DESCRIPTION (provided by applicant: Cholera is a waterborne diarrheal disease caused by Vibrio cholerae of serogroup O1 and O139. The serogroup O1 can be divided in two biotypes: classical and El Tor. The current and seventh cholera pandemic is characterized by the predominance of the El Tor biotype. Recently, hybrid V. cholerae O1 clinical isolates (the Hybrid O1 and the El Tor variant) with attributes of both biotypes and enhanced virulence have been identified. Lateral gene transfer, transcriptional silencing of xenogenic DNA and gain of function through anti-silencing could play an important role in biotype evolution. The histone-like nucleoid structuring protein (H-NS) is a global regulator known to silence the transcription of laterally- acquired genes. We have shown that H-NS and quorum sensing differentially regulates the transcription of the vieSAB operon encoding a three-component regulatory system in V. cholerae biotypes. The VieSAB regulatory system is expressed in classical biotype V. cholerae where it regulates the expression of 10 % of the genome. Genes regulated by this system include those required to make cholera toxin and biofilm development. Thus, the VieSAB regulatory system is a major contributor to biotype-specific phenotypic differences impacting V. cholerae pathogenesis, environmental fitness and transmission. We found that H-NS binds with similar affinity to the promoter of classical and El Tor biotype vieSAB. However, H-NS was found to function as weaker repressor of vieSAB in the classical biotype. In this application we propose to characterize the regulation of vieSAB transcription and establish the basis for its differential expression in V. cholerae biotypes. In aim 1, we will determine the molecular basis for the differential transcriptional silencing of vieSAB by H-NS in classical and El Tor biotype V. cholerae. In aim 2, we will determine the connection between quorum sensing and vieSAB transcription in the El Tor biotype. Our study will likely identify new regulators that modulate V. cholerae pathogenesis by acting through the VieSAB regulatory system. Further, a better understanding of the molecular determinants of biotype-specific behavior could support the development of more effective cholera management strategies and perhaps even predict the emergence of new pathogenic variants.