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Association between Key-Gaskell syndrome

and infection by Clostridium botulinum
type CID

F. NUNN, T. A. CAVE, C. KNOTTENBELT, 1. R. POXTON

There is growing evidence that equine dysautonomia is a toxicoinfection with Clostridium botulinum type C.
The possibility that feline dysautonomia has the same aetiology was investigated by attempting to detect
botulinum type C neurotoxin in the food, faeces and the contents of the ileum of affected cats, and by
serology. The toxin was detected directly in four of eight affected cats and after enrichment in seven of
them, and in their dried food. No toxin was detected in healthy control cats or in their tinned food. Recent
exposure to the organism was assessed by the detection of immunoglobulin A (IgA) in the faeces of
healthy control cats and affected cats. The levels of IgA antibodies to the toxin and to surface antigens of
C botulinum type C in the faeces of the affected cats 14 weeks after the outbreak were significantly higher
than in the faeces of the control cats.

FELINE dysautonomia is a disease characterised by extensive and C2 showed no clinical signs, but a fluoroscopic exami-
degeneration of the autonomic nervous system (Key and nation 96 days after the beginning of the outbreak showed
Gaskell 1982), and dysautonomia has also been reported in subjective evidence of reduced oesophageal function; they
horses, dogs, rabbits and hares. Most of the cases of feline, may have suffered a subclinical form of the disease. The cats
equine and leporine dysautonomia have been reported in the were fed a single brand of tinned food from a communal bowl
UK (Whitwell 1991, Whitwell and Needham 1996), whereas and a single brand of dry food from two communal hoppers.
most cases of canine dysautonomia have been reported in the At the time of the outbreak the batch of dry food had been
USA (Berghaus and others 2001). in use for six days before the index case. On average it took 10
In horses with grass sickness, C I neurotoxin from days for the cats to consume all the dried food contained in
Clostridium botulinum type C (BoNT/C) has been detected in the hoppers. Fresh tins of food were opened daily. The cats'
74 per cent of acute cases and 67 per cent of both subacute food and pattern of feeding had not changed during the pre-
and chronic cases, but in only 10 per cent of control horses ceding 12 months.
(Hunter and others 1999). During an outbreak of dysautono-
mia in cats, the same toxin was detected indirectly from ileal Control cats
samples from 11 of the 16 affected cats, and after enrichment Eleven cats over two years old were used as controls. They
from nine of them. The levels of specific immunoglobulin A were from the same geographical area (Glasgow) and had
(IgA) to both EDTA-extracted C botulinum surface antigens unrestricted access to the outdoors. They were owned by staff
and a BoNT/( -toxoid complex have also been found to be sig- from Glasgow Veterinary School, who considered that they
nificantly higher in horses with grass sickness than in con- were in good health; their dietary history was unknown.
Veterinary Record (2004) trol horses (Hunter 2001).
