogen and Omega genomic DNA extraction kit and the genomic DNA was analyzed by
electrophoresis on 1% agarose gel. In Species –Specific PCR the ITS region on rDNA was
amplified by using primers ITS1 and ITS4 which resulted in PCR product size in the range
between 550 inT.tonsurans and 740bp in M. canis and E.flocosumas shown in table (3). The
PCR products were obtained previously by amplification of the ITS region subjected to the
digestion by BstN1 restriction enzyme. This analysis showed the polymorphism of ITS region
for different species of dermatophytes. Digestion of ITS PCR products revealed a number of
recognition patterns with the exception of M.audouinii which was none. The size of ITS PCR
products before and after digestion demonstrated in table (3) and figure (3).
A
Figure 3. (A) PCR amplified of ITS regions. (B) Digestion of amplified ITS product by BstN1
restriction enzyme (PCR-RFLP) for 23 clinical isolates of dermatophytes. All products were
electrophoresd in 2% agarose gel with ethidiumbromide.Lanes: 1,T.rubrum; 2, T.tonsurans;
3,T.rubrum; 4, M.audouinii; 5, T.concentricum; 6, T.rubrum7, M.gypseum; 8, M.canis;
9,T.mentagrophytes; 10, T.schoenleinii; 11,T.rubrum; 12,T.terrestre; 13,M.canis; 14,
T.interdigitalae; 15,T.rubrum; 16, T.rubrumvar.rubitiskii; 17,E.flocosum; 18,T.mentagrohytes;19,
T.soudanense; 20,T.soudanence; 21,T.mentagrophytes; 22, T.mentagrophytes; 23, T.verucosum
Table 3. PCR product of ITS region in dermatophyte species before and after digestion
Size of amplified ITS Size of digested amplified
N. Dermatophyte species
region befor digestion ITSproduct
1 T.rubrum 690bp 380, 180 , 100 and 30bp.
2 T.mentagrophytes 690bp 250,180,160 and 120bp.
3 T.tonsurans 550bp 280,100,100 and 70bp
4 T. soudanense 680bp 350,180,100 and 50bp
5 T.concentricum 650bp 360 and 290bp
6 T.verucosum 650bp 380,180,100 and 10bp.
7 T.schoenlenii 650bp 400 and 250bp
8 T.interdigitalae 700bp 250,180,150,120
9 T.terrestre 630bp 250,190,120 and 80bp
10 M. canis 740bp 440,160 and 100 and 40bp.
11 M.audouinii 600bp 600bp
12 M.gypseum 650bp 400,250bp
13 E.flocosum 740bp 400,250 and 180bp.
DISCUSSIONS
The results we have obtained revealed that among 220 suspected patients with clinical
symptoms of dermatophyte infection, 200 cases (90.9%) were found to be positive by KOH
examination while the positive cultures were 80 cases (36.4%) and the remaining cultures
were suspected to be contaminated by Rhizopus , Aspergillus , Penicillium Alternaria and
Fusarium. These data revealed that the KOH examination was false positive for 120 cases
and this result is unacceptable. This variation between the results of KOH test and laboratory
culture may be due to that the microscopic examination by KOH is a simple technique which
giving fast presumptive diagnosis (Sarika et al., 2014) and therefore the laboratory culture
was more sensitive than the direct examination by KOH. This study showed similarity with
the results of Singh and Beena (2003) who's found that KOH test was less sensitive than the
laboratory culturing of dermatophytes. On the other hand the negative results were obtained
from the laboratory culture of dermatophytes may be due to error in the sampling process or
inadequate specimen which subjected to splitting to perform microscopic examination and
laboratory culturing. In addition, the presence of saprophytic fungi which accompanied with
the dermatophytes at the infected site, possibly compete with it and prevent the
dermatophytes from growing in the agar medium plate (Milne,1996 ; Robson, 2012). The
most common cause of negative culture was belonged to inappropriate topical use of
corticosteroid drug which had been taken from the patient (Colleeet al.,1996;Hayette and
Sacheli, 2015). In the present study Trichophyton species represented 82.5% (66 cases)
