Pathotyping and virotyping

Pathotyping

The initial determination of the presence of pathogenic E. coli in a sample is based primarily on the detection of appropriate virulence factors by the PCR technique. Appropriate samples may be intestinal samples or fecal swabs from animals with diarrhea, extraintestinal tissues from animals with septicemia or localized infections, water, soil, effluent, sludge, or organic wastes, from the environment, or contaminated foods. These samples are tested directly following an overnight enrichment. E. coli cultures may also be submitted for testing and pathotype identification. In samples from sick animals, we will test for the appropriate virulence factors to enable us to identify the pathotypes most commonly associated with the animal species and disease symptoms (see Tables in sections Pathogenic E. coli - Diseases and Virulence factors and serogroups). For environmental samples, if the source of the potential contamination is known, we can test for the virulence factors associated with the suspected animal species. If the source of the potential contamination is not known, or if you wish to eliminate the possibility of pathogenic E. coli contamination, we can test for a larger number of virulence factors, representing all of the known pathogenic E. coli. Please contact us for further information.

Suggested virulence factors to test and prices for pathotyping
Animal species	Disease syndrome	Virulence factor	Price (CAN $)*
			1st sample	Each add. sample
Any	Diarrhea	STa, STb, LT, Eae, Stx1(VT1) and Stx2(VT2)	40 $	35 $
	Extraintestinal samples	STa, STb, LT, Eae, Stx1(VT1), Stx2(VT2), CNF, P fimbriae, Aerobactin and Tsh (Tsh for avian samples)	60 $	55 $
Pig	Diarrhea	STa, STb, LT, Eae and F4(K88)	35 $	30 $
	Edema disease	Stx2e(VT2e), F18	15 $	13 $
Bovine (calf)	Diarrhea or hemorrhagic dysentery	STa, Eae, Stx1(VT1) and Stx2(VT2)	35 $	30 $
Poultry	Colisepticemia or cellulitis	Aerobactin, P fimbriae and Tsh	30 $	25 $
Dog and cat	Diarrhea	STa, Eae, CNF and P fimbriae	35 $	30 $
	Urinary tract disease	CNF, P fimbriae and Aerobactin	30 $	25 $
Rabbit	Diarrhea	Eae	10 $	8 $
Human	Diarrhea	STa, LT, Eae, Stx1(VT1) and Stx2(VT2)	40 $	35 $
	Urinary tract disease and septicemia	CNF, P fimbriae, Aerobactin and Hemolysin	35 $	30 $
		Additional fimbrial adhsins or other factors	8 $	6 $

* Prices subject to change

Turn-around time is 1-3 working days.

If you are submitting isolates from a country other than Canada, you will require a permit for entry of the isolate(s) into Canada. The application for this permit will be made by the EcL, based on the information obtained from you, and is usually received within a week of the application, at a cost of 35 CAN $* for a single entry or 60 CAN $* for multiple entries for one year, in both cases, irrespective of the number of isolates. The cost for obtaining this permit will be charged to the client. Please contact our technical service for more information.

A brief history (clinical signs, animal species, age, sex, origin of the sample) would help us in the choice of virulence factors to test, interpretation of the results, and for epidemiological purposes.

Please do not hesitate to contact us for further information or to discuss the possibility of special pricing if you wish to test a larger number of samples or virulence factors in herd problems, in vaccine control tests, etc.

* Prices subject to change

Virotyping

If an E. coli strain or isolate is submitted to the EcL, or we isolate individual colonies from a sample that has been submitted to us, we can test the strain or isolate for different virulence factors, and thus determine its virotype. We may test for the appropriate virulence factors by PCR or colony hybridization, depending on the number of virulence factors and isolates and, in certain cases, on the specific test required. The standard reference test is the PCR, which will be used for the definitive virotyping of strains. However, when a large number of virulence factors and isolates are to be tested, it may be more economical and hence less costly to use colony hybridization. This option would be more attractive for larger epidemiological surveillance studies.

Virotyping
Price	For PCR, the price will depend on whether Multiplex or simple PCR is used for the particular factor: 7 $* per virulence factor for the first sample 5 $* per virulence factor for additional sample
Turn-around	1-3 working days

* Prices subject to change (CAN $)

Choice of virulence factors to be tested

In routine diagnostic testing, we use colony hybridization and test for the following 20 virulence factors, at a price of 60 CAN $* per sample:

Toxins: LT, STa, STb, Stx1(VT1), Stx2(VT2), CNF, EAST1, Aerobactin, Tsh Fimbriae/adhesins: F4(K88), F5(K99), F6(987P), F41, P, AFA, F17, F18ab/ac(F107), Eae, Paa, AIDA

* Price subject to change

This will enable us to identify the virotypes most commonly associated with the most common animal species and disease symptoms (see Tables in sections Pathogenic E. coli - Diseases and Virulence factors and serogroups). For isolates from contaminated environmental or other samples, if the source of the potential contamination is known, we can test for the virulence factors associated with the suspected animal species. If the source of the potential contamination is not known, or if you wish to eliminate the possibility of pathogenic E. coli contamination, we can test for a larger number of virulence factors, representing all of the known pathogenic E. coli.

The following is a table showing pathotypes and associated virulence factors which would encompass most of the known pathogenic E. coli.

We recommend testing for the virulence factors listed in this table for complete virotyping or virulence factor analysis. Please contact us for further information.

Virulence factors commonly associated with the various pathotypes of E coli
Category of pathogenic E. coli		Virulence factor
CDEC	cell-detaching E. coli	Cytolethal distending toxins: CDT3, CDT4 Cytotoxic necrotizing factors: CNF1, CNF2
DAEC	diffuse adherence E. coli	Adhesins involved in diffuse adherence: AIDA, daaE
ETEC	enterotoxigenic E. coli	Adhesion factors shared by human ETEC: CFAI, CFAII(CS1coo), CFAII(CS3cst) Adhesion factors shared by porcine and bovine: F4(K88), F5(K99), F18, F41 Enterotoxins: heat-labile toxin (LT), human heat-stable toxin (STaH), porcine heat-stable toxin (STaP), heat-stable toxin type II (STb)
EPEC	enteropathogenic E. coli	Attaching and effacing factor: Eae
EHEC	enterohemorragic E. coli	Shiga-toxins: Stx1, Stx2, VT2vpl, VT2vh Flagellar antigen: FliC
EaggEC	enteroaggregative E. coli	Adhesion factors: aggR, AAF/1 Toxine: EAST1
EHEC EPEC	enterohemorragic E. coli enteropathogenic E. coli	Attaching and effacing gene: Paa
EHEC UPEC	enterohemorragic E.coli uropathogenic E. coli	Hemolysins: HlyA, HlyC, Ehx
SEPEC MENEC UPEC	septicemic E. coli meningitis associated E. coli uropathogenic E. coli	Aerobactin: iuc
SEPEC UPEC	septicemic E. coli uropathogenic E. coli	Adhesion factors: PapC, PapG
UPEC	uropathogenic E. coli	Adhesion factors: AFA, SFA, Type 1(FimA)
ExPEC	extraintestinal E. coli	Adhesion factor: CS31a Autotransporter: Tsh

Subtyping of certain fimbrial adhesins can also be done by PCR. For example, for F4(K88), we can identify the subtypes ab, ac, and ad. For P fimbriae, we can identify the subtypes F7 to F16, and F165. Please contact us for more information.

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