Vibrio cholerae Type VI Secretion System Auxiliary Cluster 3 is a Pandemic-associated Mobile Genetic Element

Abstract

All sequenced Vibrio cholerae isolates encode a contact-dependent type VI secretion system (T6SS) in three loci that terminate in a toxic effector and cognate immunity protein (E/I) pair, allowing for competitor killing and clonal expansion in aquatic environments and the host gut. Recent studies have demonstrated variability in the toxic effectors produced by different V. cholerae strains and the propensity for effector genes to undergo horizontal gene transfer. Here we demonstrate that a fourth cluster, auxiliary cluster 3 (Aux3), encoding the E/I pair tseH/tsiH, is located directly downstream from two putative recombinases and is flanked by repeat elements resembling att sites. Genomic analysis of 749 V. cholerae isolates, including both pandemic and environmental strains, revealed that Aux3 exists in two states: a [~]40 kb prophage-like element in nine environmental isolates and a [~]6 kb element in pandemic isolates. These findings indicate that Aux3 in pandemic V. cholerae is evolutionarily related to an environmental prophage-like element. In both states, Aux3 excises from the chromosome via site-specific recombination to form a circular product, likely priming the module for horizontal transfer. Finally, we show that Aux3 can integrate into the Aux3-naive chromosome in an integrase-dependent, site-specific manner. This highlights the potential of Aux3 to undergo horizontal transfer by a phage-like mechanism, which based on pandemic coincidence may confer currently unknown fitness advantages to the recipient V. cholerae cell. Significance StatementV. cholerae is a human pathogen that causes pandemics affecting 2.8 million people annually (1). The O1 El Tor lineage is responsible for the current pandemic. A subset of non-O1 strains cause cholera-like disease by producing the major virulence factors cholera toxin and toxin co-regulated pilus but fail to cause pandemics. The full set of V. cholerae pandemic factors is unknown. Here we describe the type VI secretion system (T6SS) Aux3 element as a largely pandemic-specific factor that is evolutionarily related to an environmental prophage-like element circulating in non-pathogenic strains. These findings shed light on V. cholerae T6SS evolution and indicate the Aux3 element as a pandemic-enriched mobile genetic element.