Extensive Within-Host Diversity in Fecally Carried Extended-Spectrum-Beta-Lactamase-Producing Escherichia coli Isolates: Implications for Transmission Analyses

Studies of the transmission epidemiology of antimicrobial-resistantEscherichia coli, such as strains harboring extended-spectrum beta-lactamase (ESBL) genes, frequently use selective culture of rectal surveillance swabs to identify isolates for molecular epidemiological investigation. Typically, only single colonies are evaluated, which risks underestimating species diversity and transmission events. We sequenced the genomes of 16E. colicolonies from each of eight fecal samples (n =127 genomes; one failure), taken from different individuals in Cambodia, a region of high ESBL-producingE. coliprevalence. Sequence data were used to characterize both the core chromosomal diversity ofE. coliisolates and their resistance/virulence gene content as a proxy measure of accessory genome diversity. The 127E. coligenomes represented 31 distinct sequence types (STs). Seven (88%) of eight subjects carried ESBL-positive isolates, all containingblaCTX-Mvariants. Diversity was substantial, with a median of four STs/individual (range, 1 to 10) and wide genetic divergence at the nucleotide level within some STs. In 2/8 (25%) individuals, the sameblaCTX-Mvariant occurred in different clones, and/or differentblaCTX-Mvariants occurred in the same clone. Patterns of other resistance genes and common virulence factors, representing differences in the accessory genome, were also diverse within and between clones. The substantial diversity among intestinally carried ESBL-positiveE. colibacteria suggests that fecal surveillance, particularly if based on single-colony subcultures, will likely underestimate transmission events, especially in high-prevalence settings.