155,111-115 In November 2001, there was an outbreak of feline dys- Sample collection and storage
autonomia in a closed non-breeding colony of eight pet cats Faecal samples of 2 g from the cases and controls were
F. Nunn, BSc, (Cave and others 2001) which was investigated to test the collected in sterile universal containers immediately after
I. R. Poxton, BSc, PhD, hypothesis that the aetiology of feline dysautonomia might defecation and frozen at -70°C until processed. The cases
D)Sc, be similar to that of equine grass sickness, and that it is were sampled 14 weeks after the onset of clinical signs in the
Medical Microbiology, caused by toxicoinfection by C botulinum type C/D. The pro- index case.
Centre for Infectious cedures used to investigate equine grass sickness were The ileum and its contents were obtained from case
Diseases, College of applied: the detection of the toxin in food, faeces and gut A3 postmortem; the ileum was tied off proximally and
Medicine and Veterinary contents by ELISA; the isolation of C botulinum from food and distally with nylon suture, removed and frozen at -20°C until
Medicine, University of faeces; and the detection of specific IgA antibodies in the assayed.
Edinburgh, Teviot Place, faeces and ileal contents to clostridial surface antigens and A sample of the dry food fed to the cases at the start of the
Edinburgh EH8 9AG BoNT/C. outbreak was stored at -70°C, and unopened cans of the
T. A. Cave, BVSc, CertSAM, tinned food fed to them were stored at room temperature.
M RCVS,
C. Knottenbelt, BVSc, MATERIALS AND METHODS Sample preparation
MSc, DSAM, MRCVS, Preparation of faeces with protease inhibitors for the
Department of Veterinary Cases IgA ELISAS All reagents were stored at -20°C and kept on ice
Clinical Studies, Six of a colony of eight house-kept cats, none of which had during the procedure. Two parts protease inhibitor solution
University of Glasgow access to the outdoors, were clinically affected. The charac- (soybean trypsin inhibitor 1 mg/ml in phosphate-buffered
Veterinary School, teristics of the cases are shown in Table 1, and the clinical signs saline [PBS], 50mM EDTA containing 0-05 per cent Tween 20)
Bearsden Road, Bearsden, and outcomes are summarised in Table 2. The index case (Al) were added to one part weighed sample (approximately 1 g).
Glasgow G61 I QH was admitted to Glasgow University Veterinary School five Phenylmethylsulphonyl fluoride (PMSF; Sigma-Aldrich), 0. 1M
days after first showing clinical signs, and cats B1, B2 and Cl in ethanol, was added to a final concentration of 1mM. The
Mr Cave's current address were admitted five to eight days after first showing clinical mixture was vortexed for 30 to 60 seconds and then cen-
is Vale Reterrals, signs. Cats Al and A2 died and A3 was euthanased in trifuged at 3800 g for 10 minutes. The supernatant was
The Animal Hospital, extremis. Only Al and A3 were examined postmortem, and removed and PMSF was added to a final concentration of 1 per
Stinchcombe, Dursley, dysautonomia was diagnosed histopathologically. Clinical cent (v/v). After mixing well, the mixture was allowed to stand
Gloucestershire GL 1I 6AJ signs persisted in the surviving cats (B1, B2 and C1). Cats B3 for 15 minutes on ice. Heat-inactivated fetal calf serum (FCS)

