The recent rapid rise of multi-drug resistant Enterobacteriaceae (MDR-E) is threatening the treatment of common infectious diseases. Infections with such strains lead to increased mortality and morbidity. Using a cross-sectional study, we aimed to estimate the prevalence of gut colonization with extended spectrum beta-lactamase (ESBL) producing Enterobacteriaceae among healthy infants born in Pakistan, a setting with high incidence of MDR-E infections. Stool samples were collected from 104 healthy infants between the ages of 5 and 7 months. Enterobacteriaceae isolates were screened for resistance against several antimicrobial classes. Presence of ESBL and carbapenemase genes was determined using multiplex PCR. Sequence types were assigned to individual strains by multi-locus sequence typing. Phylogenetic analysis of Escherichia coli was done using the triplex PCR method. Forty-three percent of the infants were positive for ESBL-producing Enterobacteriaceae, the majority of which were E. coli. We identified several different ESBL E. coli sequence types most of which belonged to the phylogenetic group B2 (23%) or D (73%). The widespread colonization of infants in a developing country with ESBL-producing Enterobacteriaceae is concerning. The multiple sequence types and reported non-human sources support that multiple non-epidemic MDR lineages are circulating in Pakistan with healthy infants as a common reservoir.

Background The rapid global spread of extended spectrum beta lactamase-producing Escherichia coli (ESBL-E) strains threatens our ability to the treat many common infections and have become a major threat to public health. Some ESBL-E have a fitness advantage allowing them a competitive edge in gut colonization contributing to their global spread. We aimed to conduct in silico molecular characterization of virulence factors that may contribute to this fitness advantage. Methods For this observational study, we report data from fifteen whole-genome sequenced ESBL-E isolates found in the stool of a cohort of otherwise healthy infants. These strains were compared to MG1655 (commensal E. coli) and UTI89 (pan-sensitive uropathogenic E. coli). Phenotypic growth curves were done in minimal media with glucose as the only carbohydrate source. The genome sequences were assembled and annotated using Pathosystems Resource Integration Center (PATRIC) database and used to predict antibiotic resistance genes (ARGs) as well as virulence factors that may be driving the competitive advantage of these strains. Results All ESBL E. coli strains encoded multiple ARGs including those that target beta-lactams, aminoglycosides, fluoroquinolones, tetracyclines and macrolides. Growth curves in minimal media showed enhanced growth of some ESBL E. coli compared to control strains (Figure 1). ESBL-E strains 7 and 8 were also shown to have a higher copy number of carbohydrate metabolism genes. Proteome comparison of ESBL-E to MG1655 or UTI89 identified 93 and 321 proteins, respectively, with < 50% homology to the corresponding protein in the comparator strains (Figure 2). However, only 29 proteins across all ESBL-E were showed non-homology to both MG1655 and UTI89. These included both fimbrial and phosphotransferase system proteins. Figure 1 Figure 2A Figure 2B Conclusion ESBL-E strains may use a combination of attributes to out-compete commensal or non-resistant E. coli. These may include enhanced carbohydrate metabolism, increased adherence to the epithelial cells (via fimbrial proteins) or greater efficiency of carbohydrate uptake from the environment (via the phosphotransferase system). Further in vitro and in vivo studies are in progress to verify these in silico observations. Disclosures All Authors: No reported disclosures

Antimicrobial resistance is a global public health emergency. Infants, especially preterm infants and those in the neonatal intensive care unit, immunocompromised hosts, and those with chronic illnesses are at highest risk of adverse outcomes from invasive infections with antimicrobial-resistant strains.