Allelic variations and gene cluster modularity act as nonlinear bottlenecks for cholera emergence

The underlying factors that lead to specific strains within a species to emerge as human pathogens remain mostly enigmatic. The diarrheal disease cholera is caused by strains from a phylogenetically confined group within the <i>Vibrio cholerae</i> species, the pandemic cholera group (PCG), making it an ideal model system to tackle this puzzling phenomenon. Comprehensive analyses of over 1,840 <i>V. cholerae</i> genomes, including environmental isolates from this study, reveal that the species consists of eleven groups, with the PCG belonging to the largest and located within a lineage shared with environmental strains. This hierarchical classification provided us with a framework to unravel the ecoevolutionary dynamics of the genetic determinants associated with the emergence of toxigenic <i>V. cholerae</i>. Our analyses indicate that this phenomenon is largely dependent on the acquisition of unique modular gene clusters and allelic variations that confer a competitive advantage during intestinal colonization. We determined that certain PCG-associated alleles are essential for successful colonization whereas others provide a nonlinear competitive advantage, acting as a critical bottleneck that clarifies the isolated emergence of PCG. For instance, toxigenic strains encoding non-PCG alleles of a) <i>tcpF</i> or b) a sextuple allelic exchange mutant for genes <i>tcpA</i>, <i>toxT</i>, <i>VC0176</i>, <i>VC1791</i>, <i>rfbT,</i> and <i>ompU</i>, lose their ability to colonize the intestine. Interestingly, these alleles do not play a role in the colonization of newly established model environmental reservoirs. Our study uncovers the evolutionary roots of toxigenic <i>V. cholerae</i> offering a tractable approach for investigating the emergence of pathogenic clones within an environmental population.