The Veterinary Record, July 24, 2004 ill

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PAPERS & ARTICLES

was added to a final concentration of 4 per cent (v/v) and the

mixture was centrifuged at 15,700 g for five minutes. The Group Cat
supernatant was removed and stored at -700C. Relationship Breed Age (years) Sex
A 1 Same parents, same litter Birman 5 M (N)
Preparation of ileal tissue with saponin for the IgA 2 5 F(E)
ELISAS Antibody was extracted from the ileum with saponin 3 5 F(E)
as described by Bergquist and others (2000); 1 g of frozen B 1 Same parents, different litters Birman 9 M (N)
2 14 F (N)
tissue was removed aseptically and thawed for eight hours 3 Shared queen with Bl and B2
at 4°C in 2 ml PBS containing 2 per cent saponin (w/v), but different tom and litter 14 F (N)
1 mg/ml soybean protease trypsin inhibitor, 50mM EDTA, 0-05 C 1 Same parents, same litter Colourpoint 11 M (N)
2 11 M (N)
per cent Tween 20, 2mM PMSF, 0-2 mg/ml sodium azide
and 4 per cent FCS (v/v). After thawing, the samples were agi- M Male, F Female, N Neutered, E Entire
tated with sterile forceps and then vortexed for 30 seconds.
The samples were centrifuged at 15,000 g for five minutes and
the supernatants were collected and stored at -700C until Samples were diluted 1 in 4 with PBS-TG before the addi-
used. tion of 100 Il/well in quadruplicate. They were incubated
overnight at room temperature and then washed four times
Preparation of faeces and food samples for the ELISA in ELISA wash buffer.
for BoNT/C and bacteriology For non-enriched samples, Rabbit anti-cat IgA (Nordic Immunologicals) was diluted 1
approximately 1 g of the sample was added to 10 ml of PBS, in 400 with PBS-TG and 100 1d was added to each well. The plates
pH 7 2, containing 0-2 per cent gelatin (PBS-G), and the sam- were incubated for three hours at 37°C while being shaken, and
ples were incubated overnight at 40C. were then washed four times with ELISA wash buffer.
For enriched samples, approximately 1 to 2 g was added Anti-rabbit IgA conjugated to alkaline phosphatase
to 15 ml of prereduced NZ-CASE-CMB medium (Hunter and (Sigma) was diluted 1 in 2000 with PBS-TG and 100 pi was
others 1999) and incubated anaerobically at 300C for added to each well. The plates were incubated for three hours
five days. After vortexing thoroughly, the samples were at 37°C while being shaken, and were then washed four times
centrifuged at 3800 g for 20 minutes and the supernatant was with ELISA wash buffer.
collected and stored at -200C. Alkaline phosphatase substrate tablets (104-105 phos-
phatase tablets, p-nitrophenyl phosphate 5 mg/ml; Sigma)
EDTA extraction of C botulinum surface antigens Cul- were diluted 1 in 10,000 with substrate solvent (0-05M
tures of C botulinum (20 ml) were prepared in Fastidious sodium carbonate solution, pH 9-8, with 1mM magnesium
Anaerobe Broth (Lab M) and incubated anaerobically at 300C chloride) to give a concentration of 1 mg/ml and 100 Id of this
for 18 hours. The strain used was a toxin-negative C botu- solution was added to each well. The plates were then incu-
linum type C (NCTC 3732). EDTA extracts were prepared as bated at room temperature for 30 minutes and read with an
described by Poxton (1984) and stored at -200C, and their Anthos plate reader at 405 nm, referenced at 620 nm.
protein concentration was determined as described by Lowry
and others (1951). Bacteriological isolation of organisms from
faeces and food
ELISA detection of anti-BoNT/C toxoid IgA and Tenfold dilutions (10-l to 10-4) were made from the non-
anti-C botulinum surface antigen IgA from faeces enriched and enriched samples in pre-reduced nutrient broth
and ileal tissue after 24 and 48 hours, and plated on to fastidious anaerobe
Microwell plates (Nunc; Fisher Scientific) were coated with agar (FAA) (Lab M) containing 5 per cent egg yolk emulsion
BoNT/C toxoid (Metabiologicals) at 5 [ig/ml or surface antigens (Oxoid) and 10 [ig/ml gentamicin. The plates were incubated
at 30 [1g/ml diluted in coating buffer (0.05M sodium carbon- for up to five days and examined for lecithinase and lipase
ate buffer, pH 9 6, 0 02 per cent w/v sodium azide) and incu- activity. Likely colonies were re-streaked for purity and also
bated overnight at 4°C or at room temperature, respectively. incubated aerobically on Columbia blood agar (Oxoid CBA
The plates were washed four times with ELISA wash buffer base with 5 per cent defibrinated horse blood) to check for
(one PBS BR14a tablet/litre [Oxoid], 15mM sodium chloride, aerobic growth. The colonies were also examined by Gram
2mM potassium chloride, 0-05 per cent Tween 20, pH 7.3). stain and phase-contrast microscopy.
The plates were blocked with PBS containing 3 per cent
teleostean gelatin (Sigma) (PBS-TG) and 0-02 per cent sodium ELISA detection of BoNT/C toxin from cat food,
azide (200 p1 per well) and incubated while being shaken for faeces and ileum
four hours at 370C, before being washed four times with ELISA A polyvalent guinea pig antiserum (CAMR) raised against
wash buffer. They were stored in sealed polythene bags at purified BoNT/C was used as a capture antibody. Optimal dilu-
-20°C until used. tions of the reagents were determined by using a checker-

