The Use of PCR to Aid in the Rapid Identification of Enterovirulent Escherichia coli in Shrimp Farms

Abstract

Polymerase Chain Reaction (PCR) technique allows the detection of the most enterovirulent Escherichia coli strains: enterotoxigenic (ETEC), enteroinvasive (EIEC), enterohaemorrhagic (EHEC) and enteropathogenic (EPEC). The prospective shrimp sector is facing a threat of disease. However, the study on the virulence of E. coli in shrimp habitat is still scarce. The aim of the present study was to detect enterovirulent strains of E. coli in the ten different shrimp farms which was categorized on the basis of unhygienic (UF₁, UF₂, UF₃ UF₄, UF₅and UF₆) and hygienic (HF₁, HF₂, HF₃and HF₄) condition of farms in Kalapara, Patuakhali, Bangladesh. Water, sediment and shrimp samples were examined from each farm. Six pairs of oligonucleotide primers were used for the amplification and thereby detection of target genes viz. the alkaline phosphatase (Pho-A), the heat-labile toxin1 (LT1), heat-labile toxin2 (LT2), heat-stable toxin1 (ST1), verotoxin (VT) and attachment and effacement (EAE) virulence genes. Finally, PCR products were analyzed and detection of different strains was confirmed by agarose gel electrophoresis and restriction enzyme analysis with AluI enzyme. The results revealed the presence of EPEC groups were common in most of the samples of shrimp farms having unhygienic condition. EPEC was present in water of UF₁ and UF₆. Sediment of UF₂ and shrimp of UF₆ showed EPEC positive. Whereas ETEC groups were found in shrimp of UF₂ and UF₆ and also water and sediment of farm UF₅. E.coli was absent in UF₃ and harmless E.coli was present in UF₄ .In case of farms in hygienic condition, EEC groups was absent in HF₂, HF₃ and HF₄ where harmless E. coli was observed in water and shrimp of HF₁. These findings of the study suggest that lack of enough sanitation system, polluted water source, unhygienic condition as well as poor management practices might be responsible for exposure of enterovirulent E. coli strains in this area. This knowledge will help in production of E.coli free shrimp and rapid detection of E.coli polluted shrimp in processing industry.