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Virulence Testing by PCR

Determine the presence of 25 virulence genes in E. coli to determine what type of pathogenic strains they are.

Presence of genes encoding for different virulence factors are tested by PCR using the protocols published in the following paper.

DebRoy, C. and Maddox, C. 2001. Assessing virulence of Escherichia coli isolates of veterinary significance. Animal Health Research Reviews 1: 129-140.

Detection of different pathogenic types of E. coli

  • Enterohemorrhagic E. coli (EHEC) or Shiga-toxin producing E. coli (STEC)
  • Enteropathogenic E. coli (EPEC)
  • Enterotoxigenic E. coli (ETEC)
  • Enteroinvasive E. coli (EIEC)
  • Enteroaggregative E. coli (EAEC)
  • Necrotoxigenic E. coli (NTEC)
  • Extraintestinal Pathogenic E. coli (ExPEC)

Detection of Toxins associated with E. coli

  • STa: Associated with cultures from bovine
  • STb: Mostly associated with cultures from porcine
  • LT: Mostly associated with cultures from porcine
  • Stx1: Associated with EHEC and STEC strains
  • Stx2: Associated with EHEC and STEC strains
  • CNF1: Associated with  pathogenic strains
  • CNF2: Associated with pathogenic strains

Detection of fimbriae and adhesin genes associated with E. coli

 

  •  F4 (K88): commonly associated with ETEC isolates from porcine.
  • F5 (K99): associated with ETEC strains from porcine, bovine, and sheep.
  • F6 (987P): associated with strains from porcine.
  • F41: associated with  strains isolated from porcine, bovine, and sheep.
  • F18 (F107): associated with strains from porcine edema disease and postweaning diarrhea.
  • F1845: associated with strains from bovine.
  • CS31A: associated with septicemic and enterotoxigenic strains found in bovine and recently isolated from diarrheagenic strains from humans.
  • Eae (intimin): associated with EHEC and EPEC strains.
  • BFP: bundle forming pilus associated with E. coli causing diarrheal diseases

Suggested PCR screening tests for detecting virulence factors in E. coli isolates from different host

Host LT STa STb Stx1 Stx2 CNF1 CNF2 EAE K88 K99
Bovine   X   X X X X X   X
Porcine X X X X X X X X X X
Equine X X X X X X X X    
Canine X X X X X X X X    
Feline X X X X X X X X    
Human X X X X X X X X    
Avian       X X X X X    

 

Host 987P F107 CS31A F1845 BFP Iss IutA IiroN HlyF OmpT
Bovine     X X            
Porcine X X






 
Equine








 
Canine








 
Feline








 
Human



X



 
Avian



X X X X X X

Detection of genes encoding virulence factors: toxins

Toxins:

  • Shiga toxins 1 and 2: Stx1 and Stx2
  • Heat Stable toxins a and b: STa and STb
  • Heat-labile toxin: LT
  • Cytotoxic Necrotizing Factors 1 and 2: CNF-1 and CNF-2
  • Hemolysins: HlyA, HlyD, HlyE

Detection of genes encoding virulence factors: fimbriae/adhesins

Fimbriae:

  • K88 (F4)
  • K99 (F5)
  • 987P (F6)
  • F107 (F18)
  • F1845
  • CS31a
  • BFP
  • EAE
  • TIR

Detection of genes encoding virulence factors for avian sp.and ExPEC

Avian Specific:

  • Iss
  • IutA
  • IroN
  • HlyF
  • OmpT
  • Adhesins associated with ExPEC