..0 0 *0~R IM -ciIMlIT-M -7

Regurgitation/ Ufinary Bilateral Histopathological
Cat Lethargy vomiting Appetite Dysphagia Constipation retention pupillary dilatation Outcome confirmation
Al + + Anorexic + + + + Died +
A2 + + Anorexic + + + + Died
A3 + + Anorexic + + + + Euthanased +
Bi + + Reduced - - - + Survived
B2 + + Anorexic - - + + Survived
B3 - + Normal - - - - Survived
C1 - + Normal - - - Survived
C2 - - Normal - - - - Survived
+ Present, - Absent

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(a)
2
Case Directly (ng/kg) After enrichment
Al 15.6 +
FIG 1: Box and whiskers A2 ND +
A3 2-1 +
plots of faecal BI ND +
immunoglobulin A (IgA) B2 0-8 +
expressed in terms of B3 ND ND
the optical density (OD) Cl ND +
in the ELISA against 0 C2 1-6 +
(a) botulinum type C Dried food ND +
neurotoxin toxoid ND Not detected, + Present
complex and (b) against
EDTA-extracted surface
antigens of a 0-
toxin-negative
C botulinum type C Affected cats Controls controls and to normalise the results in later assays. Negative
strain. The affected cats controls consisted of coated wells with no sample but
were sampled 14 weeks (b) with conjugate, and coated wells with sample but no con-
after the onset of jugate.
clinical signs (with the The results were analysed by using sPss software (spss
exception of the lowest Science) and the Mann-Whitney U test. Significance was set
value, which was from
an ileal sample from at the 5 per cent level.
1-
a cat which was
euthanased in the third 0
week of the outbreak). 0
RESULTS
The control group 0
consisted of 11 healthy, ELISA to detect IgA antibodies to BoNT/C toxoid
free-roaming cats. The and C botulinum surface antigens from faeces
affected group of cats and ileal tissue
had significantly higher 0
Specific IgA antibody to BoNT/C and surface antigens was
levels of faecal IgA than
the control group detected mainly in the affected cats which had significantly
(P<0-001 and P<0.005, I S higher mean specific IgA levels than the healthy control cats.
respectively) in both 0 - The lowest value in the affected cats (Fig 1) was observed in
assays Affected cats Controls the ileum of case A3, in a sample taken during the outbreak
(week 3) and not 14 weeks afterwards like the others.
The ELISA values for IgA antibody to BoNT/C toxoid in the
board titration. Microwell plates (Nunc) were coated with free-roaming control cats (median 0-23, range 0-6 to 0.03)
100 [d per well of the antiserum diluted 1 in 10,000 in 0-05M were significantly lower than in the surviving cases (median
sodium carbonate buffer, pH 9-6, 0-02 per cent w/v sodium 1-5, range 1-7 to 1-3) [P<0-00l]). This analysis excludes the
azide, and incubated overnight at 4°C. The plates were then ileum sample from A3, but if it is included the results are
washed three times with ELISA wash buffer. still significant (median 1-45, range 1-7 to 0-7 [P<0-001]).
They were then postcoated with PBS-TG (200 p1 per well) Similarly, the levels of IgA against EDTA-extracted surface anti-
for two hours at 370C in a shallow water bath, and washed gens were much lower in the control cats (median 0-23, range
three times in ELISA wash buffer. 0-85 to 0.06) than in the surviving affected cats (median 0-92,
The non-enriched samples were added (100 [I per well) range 1-35 to 0-81 [P<0-001 ]). Again, if the ileal sample from
either neat or diluted 1 in 2 and 1 in 4 in PBS-TG, and the A3 is included, the results are still significant (median 0-87,
enriched samples (100 pd per well) were added neat or diluted range 1-35 to 0-34 [P<0.05]).
1 in 5 and 1 in 25 in PBS-TG, and incubated for three hours
at 37°C in a shallow water bath, and then washed three times Bacteriological isolation of organisms from
in ELISA wash buffer. The guinea pig anti-BoNT/c horse- faeces and food
radish peroxidase conjugate (CAMR) was diluted 1 in 300 in On the basis of the colony morphology the faecal samples did
PBS-TG, added to the plates (100 pl per well) and incubated not yield any likely colonies, and there was no growth anaer-
for three hours in a shallow water bath. After washing three obically from the tinned cat food. Likely colonies did appear
times in PBS containing 0-05 per cent Tween 20, substrate in the cultures of the dry food, but subculture was unsuc-
solution (3,3', 5,5'-tetramethyl-benzidine dihydrochloride cessful despite several attempts.
tablets [Sigma] dissolved in phosphate citrate buffer, pH 5-0,
with 2 pl of 30 per cent hydrogen peroxide per 10 ml) was ELISA detection of BoNT/C toxin in cat food, faeces
added at 100 pl per well. The reaction was allowed to develop and ileum
at room temperature for up to 60 minutes and was stopped Toxin was detected without enrichment in faecal samples
by the addition of 50 pI of 2M sulphuric acid to each well. from three of the six clinical cases, and one of the two sub-
A standard curve of purified BoNT/C (CAMR) was run on each clinical cases (Cl). The index case (Al) had the highest fae-
plate. The limits of detection of this assay were 0-8 ng/ml cal concentration of toxin. No toxin was detected in the faeces
and a standard curve of purified neurotoxin with concen- of the control cats or in the cat food by this method. After
trations ranging from 200 ng to 0-8 ng was included on each enrichment, toxin was detected in faecal samples from all the
plate. clinical cases except B3, but none was detected in faeces from
the control cats. Toxin was detected in the dry cat food but
Controls and statistical analysis not in the tinned cat food. Table 3 summarises these results
No positive control sera were available, but positive signals and shows the amounts of toxin detected by the direct detec-
were identified in the initial assays and these were used as tion method.