followed by Microsporum species 15% (12 cases) and Epidermophytonflocosum which
conistitute 2.5% (2 cases) from the positive culture. Among Trichophyton species, T.rubrum
was the predominant dermatophytes 35% and this is similar to the result of Hay, (1999);
Enemuor and Amedu, (2009); Salman, et al., (2013) and Bhatia and Sharma, (2014) followed
by T.mentagrophyte 17.5%, M.canis 10% and T.tonsurans 7.5% and the same results were
reported by Hanumanthappa et al.,(2012). From 28 clinical isolates of T.rubrum, 6 isolates
were identified as downy types which characterized by absence of macroconidia, 22 isolates
were granular types which revealed oval shaped microconidia and cylindrical shaped
macroconidia with appendix or tail. Of the 14 clinical isolates of T.mentagrophytes, 5 isolates
were identifies as downy type and 9 isolates were granular type. Morphologically, the
granular type revealed powdery to granular colonies with white color at the surface and
brown color at the reverse, while the downy variant exhibited white colonies with velvety to
cottony texture and no pigmentation at the reverse and microscopically showed typical coiled
and areal hyphae which absent in the granular type. The alterations which have been seen in
the physiological and morphological properties of dermatophytes complicate or hamper the
diagnosis process. This variation may be due to the culture technique, incubation temperature
and the use of pharmaceutical (Brilhante et al., 2005). In the present research there were
many difficulties in the phenotype identification of dermatophytes. So, molecular techniques
were employed to confirm the identification. Fungal genomic DNA of was extracted by using
Omega DNA kit in addition to the liquid nitrogen which utilized to lysis the fungal cell wall.
Later, the genomic DNA was analyzed by electrophoresis on 1% agarose gel. Molecular
identification of the positive cultures of dermatophytes was achieved by gene- specific PCR
followed by PCR-RFLP for digestion the PCR products which resulted from amplification of
ITS region in rDNA gene. In the species or gene specific PCR, the ITS region was amplified
by using the primers ITS1 and ITS4 which resulted in PCR products ranged between 550-
740bp. The variation which have been seen in the length of ITS region among dermatophytes
confirmed the results of laboratory conventional identification and these results showed good
similarity with the results of Jackson et al., (1999); Li et al.,(2008) , Al- Khafajii (2014) ,
Ahmadi et al., (2015) ,Ghojogh et al., (2015). Several studies showed
that PCR-RFLP analysis by using single restriction enzyme BstN1 was able to distinguish all
species of dermatophytes during the formation of different band patterns. The PCR- RFLP
targeting ITS region is a rapid, inexpensive, easy to application and reliable method for
dermatophyte differentiation at species level (Mochizuki et al., 2005;Ahmadi et al., 2015). In
this work, the amplified products of ITS regions from all species were subjected to digest by
the BstN1 endonuclease restriction enzyme provides between two to four recognition sites
with the exception of M.audouinii which was none and only revealed the original band
600bp. In addition, the morphological and microscopic examination of T.interdigatale
revealed similarity with those in T.mentagrophytes, so much often difficulty to distinguish
between them however, the patterns were obtained from ITS RFLP in T.interdigitale
exhibited variation from those obtained by T.mentagrophytes. The profiles of electrophoretic
analysis of patterns were obtained from ITS- RFLP in all clinical isolates showed similarity
with those of Jackson et al., (1999); Mochizuki et al., (2003); Elavarashiet al., (2013); Al-
Khafajii (2014); Ahmadi et al., (2014); Ghojoghi et al., (2015) who have identified
