Genomic diversity of ten Escherichia coli strains associated with bloodstream infections.

Escherichia coli are usually regarded as a harmless human colonic flora. However, pathogenic strains of E. coli have been associated with infections that could range from infected mucosal surfaces by intestinal pathogenic E. coli to the more severe cases of disseminated infections throughout the body by the extraintestinal pathogroups. The main focus of this project was to investigate the genomic contents of pathogenic bloodstream infection (BSI)-associated E. coli strains. This is because the genome contents of the E. coli BSI-associated isolates have not been well studied, with only few reports indicating that the pathogenincity of these strains could be attributed to horizontally acquired DNAs known as genomic islands (GEIs). The genomic contents of 10 clinical BSI-associated E. coli strains, isolated at the Leicester Royal Infirmary were investigated in this study. The first approach used to investigate the genomic contents of these strains was by interrogating the downstream ends of tRNA genes for their GEI contents by the sequential PCR strategy tRIP-PCR (tRNA interrogation for pathogenicity islands) followed by the SGSP-PCR (single genome specific primer-PCR). In this approach the flanking regions of the tRNA sites were used to first screen the tRNA genes for their GEIs followed by amplifying the boundaries of the identified GEIs. In the second approach termed Microarray-Assisted mobilome Prospecting (MAmP), the physical genome size of the tested strains obtained by the pulsed-field gel electrophoresis (PFGE) is compared to the sum total of the bits of the genome detected or visualized by the array. The difference between the two measurements is used to estimate the size of the novel, non-microarray-represented mobile genome (mobilome) present in the tested strains. Remarkably, despite only studying 10 E. coli strains, associated with a single disease type the tRIP-PCR method has identified at least 3 GEIs that contain novel sequences, and 46 GEIs, resembling uropathogenic E. coli CFT073-like entities. One particular strain E105 had 13 tRNA sites occupied with GEIs. On the other hand, an average novel, non-microarray-borne mobilome of (219 kb /strain) was obtained by the MAmP which, corresponds with previous studies. The strategies used in this study had proved successful in addressing and identifying mobilome-rich strains. Therefore, using such approaches in combination with whole genome sequencing progects could prioritize the strains and the genomic regions that need to be sequenced. Such prioritization would avoid sequencing of hundreds of isolates to identify their novel gene pool and would reduce the cost of genomic sequencing. Moreover, applying such approaches for the identification of new virulence genes and/or pathogenic mechanisms could lead to significant improvements in the treatment of E. coli infections.