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DISCUSSION tive, and there were only a small number of cats; a case-
control study would be useful, particularly of animals fed
There is evidence that equine dysautonomia, or grass sickness, similar diets.
is caused by a toxicoinfection with C botulinum group III phe- It is not known whether the gut stasis is caused directly
notype (serotype C/D) (Hunter and others 1999, Hunter by the production of the toxin. However, the detection of spe-
2001). cific IgA in gut samples from acute cases of grass sickness
C botulinum type C mainly affects animals and can pro- (Hunter 2001) indicates that the horses had been exposed to
duce three toxins: Cl (BoNT/C) is a bacteriophage-encoded the organism before the clinical signs developed, and that the
neurotoxin that inhibits the release of neurotransmitters by gut stasis may be an effect rather than a cause of the disease.
specifically proteolysing syntaxin (Schiavo and others 1995) The levels of specific serum IgG have been found to be
and SNAP-25 (Foran and others 1996) at cholinergic nerve significantly lower in horses with grass sickness than in
terminals; C2 and C3 toxins both have ADP-ribosylating healthy controls (Hunter and Poxton 2001), suggesting that
activity, and C2 can disrupt the ultrastructure of the weakly immune animals may be more susceptible to grass
cytoskeleton (Mauss and others 1990). C botulinum type D sickness.
is very similar to C botulinum type C and is distinguishable The results of this investigation provide strong circum-
only by the bacteriophage-encoded type D neurotoxin. stantial evidence of an association between feline dysautono-
Mosaic toxins consisting of type C and type D subunits also mia and C botulinum type C. The toxin and specific IgA were
occur (Morrishi and others 1989). Of these toxins, only detected in samples from the affected cats, and the toxin was
BoNT/C has been shown to be cytotoxic in vitro (Kurokawa also detected in samples of the dried food fed to the cats. Only
and others 1987) and could be the cause of the pathology a small number of samples from the affected and control cats
observed in dysautonomias. were available, but the results warrant further investigation of
It is very difficult to isolate the organism and to detect any new cases of feline dysautonomia.
the toxin. C botulinum is highly fastidious and the toxin is
encoded on an unstable bacteriophage which is easily lost
upon subculture. The presence of toxin after enrichment does ACKNOWLEDGEMENTS
not necessarily mean that toxin was present in vivo and the
absence of toxin does not necessarily mean that the organism The authors are most grateful to the cats' owners for their
was not there. These results provide evidence that there is considerable assistance in allowing the reassessment of the
an association between feline dysautonomia and a toxico- cases, and to Morag Lindon for originally referring them
infection with C botulinum type C similar to that reported to Glasgow University Veterinary School. They also thank
in equine grass sickness. Hunter and others (1999) detected Professor Os Jarrett for control samples and Matthew Golder,
toxin in healthy horses leading to the hypothesis that C botu- Companion Animal Diagnostics, Glasgow University, for
linum type C/D may be present in the gastrointestinal tract of arranging the collection of the plasma and faecal samples.
healthy horses. In these cats, the toxin was detected only in Robert Brown of the University of Edinburgh is thanked for
samples from the affected cats, and the samples containing his technical assistance. Francesca Nunn is funded by the
the highest concentrations of toxin were from the three most Equine Grass Sickness Fund. Tom Cave was funded by the
acutely affected cats (Al, A2 and A3); the one negative sam- RCVS Trust (Clarke & Sparrow) and Cats Protection.
