Allelic diversity uncovers protein domains contributing to the emergence of antimicrobial resistance

Abstract

Antimicrobial resistance (AMR) remains a major threat to global health. To date, tractable approaches that decipher how AMR emerges within a bacterial population remain limited. Here, we developed a framework that exploits genetic diversity from environmental bacterial populations to decode emergent phenotypes such as AMR. OmpU, is a porin that makes up to 60% of the outer membrane of Vibrio cholerae, the cholera pathogen. This porin is directly associated with the emergence of the bacterium and confers resistance to numerous host antimicrobials. In this study, we examined naturally occurring allelic variants of OmpU in environmental V. cholerae and established associations that connected genotypic variation with phenotypic outcome. We covered the landscape of gene variability and found that the porin forms two major phylogenetic clusters with striking genetic diversity. We generated 14 isogenic mutant strains, each encoding a unique ompU allele, and found that divergent genotypes lead to convergent antimicrobial resistance profiles. We identified and characterized functional domains in OmpU unique to variants conferring AMR-associated phenotypes. Specifically, we identified four conserved domains that are linked with resistance to bile and host-derived antimicrobial peptides. Mutant strains for these domains exhibit differential susceptibility patterns to these and other antimicrobials. Interestingly, a mutant strain in which we exchanged the four domains of the clinical allele for those of a sensitive strain exhibits a resistance profile closer to a porin deletion mutant. Finally, using phenotypic microarrays, we uncovered novel functions of OmpU and their connection with allelic variability. Our findings highlight the suitability of our approach towards dissecting the specific protein domains associated with the emergence of AMR and can be naturally extended to other bacterial pathogens and biological processes.

Modular small RNA drives pathogen emergence

“…Modularity spans many scales in biological systems from genes, to cells, to ecosystems and provides a cohesive mechanistic framework to explain biological complexity [74][75][76][77] . Recently, we determined that the porin OmpU contains modular protein domains that uniquely contribute to the emergence of antimicrobial resistance in certain alleles 38 . Here, we unveil a new layer in the complex modular nature of the ompU ORF and demonstrate that OueS also displays modular characteristics.…”

“…Modularity spans many scales in biological systems from genes, to cells, to ecosystems and provides a cohesive mechanistic framework to explain biological complexity [74][75][76][77] . Recently, we determined that the porin OmpU contains modular protein domains that uniquely contribute to the emergence of antimicrobial resistance in certain alleles 38 (NFC) of genetic information, which might be critical for the emergence of complex phenotypes. We contend this information is a) nested and fractal, as increasing the resolution within the unit ORF (nested) unveils successive layers of complexity (fractal), and b) compressed, as the overlapping of the coregulated modular porin gene with that of the modular sRNA confers a non-linear, finetuned regulation that is greater than the individual contributions of either of the two.…”

“…The presence of these virulence adaptive polymorphisms (VAPs) is associated with the emergence of toxigenic strains of V. cholerae , explaining their limited distribution to specific clades 37 . Interestingly, we determined that ompU in toxigenic V. cholerae strains encode VAPs and some of the virulence traits associated with the porin are allelic-dependent 37, 38 . Furthermore, VAP-encoding environmental alleles of ompU confer phenotypes associated with clinical outcomes such as antimicrobial resistance 37, 38 .…”

“…Interestingly, we determined that ompU in toxigenic V. cholerae strains encode VAPs and some of the virulence traits associated with the porin are allelic-dependent 37, 38 . Furthermore, VAP-encoding environmental alleles of ompU confer phenotypes associated with clinical outcomes such as antimicrobial resistance 37, 38 . These phenotypes are absent from mutant strains with ompU alleles that do not encode VAPs, indicating that these adaptations to virulence are present in their genomic background prior to host colonization 37, 38 .…”

“…Furthermore, VAP-encoding environmental alleles of ompU confer phenotypes associated with clinical outcomes such as antimicrobial resistance 37, 38 . These phenotypes are absent from mutant strains with ompU alleles that do not encode VAPs, indicating that these adaptations to virulence are present in their genomic background prior to host colonization 37, 38 . Overall, our data indicates that OmpU-associated traits are dependent on the specific allele encoded by a given strain and is directly associated with the emergence of toxigenic V. cholerae .…”