Antibiotic Resistance in Vibrio Cholerae: Mechanistic Insights from IncC Plasmid-Mediated Dissemination of a Novel Family of Genomic Islands Inserted at

Cholera remains a formidable disease, and reports of multidrug-resistant strains of the causative agent have become common during the last 3 decades. The pervasiveness of resistance determinants has largely been ascribed to mobile genetic elements, including SXT/R391 integrative conjugative elements, IncC plasmids, and genomic islands (GIs). Conjugative transfer of IncC plasmids is activated by the master activator AcaCD whose regulatory network extends to chromosomally integrated GIs. MGIHai6 is a multidrug resistance GI integrated at the 3' end of ( or ) in chromosome 1 of non-O1/non-O139 clinical isolates from the 2010 Haitian cholera outbreak. In the presence of an IncC plasmid expressing AcaCD, MGIHai6 excises from the chromosome and transfers at high frequency. Herein, the mechanism of mobilization of MGIHai6 GIs by IncC plasmids was dissected. Our results show that AcaCD drives expression of GI-borne genes, including and , involved in excision and mobilization. A 49-bp fragment upstream of was found to serve as the minimal origin of transfer () of MGIHai6. The direction of transfer initiated at was determined using IncC plasmid-driven mobilization of chromosomal markers via MGIHai6. In addition, IncC plasmid-encoded factors, including the relaxase TraI, were found to be required for GI transfer. Finally, exploration of genomes identified 47 novel related and potentially AcaCD-responsive GIs in 13 different genera. Despite sharing conserved features, these GIs integrate at , , or and carry a diverse cargo of genes involved in phage resistance. The increasing association of the etiological agent of cholera, serogroup O1 and O139, with multiple antibiotic resistance threatens to deprive health practitioners of this effective tool. Drug resistance in cholera results mainly from acquisition of mobile genetic elements. Genomic islands conferring multidrug resistance and mobilizable by IncC conjugative plasmids were reported to circulate in non-O1/non-O139 clinical strains isolated from the 2010 Haitian cholera outbreak. As these genomic islands can be transmitted to pandemic serogroups, their mechanism of transmission needed to be investigated. Our research revealed plasmid- and genomic island-encoded factors required for the resistance island excision, mobilization, and integration, as well as regulation of these functions. The discovery of related genomic islands carrying diverse phage resistance genes but lacking antibiotic resistance-conferring genes in a wide range of marine dwelling bacteria suggests that these elements are ancient and recently acquired drug resistance genes.