ple came from the least affected cat (B3). The toxin assay is
highly specific and does not detect BoNT/D or any of the other
botulinum neurotoxins. References
Given the very different diets and gastrointestinal physi- BERGHAUS, R. D., O'BRIEN, D. P., JOHNSON, G. C. & THORNE, J. T. (200 1)
ology of horses and cats it is possible that cats kept indoors Risk factors for development of dysautonomia in dogs. Journal of the
are not normally exposed to C botulinum type C/D in their American Veterinary Medical Association 218, 1285-1290
canned food. The dried food was positive for toxin after BERGQUIST, C., MATTSSON-RYDBER, G. A., LONROTH, H. & SVENNER-
enrichment, and it is possible that contamination with cereal HOLM, A. (2000) Development of a new method for the determination of
or soil in dried food may harbour C botulinum (and other immune responses in the human stomach. Journal ofImmunological Methods
234, 51-59
clostridia) and provide an environment in the gut that allows CAVE, T. A., KNOTTENBELT, C., MELLOR, D. J., LIDDON, M. & REID, S. W.
them to proliferate. Dry food is heated only to 80°C during (2001) Feline dysautonomia in a closed colony of pet cats. Veterinary Record
processing, theoretically allowing C botulinum spores to sur- 149, 779
vive. Since the investigation, the dried and tinned foods of dif- FORAN, P., LAWRENCE, G. W., SHONE, C. C., FOSTER, K. A. & DOLLY,
ferent brands fed to three other cats with Key-Gaskell J. 0. (1996) Botulinum neurotoxin Cl cleaves both syntaxin and SNAP-25 in
syndrome have been assayed for toxin and all have been neg- intact and permeabilised chromaffin cells: correlation with its blockade of
ative. These cats were allowed access to the outdoors, and catecholamine release. Biochemistry 35, 2630-2636
Berghaus and others (2001) and Harkin and others (2002) HARKIN, K. R., ANDREWS, G. A. & NIETFIELD, J. C. (2002) Dysautonomia
have shown that free-roaming dogs, known to forage, are in dogs: 65 cases (1993-2000). Journal of the American Veterinary Medical
Association 220, 633-639
more likely to develop dysautonomia. HUNTER, L. C. (2001) The role of Clostridium botulinum type C in the patho-
Local mucosal immunity may be important in protecting genesis of equine grass sickness. PhD Thesis. Edinburgh, University of
animals against the disease, especially free-roaming cats that Edinburgh. pp 220-221
may be exposed to C botulinum type C/D through hunting. HUNTER, L. C., MILLER, J. K. & POXTON, I. R. (1999) The association of
However, in house-kept cats that are not naturally exposed to Clostridium botulinum type C with equine grass sickness: a toxicoinfection?
the organism, specific IgA may be a good indicator of their Equine Veterinary Journal 31, 492-499
exposure. The affected cats had detectable levels of specific HUNTER, L. C. & POXTON, I. R. (2001) Systemic antibodies to Clostridium
IgA whereas the healthy controls did not. The control cats botulinum type C- protection from grass sickness. Equine VeterinaryJournal
were free-roaming but only three of them showed a positive 33, 547-553
KEY, T. J. & GASKELL, C. J. (1982) Puzzling syndrome in cats associated with
signal to BoNT/C toxoid, compared with all of the affected cats. pupillary dilatation. Veterinary Record 1 10, 160
No dietary history was available for the control cats. No pos- KUROKAWA, Y., OGUMA, K., YOKOSAWA, N., SYOTO, B., FUKATSO, R. &
itive control for specific IgA is available and the concentration YAMASHITA, I. (1987) Binding and cytotoxic effects of Clostridium botu-
of IgA in the samples from the affected cats may have been linum type A, C I and E toxins in primary neuron cultures from foetal mouse
increased by dehydration, although they were not clinically brains. Journal of General Microbiology 133, 2647-2657
dehydrated when sampled. The assay used was not quantita- LOWRY, 0. H., ROSEBROUGH, N. J., FARR, A. L. & RANDALL, R. J. (1951)

