avian & livestock 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
(Tyzzer's disease) bacteria by real time PCR
Qualitative detection but not
differentiation of several
common Clostridium species, including C. difficile, C. piliforme
and C. perfringens,
polymerase chain reaction. Assay DOES NOT detect Clostridium
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.
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
clinically 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.
culture 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 biologically active.
detection of C. difficile
is highly sensitive and can discriminate between toxigenic and
nontoxigenic strains of the organism by detecting its toxin
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
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
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,
is an obligate gram-negative bacteria found in necrotic foci in
spore forms. Transmission is mainly through the fecal-oral
Tyzzer’s 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.,
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
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
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 flocks or animal facilities are free of
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 and birds
Abubakar, 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. 11:1-216.
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. 215:358 361.
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.
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, cloacal swab, or 0.2 ml feces, or
food swab, or lesion swab, or environmental surface swab.
Contact Zoologix 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.
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 information.
2 business days