Rapid detection of Extra-intestinal Pathogenic Escherichia coli Multi-locus Sequence Type 127 using a specific PCR

Members of the ST127 uropathogenic E. coli (UPEC) clone are highly virulent in insect infection models, but strains of this lineage are reported in relatively low numbers in many studies. ST127 strains are also usually widely susceptible to antibiotics and, consequently, their true prevalence may be under-recognised, as they will be eradicated during empiric therapy. A genuine concern is the possibility that members of this lineage will acquire resistance, leading to a more serious threat given their virulence. The aim of this study was to design and validate a PCR assay specific to ST127. Genomic sequences obtained from various UPEC isolates from the leading clones were used in comparative genomics to allow identification of highly discriminative sequences specific to E. coli ST127. The fliC (flagellin) and a putative upaG (Autotransporter adhesin) genes were identified as meeting our criteria and were used to develop a multiplex PCR assay. A total of 143 E. coli UPEC isolates representing 99 different MLST clones from three locations (North West and south west England and Riyadh, Saudi Arabia) were used to validate the PCR assay. The three primer pair multiplex PCR readily identified all 29 E. coli ST127 isolates, but equally importantly, produced no false positives with any of the other 98 ST’s tested. We report the design and validation of a specific multiplex PCR for the rapid and reliable identification of ST127, which can be used for enhanced surveillance for this high-risk clone.