Detection and characterization of E. coli O157:H7 and induced shiga toxin-2 coding bacteriophages

Abstract

Escherichia coli 0157:H7 is classified as a member of the enterohaemorrhagic E. coli (EHEC) family. These organisms are responsible for a variety of clinical manifestations ranging from non-bloody diarrhoea to gross bloody diarrhoea with complications that include haemorrhagic colitis (HC), haemolytic uraemic syndrome (HUS) and thrombotic thrombocytopaenic purpura (TTP). Infection occurs by the ingestion of faecally contaminated food products, water sources and through person-to-person contact. Outbreaks of E. coli O157:H7 have been reported worldwide although most outbreaks seem to be from countries in the Northern hemisphere. Very little information is available on the prevalence of E. coli O157:H7 in South Africa. The only data available on E. coli O157:H7 were from a 1992 outbreak in Swaziland with some cases spreading to the adjacent provinces of South Africa. Selective methods were assessed and optimised to identify and isolate E. coli O157:H7 from food, water and faeces. These methods included culture techniques, immunomagnetic separation, immunoassays and molecular confirmation techniques. The methods optimised and assessed in this study proved to be suitable for the detection and isolation of E. coli O157:H7 from environmental water, food and faecal samples. In addition to the isolation of E. coli O157:H7 from these sources, methods were also optimised for the characterisation of E. coli O157:H7 using repetitive sequence analysis and induction of shiga toxin (Stx)¬converting phages which carry the genes coding for Stx from strains of E. coli O157:H7. The prevalence of E. coli O157:H7 in human-, bovine- and porcine faeces, sewage and recreational waters was investigated in a selected region of South Africa. Data suggested a low prevalence in sewage (0.76%), recreational waters (0%) and human faecal (0%) samples with a higher prevalence among carriers such as cattle (12.5%) and pigs (14.29%). UV-induced Stx-converting phages were examined and found to have different phage morphologies to the previously described lambdoid structure. In order to establish the host range susceptibility of these phages, all induced phages were subjected to conditions favourable for infecting E. coli O157:H7, non-O157 E. coli and other members of the enterobacteriaceae family including Salmonella, Shigella, Enterobacter, Klebsiella, and Proteus. These results have shown that Stx-phages were able to infect Salmonella cholerasuis and produce infectious progeny from these strains. Stx-converting phages propagated in Salmonella cholerasuis were able to re-infect strains of E. coli O157:H7. This study has shown that IMS in combination with molecular techniques was a sensitive tool for the isolation, identification and characterisation of E. coli O157:H7 from different sources. Results indicated that Stx-phages induced from E. coli O157:H7 demonstrated lambdoid structure as well as phages with long hexagonal heads and long non-contractile tails.