11 4 The Veterinary Record, July 24, 2004

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Protein measurement with the Folin phenol reagent. Journal of Biological immunoassay and electroblot transfer. Journal of General Microbiology 130,
Chenmistry 193, 265-275 975-981
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perfringens iota toxin. European Joturnal ofBiochemistry 194, 237-241 bond within the carboxy-terminal region of syntaxins. Journal of Biological
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botulinum, C sporogenes anid C novyi by an enzyme linked immunosorbent dysautonomia confirmed. Veterinary Record 139, 323-324

Stabilisation of scoliosis in two koi

(Cyprinus carpio)
P. D. GOVETT, N. J. OLBY, D. J. MARCELLIN-LITTLE, D. S. ROTSTEIN, T. L. REYNOLDS,
G. A. LEWBART
Two koi (Cyprinus carpio) from the same pond developed similar lesions of scoliosis. Radiographic
examinations showed that their spines had become malaligned as a result of vertebral compression
fractures involving T14 to T16. The vertebrae in both fish were stabilised with screws, k-wire and
polymethylmethacrylate. They both appeared to improve after surgery, but they began to decline and
died within three months. A postmortem examination revealed multi-organ inflammation that was
not associated with the surgical implants.

Veterinary Record (2004) SCOLIOSIS, or 'bent-back', in koi (Cyprinus carpio) is being ing 0.5 mg/g enrofloxacin (Baytril 2-27 per cent; Bayer
155,115-119 reported more often. Many causes have been suggested, Corporation) at a dose of 10 mg enrofloxacin/kg/day. When
including vitamin C deficiency (John and others 1979, Halver there was no improvement, the paste food was instead
P. D. Govett, DVM, and Hardy 1994); electrocution as a result of either lightning impregnated with 2 mg/g trimethoprim-sulphadiazine
G. A. Lewbart, MS, VMI), strike (Barlow 1993), faulty submersible pumps (Johnson (Tribrissen; Schering-Plough) and fed to provide a dose of 30
DACZM, 1997) or electrofishing (Sharber and Carothers 1988); tryp- mg/kg/day, upon a veterinarian's recommendation.
Environmental Medicine tophan deficiency (Halver and Shanks 1960, Kloppel and Post
Consortium and 1975, Poston and Rumsey 1983, Walton and others 1984, Post Clinical and radiographic findings
Department of Clinical 1993); trauma; organophosphates (Couch and others 1977, The ulcer was hyperaemic but healing, and there was mild
Sciences, Alam and Maughan 1993, Waddington 1995); and bacterial scoliosis, with a left lateral deviation of the tail at the level of
N. J. Olby, VetMB, PhD, cold water disease (Noga 1996). Although some fish continue the anal fin.
DACVIM, to do well with a noticeable curvature of the spine, others A biopsy taken from the tip of the right pectoral fin
D. J. Marcellin-Little, become debilitated and intervention becomes necessary. This had no evidence of microscopic changes. A skin scrape taken
DEDV, DACVS, DECVS, paper describes the progression of the condition and the sur- from the ulcerative lesion contained a few unidentified non-
