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Current evolutionary theory proposes that niche-adapted microbial populations might evolve through selection for favoured genotypes followed by clonal expansion (Maynard-Smith, 1991). Possible correlations between genomic variation and ecological niche in Escherichia coli isolates derived from human and animal sources were investigated by randomly amplified polymorphic DNA (RAPD) analysis. A 1.6-kb polymorphic marker was identified which was present in 60% of isolates from human clinical specimens but was present in less than 5% of isolates derived from ovine and bovine faeces. The marker maps to a region of the chromosome located immediately downstream from the gene encoding the glycine decarboxylase P-protein (gcvP). DNA sequences from marker-positive and marker-negative isolates exhibit an abrupt loss of homology immediately downstream from the transcription termination point of the gene which extends for at least 130-base pairs beyond the gcvP transcription terminator. Sequences spanning this region in marker-negative isolates exhibit similarity to the cognate sequence from E. coli K-12, while the corresponding region in marker-positive isolates bears no similarity to any other published sequence. The utility of the marker for investigating the occurrence of human-derived E. coli in the environment was studied in a rural stream. Although the stream carried a high background of animal-derived E. coli, the marker could only be detected in isolates obtained downstream of the human faecal input. The polymorphism therefore shows promise for identification of human-derived E. coli within environments containing isolates from multiple and diverse sources. The methods described here could be used to generate further markers suitable for investigating microbial population ecology.

A randomly amplified polymorphic DNA (RAPD) marker, localised to the glycine decarboxylase gene (gcvP) of Escherichia coli, has shown promise for use as a molecular marker for the identification E. coli isolates of human origin. Characterisation of the RAPD polymorphism has enabled development of target-specific primers for direct PCR detection of E. coli of human origin. A field trial was undertaken to investigate the suitability of the marker for detecting E. coli derived from sewage effluent discharged to a rural stream which already contained a substantial background level E. coli of animal origin. Multiplex PCR was performed on E. coli isolates from each sample site using the marker-specific primers in conjunction with primers directed towards a region of the β-glucuronidase (gusA) gene which served as an internal PCR control. Marker-positive strains were detected in raw sewage, treated effluent, stream water collected from immediately downstream of the effluent discharge and from a small tributary which ran down one side of the treatment plant. However, no marker-positive isolates were detected upstream of the treatment plant, despite the relatively high faecal coliform levels (mean = 3.5xl0³cfu/100ml) recorded at these sites. Further investigations on the side tributary revealed a possible entry point for sewage from the treatment system. Thus the PCR-based approach described here shows promise for the detection of human faecal contamination in rural environments and may be of use far other environmental applications such as the detection of septic tank leachate in rural catchments. [ABSTRACT FROM AUTHOR]