wildlife and zoo assay data sheet
- Qualitative detection of Clostridium difficile bacteria by polymerase chain
reaction. Assay also detects and differentiates C. dif
toxin-producing genes A and B.
Ultrasensitive qualitative detection of
Clostridium perfringens alpha toxin and enterotoxin by real time PCR
B0043 - Ultrasensitive qualitative detection of
Clostridium piliforme (Tyzzer's
disease) bacteria by real time PCR
detection but not differentiation
of several common Clostridium species, including C. difficile, C.
piliforme and C. perfringens,
chain reaction. Assay DOES NOT detect Clostridium botulinum.
is a gram positive, anaerobic, spore forming motile rod bacterium that
commonly inhabits the intestinal tract of many mammalian species,
reptiles and birds. It is also found in the environment. The bacterium
is a highly diverse organism, with more than 400 unique types, and has
several virulence factors. Exotoxin A and B are the most significant
factors, and bacterial production of exotoxins is correlated with
pathogenicity of individual strains of
C. difficile. Toxin A is
an enterotoxin, promoting fluid exudation from the intestinal mucosa,
and acts synergistically with the cytotoxic toxin B through attachment
to specific receptors on the surface of enterocytes. The combined
action of these toxins results in necrosis of superficial epithelium
and edema in affected areas of intestine.
The organism is an
important cause of enteric disease in laboratory rodents and horses.
Hamsters, guinea pigs and mice may be affected by pseudomembranous
colitis induced by antimicrobial therapy. In neonatal foals,
C. difficile has been
associated with hemorrhagic necrotizing enterocolitis and diarrhea.
The lack of an established intestinal microflora may make foals more
susceptible to colonization by this bacterium. Adult horses may
develop typhlocolitis and outbreaks of nosocomially acquired diarrhea
have been reported (Donaldson and Palmer, 1999; Madewell et al., 1995;
Perrin et al., 1993).
has also recently been implicated as a cause of typhlocolitis in
nursing piglets, chronic diarrhea in dogs and enterotoxemia in
normal patients, an established intestinal microflora is thought to
competitively prevent proliferation of
C. difficile and
subsequent toxin attachment. Alteration of intestinal microbial
balance with antibiotic use and increased exposure to the organism in
a hospital setting allows C.
difficile to colonize the gut in susceptible individuals.
of C. difficile is not
highly sensitive and does not differentiate the pathogenic and
non-pathogenic strains. Specific tests for
C. difficile toxins used
in the diagnostic laboratory include cell culture, which relies on the
presence of biologically active toxin, and an ELISA assay which
detects immunologically active toxin that may or may not be
PCR detection of
C. difficile is highly
sensitive and can discriminate between toxigenic and nontoxigenic
strains of the organism by detecting its toxin producing genes.
perfringens is a Gram-positive, rod-shaped, anaerobic, spore-forming bacterium found
as a normal component of decaying vegetation, marine sediment, the
intestinal tract of humans and other vertebrates, insects and soil.
Infections due to C. perfringens
can result in tissue necrosis, bacteremia, emphysematous cholecystitis
and gas gangrene. The bacteria can secrete α-toxin which results in
gangrene formation. If patients ingest the bacteria, colic, diarrhea
and sometimes nausea can result.
Food poisoning due to C. perfringens
bacteria is one of the common causes of food-borne illness. Poorly
prepared meat and poultry are commonly the sources of food poisoning.
The enterotoxin (CPE)
secreted by the bacteria, which mediates the food poisoning, is
heat-resistant and cannot be destroyed easily. Furthermore, the
bacteria themselves form spores that can withstand cooking
temperatures. If these spores are then left at room temperature,
germination may begin and infective bacterial colonies develop.
Generally, the incubation time of these spores is 6 to 24 (commonly 10
to 12) hours after ingestion of contaminated food. Since meat and
poultry are often prepared in advance of consumption, this allows good
opportunities for the spores to germinate.
People ingesting these bacteria can develop abdominal cramping and diarrhea.