Department of Clinical gical stabilisation of scoliosis in two koi. pathogenic protozoal organisms and one free-living nema-
Sciences, tode. The fish was anaesthetised in 200 mg/litre of buffered
D. S. Rotstein, DVM, tricane methanesulphonate (MS222) (Finquel; Argent
MSPVM, DACVP, CASE HISTORY 1 Chemical Laboratories) for the purpose of radiographic
T. L. Reynolds, DVM, examination (Love and Lewbart 1997) and maintained by
Department of In July 2001, an approximately two-year-old, 56 cm, 1314 g, delivering the anaesthetic-containing water through a syringe
Population Health and female doitsu sanke koi developed a 3 cm chevron-shaped over its gills as needed. Dorsoventral radiographs revealed a
Pathobiology, College of ulcer, dorsal to its right pectoral fin. The fish shared a 56,775 right laterodorsal angulation involving the 14th, 15th and
Veterinary Medicine, litre pond, with a maximum depth of 137 cm, with 30 other 16th trunk vertebrae (T14 to T16) at the level of the caudal
North Carolina State koi and 10 goldfish (Carassius auratus). The water quality had swimbladder. The vertebrae at this site appeared to be
University, 4700 been good, but in the previous three months some of the fish osteopenic and the intervertebral spaces were not well delin-
Hillsborough Street, had developed ulcerative skin disease, and when this koi eated (Fig 1). Lateral views revealed a dorsolateral subluxa-
Raleigh, NC 27606, USA began separating itself from its school the ulcer was first tion of the spinal column just caudal to the last rib, involving
noticed. The koi had been treated by the owner with a series T 14 and T 15. The lateral angulation was considered to be due
Dr Rotstein's present of three injections of 5-7 mg/kg amikacin sulphate (Ami- to a combination of a fracture and a subluxation. Several cal-
address is Department of glyde-V, 50 mg/ml; Fort Dodge) administered intracoelomi- lused, mid-body fractures were apparent in the caudal ribs.
Pathobiology, University cally every 48 hours. When the injections were given, the ulcer The koi recovered uneventfully in fresh dechlorinated water;
of Tennessee, College of was cleansed with 10 per cent povidone-iodine solution it resumed swimming with the group of fish and the ulcer
Veterinary Medicine, 2407 (Betadine; Purdue Frederick) and then treated with a topical started to heal; it was treated with trimethoprim-sulpha-
River Drive, Knoxville, triple antibiotic ointment (Neosporin; Warner Lambert). All diazine for two more weeks, but no further treatment was
TN 37996, USA the fish in the pond were provided with paste food contain- considered necessary.

The Veterinary Record, July 24, 2004 11 5

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Association between Key-Gaskell syndrome

and infection by Clostridium botulinum type
CID
F. Nunn, I. R. Poxton, T. A. Cave and C. Knottenbelt

Veterinary Record 2004 155: 111-115

doi: 10.1136/vr.155.4.111

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