dermatophytes by PCR RFLP and showed the same patterns of ITS RFLP. However, an
important fact which observed in the present study during the performance of PCR- RFLP
and differ from the previous studies, that the patterns bands were below 100bp not revealed
on the gel during the electrophoretic analysis and this is may be due to the small molecular
weight of these packages bands and cannot be shown which needs to increase the molecular
weight of the agarose gel.. Furthermore, the results were obtained from ITS RFLP in
T.concentricum showed two bands 360 and 290bp and the clinical isolate number 6 of
T.rubrum revealed three bands and these results differ from the results of the other isolates in
the same species of our study and also showed variation from the previous studies. This
alteration may be due to mutation in the ITS region or mistakes in the identification process.
Consequently, the analysis of ITS region by BstN1endonuclease enzyme was provided simple
method for dermatophytes characterization and this analysis showed the polymorphism of
ITS region for strains and species of dermatophytes (Jackson et al., 1999).
REFERENCES
[1] Ahmadi, B., Mirhendi, H., Shidfar, M. R., Nouripour-Sisakht, S., Jalalizand, N.,
Geramishoar, M., & Shokoohi, G. R. (2015). A comparative study on morphological
versus molecular identification of dermatophyte isolates. J Mycol Med, 25 (1), 29-35.
[2] Al-Khafajii, K. (2014). Myco-epidemiologic and genetic study of rermatophytosis and
non dermatophytes in middle euphrates Iraq. African Journal of Microbiology
Research, 8 (24), 2381-2386.
[3] Bhatia, V. K., & Sharma, P. C. (2014). Epidemiological studies in human patients in
Himachal Pradesh, India. J. Springer Plus, 3, 134.
[4] Brilhante, R. S. N., Rocha, M. F. G., Cordeiro, R. A., Rabenhorst, S. H. B., Granjeiro,
T. B., Monteiro, A. J., & Sidrim, J. J. C. (2005). Phenotypical and molecular
characterization of Microsporum canis strains in north-east Brazil. J ournal of Applied
Microbiology, 99, 776-782.
[5] Cafarchia, C., Iatta, R., Latrofa, M. S., Graser Y., & Otranto, D. (2013). Molecular
epidemiology, phylogeny and evolution of dermatophytes. Infection, Genetics and
Evolution Journal, 20, 336-351.
[6] Collee, J. G., Marmion, B. P., Fraser, A. G., & Simmon, A. (1996). Practical medical
microbiology (14th Ed.). Singapore: Longman Singapore Publishers (pte) Ltd.
[7] Davison, F. D., Mackenzie, D.W. R., & Owen, R. J. (1980). Deoxyribonucleic acid
base compositions of dermatophytes. J. Gen Microbial, 118, 465-470.
[8] Elavarashi, E., Kindo, A. J., & Kalyani, J. (2013). Optimization of PCR RFLP
directly from the skins and nails in Cases of Dermatophytosis, Targeting the ITS and
the 18S Ribosomal DNA regions. J Clin Diagn Res, 7 (4), 646-651.
[9] Enemuor, S. C., & Amedu, A. S. (2009). Prevalence of superficial mycoses in
primary school children in Anyigba, Kogi State, Nigeria. African J. Microbiol Resch,
3 (2), 62-65.
[10] Ghojoghi, A., Falahati, M., Paghehm A. S., Abastabar, M., Ghasemi, Z., Ansari, S.,
Farahyar, S., & Roudbary, M. (2015). Molecular identification of epidemiological
aspect of Dermatophytosis in Tehran, Iran. J. Research in Molecular Medicine, 3, 11-
16.
[11] Graser, Y., El Fari, M., Presber, W., Sterry, W., & Tietz, H. J. (1998). Identification
of common dermatophytes (Trichophyton, Microsporum, Epidermophyton) using
polymerase chain reactions. Br. J. Dermatol, 138, 576-582.
[12] Hanumanthappa, S. B., Sarojini, S. B., Shilpashree, S. B., & Muddapur, S. B. (2012).
Clinicomycological study of 150 cases of dermatophytosis in a tertiary care hospital
in South India. Indian J Dermatol, 57, 322–3.
[13] Hay, R. J. (1999). The Achilles project: How common are fungal foot in Europe.
Satellite symposium. Denmark: English Congress of European Academy of