Vomiting and fever are unusual. Illness usually resolves within 24
hours. It is also possible that many cases of C. perfringens
food poisoning remain subclinical, as antibodies to the toxin are
common among humans. This has led to the conclusion that most of the
population has experienced food poisoning due to C. perfringens.
Detection of C. perfringens by culture is slow
and not very sensitive. PCR detection is the method of choice for
rapid, sensitive and specific detection of this pathogen (Abubakar,
Infection with Clostridium
piliforme results in Tyzzer’s disease, which is
characterized by necrotic lesions in the liver, digestive organs and
heart. A number of animal species are susceptible to this organism,
including mice, rats, rabbits, dogs, cats, primates, and horses.
The organism is an
obligate gram-negative bacteria found in necrotic foci in spore forms.
Transmission is mainly through the fecal-oral route.
is a severe disease in many animal species, infected mice often do not
exhibit clinical symptoms. These mice become carriers of the disease
and spread the pathogen to other mice and other animal species.
Interestingly, different mouse strains differ in their susceptibility
to the pathogen (Waggie et al., 1981).
cannot be cultivated in artificial media, so diagnosis may be based on
microscopic examination of tissues, serological assays or steroid
challenge tests; these methods all require blood or necropsy samples.
When steroid challenge assays are performed, extreme care must be
taken to avoid spreading the pathogen. Moreover, microscopic
examination, serology and steroid challenge all suffer from a lack of
sensitivity and are labor intensive.
Detection of this
pathogen by polymerase chain reaction is highly sensitive and
specific. The test can be performed on fecal specimens rather than
blood or tissue, resulting in less trauma and risk to animals.
Help confirm the disease causing agent
Shorten the time required to confirm a clinical
diagnosis of Clostridium
infection to the species level
Help ensure that animal groups and
facilities are free
of these bacteria
Early prevention of spread of these bacteria
Minimize personnel exposure to these bacteria
Safety monitoring of biological products and vaccines
that derive from susceptible animals
I., Irvine, L., Aldus, C.F., Wyatt, G.M., Fordham, R., Schelenz, S.,
Shepstone, L., Howe, A., Peck, M. and Hunter, P.R. (2007) A systematic
review of the clinical, public health and cost-effectiveness of rapid
diagnostic tests for the detection and identification of bacterial
intestinal pathogens in faeces and food. Health. Technol. Assess.
Donaldson, M.T. and Palmer, J.E. (1999) Prevalence of
Clostridium perfringens enterotoxin and Clostridium difficile toxin A
in feces of horses with diarrhea and colic. J. Am. Vet. Med. Assoc.
Madewell, B.R., Tang, Y.J., Jang, S., Madigan, J.E.,
Hirsh, D.C., Gumerlock, P.H. and Silva, J. (1995) Apparent outbreaks
of Clostridium difficile associated diarrhea in horses in a veterinary
medical teaching hospital. J. Vet. Diagn. Invest. 7:343 346.
Perrin, J., Cosmetatos, I., Gallusser, A., Lobsiger, L., Straub, R.
and Nicolet J. (1993) Clostridium difficile associated with
typhlocolitis in an adult horse. J. Vet. Diagn. Invest. 5:99 101.
Waggie, K.S., Hansen, C.T. Ganaway, J.R. and Spencer, T.S.
(1981) A study of mouse strains susceptibility to Bacillus piliformis
(Tyzzer's disease): the association of B-cell function and resistance.
Lab. Anim. Sci. 31:139-142
Rectal swab, or 0.2 ml feces, or food swab, or
lesion swab, or environmental surface swab.
if advice is needed to determine an appropriate specimen type
for a specific diagnostic application. For specimen
types not listed here, please contact Zoologix to confirm
specimen acceptability and shipping instructions.
For all specimen
types, if there will be a delay in shipping, or during very warm
weather, refrigerate specimens until shipped and ship with a cold pack
unless more stringent shipping requirements are specified. Frozen
specimens should be shipped so as to remain frozen in transit. See
shipping instructions for more
2 business days