DETECTION SYSTEM FOR ESCHERICHIA COLI-SPECIFIC VIRULENCE GENES - ABSENCE OF VIRULENCE DETERMINANTS IN B-STRAIN AND C-STRAIN

We describe a rational approach to simultaneously test Escherichia col i strains for the presence of known virulence genes in a reverse dot b lot procedure, Specific segments of virulence genes off. coli designed to have similar hybridization parameters were subcloned on plasmids a nd subsequently amplified by PCR as unlabeled probes in amounts suffic ient to be bound to nylon membranes, Various pathogenic isolates and l aboratory strains of E. coli were probed for the presence of virulence genes by labeling the genomic DNA of these strains with digoxigenin a nd then hybridizing them to the prepared nylon membranes, These hybrid ization results demonstrated that besides the E. coli K-12 safety stra in derivatives, E. coli B and C strains are also devoid of genes encod ing any of the investigated virulence factors, In contrast, pathogenic E. coli control strains, used to evaluate the method, showed typical hybridization patterns, The described probes and their easy applicatio n on a single filter were shown to provide a useful tool for the safet y assessment of E. coli strains to be used as hosts in biotechnologica l processes, This approach might also be used for the identification a nd characterization of clinically significant E. coli isolates from hu man and animal species.