Robert William O'Hara1, Peter J Jenks2, Matthew Emery3, Mathew Upton1,4. 1. 1Faculty of Medicine and Dentistry, University of Plymouth, Plymouth, UK. 2. 2Department of Microbiology, University Hospitals Plymouth, Derriford, Plymouth, UK. 3. 3School of Biological and Marine Sciences, University or Plymouth, Plymouth, UK. 4. 4+441752584466.
Abstract
PURPOSE: Members of the ST127 uropathogenic E. coli (UPEC) clone have a high virulence potential and are also highly virulent in insect infection models. However, 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-recognized as they will be eradicated during empirical therapy. A genuine concern is the possibility that members of this highly virulent lineage will acquire resistance, leading to a more serious threat. The aim of this study was to design and validate a PCR assay specific to ST127. METHODOLOGY: Genomic sequences obtained from various UPEC isolates from the leading clones were used in comparative genomic analyses to allow identification of highly discriminatory sequences specific to E. coli ST127. The fliC (flagellin) and a homologue of the upaG (autotransporter adhesin) gene were identified as meeting our criteria and were used to develop a multiplex PCR assay. A total of 143 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. RESULTS: The multiplex PCR readily identified all 29 E. coli ST127 isolates but, equally importantly, produced no false positives with representatives of any of the other 98 STs tested. CONCLUSION: 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.
PURPOSE: Members of the ST127 uropathogenic E. coli (UPEC) clone have a high virulence potential and are also highly virulent in insect infection models. However, 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-recognized as they will be eradicated during empirical therapy. A genuine concern is the possibility that members of this highly virulent lineage will acquire resistance, leading to a more serious threat. The aim of this study was to design and validate a PCR assay specific to ST127. METHODOLOGY: Genomic sequences obtained from various UPEC isolates from the leading clones were used in comparative genomic analyses to allow identification of highly discriminatory sequences specific to E. coli ST127. The fliC (flagellin) and a homologue of the upaG (autotransporter adhesin) gene were identified as meeting our criteria and were used to develop a multiplex PCR assay. A total of 143 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. RESULTS: The multiplex PCR readily identified all 29 E. coli ST127 isolates but, equally importantly, produced no false positives with representatives of any of the other 98 STs tested. CONCLUSION: 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.
Authors: Robert William O'Hara; Benjamin Brown; Angela Hughes; Ashley McEwan; Andrew Birtles; Adam Hawker; Emma Davies; Hamzah Z Farooq; Peter Tilston; Dominic Haigh; Louise Hesketh; Andrew Dodgson; Kirsty Dodgson; Ahmad Shazaad; Malcolm Guiver; Nicholas Machin Journal: J Clin Virol Plus Date: 2022-07-16