Dermatology and Venereology (EADV).
[14] Hayettel, M. P., & Sachelil, R. (2015). Dermatophytosis, trends in epidemiology and
diagnostic approach. Curr Fungal Infect Rep, 9, 164-179.
[15] Jackson, C. J., Barton, R. C., & Evans, G. V. (1999). Species identification and strain
differentiation of Dermatophyte fungi by analysis of Ribosomal- DNA interginic
spacer regions. Journal of Clinical Microbiology, 37 (4), 931-936.
[16] Kane, J., Summerbell, R. C., Sigler, L., Krajden, S., & Land, G. (1997). Laboratory
handbook of dermatophytes. Belmont CA: Star Piblishers.
[17] Li, H. C., Bouchara, J. P., Hsu, M. M., Barton, R., Su, S., & Chang, T. C. (2008).
Identification of dermatophytes by sequence analysis of the of the rDNA gene internal
transcribed spacer regions. J. Med Microbial, 57, 592-600.
[18] Matehkolaei, A. R., Makimura, K., & Shidfar, M. R. (2012). Use of single-enzyme
PCR- restriction digestion barcode targeting the internal transcribed spacer (ITS
rDNA) to identify dermatophyte species. Iran J. Publ. Health, 41 (3), 82-94.
[19] Milne, L. J. R. (1996). Fungi. In Practical microbiology by Collee, J. G., Marmion, B.
P., Frasser, A. G., & Simmons, A. (14th Ed.). Singapore: Longmen Singapore
Publishers Ltd.
[20] Mochizuki, T., Ishizaki, H., Barton, R. C., Moore, M. K., Jackson, C. J., Kelly, S. L.,
& Evans, G. V. (2003). Restriction fragment length Polymorphism analysis of
Ribosomal DNA Intergenic regions is useful for differentiating strains of
Trichophyton mentagrophytes. Journal of Clinical Microbiology, 41 (10), 4538-4588.
[21] Nenoff, P., & Haustein, U. F. (1997). Tinea corporis due to Trichophyton tonsurans
Malmsten-report of a patient from Zaire. J. Mycosis, 40 (3-4), 127-129.
[22] Robson, J. (2012). Focus on Dermatophytes. Queensland: Sullivan Nicolaides
Pathology, University of Queensland.
[23] Salman, Z. A., Hashim, A. J., & Karhoot, J. M. (2013). Identification of Trichophyton
rubrum isolates by using traditional methods and RAPD-PCR. Iraq: University of
Baghdad.
[24] Sarika, G., Purva, A., Rahul, R., & Saksham. (2014). Prevalence of Dermatophytic
infection and determining sensitivity of diagnostic procedures. International Journal
of Pharmacy and Pharmaceutical Sciences, 6 (3), 35-38.
[25] Sharma, R., & Swati. (2012). Effect of Keratin Substrates on the growth of
Keratinophilic fungi. Journal of Academia and Industrial Research, 1, 170-192.
[26] Singh, S., & Beena, P. M. (2003). Profile of Dermatophytes infections in Baroda.
Indian Journal of Dermatology, Venereology and Leprology, 69, 281-283.
[27] Weitzman, I., & Summerbell, R. (1995). The dermatophytes. Microbiology Review, 8
(2), 240-259.
View publication stats

Molecular Identification of Dermatophytes Among Clinical Isolates

Uploaded by

Copyright:

Available Formats

Molecular Identification of Dermatophytes Among Clinical Isolates

Uploaded by

Document Information

Original Title

Copyright

Available Formats

Share this document

Share or Embed Document

Sharing Options

Did you find this document useful?

Is this content inappropriate?

Copyright:

Available Formats

Molecular Identification of Dermatophytes Among Clinical Isolates

Uploaded by

Copyright:

Available Formats

See discussions, stats, and author profiles for this publication at: https://www.researchgate.

MOLECULAR IDENTIFICATION OF DERMATOPHYTES AMONG CLINICAL

Article · June 2018

The user has requested enhancement of the downloaded file.

MOLECULAR IDENTIFICATION OF DERMATOPHYTES AMONG

MATERIALS AND METHODS:

Total Number Direct Examination by KOH Laboratory Culture

N. Isolates Number of isolates %

Figure 2: microscopic morphology of dermatophytes showing micro and macroconidia (40X). A) 1)

View publication stats

You might also like

Pfad - The Proxy pFad of © 2024 Garber Painting. All rights reserved.