{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"institution":[{"name":"bioRxiv"}],"indexed":{"date-parts":[[2026,10,3]],"date-time":"2026-10-03T11:52:49Z","timestamp":1791028369907,"version":"4.1.0"},"posted":{"date-parts":[[2026,9,29]]},"group-title":"Microbiology","reference-count":89,"publisher":"openRxiv","license":[{"start":{"date-parts":[[2026,9,29]],"date-time":"2026-09-29T00:00:00Z","timestamp":1790640000000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by-nc-nd\/4.0\/"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"accepted":{"date-parts":[[2026,9,28]]},"abstract":"<jats:title>Abstract<\/jats:title>\n                <jats:p>\n                  <jats:italic>Vibrio cholerae<\/jats:italic>\n                  , the facultative pathogen underlying cholera, naturally inhabits warm aquatic estuaries. Environmental persistence is enhanced by the ability of\n                  <jats:italic>V. cholerae<\/jats:italic>\n                  to colonize host reservoirs and form multicellular biofilms, causing seasonally endemic outbreaks in many tropical regions. Most toxigenic strains utilize the type IVa mannose-sensitive hemagglutinin (MSHA) pilus for host reservoir colonization and biofilm formation. Temperature and salinity can alter\n                  <jats:italic>V. cholerae<\/jats:italic>\n                  biofilm formation, yet their impact on MSHA production specifically remains largely unknown. Here, we utilized transcriptional reporters of predicted\n                  <jats:italic>msh<\/jats:italic>\n                  promoters (\n                  <jats:italic>msh<\/jats:italic>\n                  -P1\/\n                  <jats:italic>msh<\/jats:italic>\n                  -P2\/\n                  <jats:italic>msh<\/jats:italic>\n                  -P3) and functional assays, to determine temperature and salinity impacts on\n                  <jats:italic>msh<\/jats:italic>\n                  expression and pilus biogenesis. Under standard laboratory conditions (30\u00b0C, 1% NaCl) only\n                  <jats:italic>msh<\/jats:italic>\n                  -P1\/P2 are active and inversely-coordinated with one another. Both\n                  <jats:italic>msh<\/jats:italic>\n                  -P1\/P2 activity were elevated by high temperature (37\u00b0C) and low salinity (0.25%\/0.5% NaCl), and reduced by low temperature (20\u00b0C\/25\u00b0C) and high salinity (2%\/3% NaCl). Temperature-mediated alterations in promoter activity were not immediately reflected in changes to cell-surface MSHA levels, whereas high salinity led to decreased MSHA production. Combining high temperature (37\u00b0C) and high salinity (2%\/3% NaCl), attenuated the salinity-mediated reduction of\n                  <jats:italic>msh<\/jats:italic>\n                  -P1\/P2 activity. Biofilm biomass levels were only substantially heightened at 25\u00b0C and 20\u00b0C, likely a result of no temperature-dependent changes in cell-surface MSHA, and additional temperature-controlled biofilm regulation previously described. We also found\n                  <jats:italic>msh<\/jats:italic>\n                  -P1\/P2 promoter activity and MSHA production varies widely across toxigenic O1 and O139 serogroups despite complete sequence homology. These results shed new light on how key signals regulate MSHA pilus production to support\n                  <jats:italic>V. cholerae<\/jats:italic>\n                  persistence in aquatic environments.\n                <\/jats:p>\n                <jats:sec>\n                  <jats:title>Importance<\/jats:title>\n                  <jats:p>\n                    Toxigenic\n                    <jats:italic>Vibrio cholerae<\/jats:italic>\n                    bacteria, are the causative agent of the life-threatening pandemic diarrheal disease cholera. Along with sporadic outbreaks often following catastrophic natural disasters or large-scale breakdowns of civil sanitation infrastructure, cholera is endemic to \u223c50 countries across primarily Asia and Africa, driven by\n                    <jats:italic>V. cholerae<\/jats:italic>\n                    \u2019s survival in brackish estuary waters. This survival is enhanced by the ability of\n                    <jats:italic>V. cholerae<\/jats:italic>\n                    to interact with estuarial surfaces, via the type IVa mannose-sensitive hemagglutinin (MSHA) pilus. This study identifies the impacts of key estuarial signals, temperature and salinity, on\n                    <jats:italic>msh<\/jats:italic>\n                    gene promoter activity and pilus production. Our results indicate that promoter activity and pilus production is reduced at lower temperatures and higher salinities, which perhaps helps to explain\n                    <jats:italic>V. cholerae<\/jats:italic>\n                    \u2019s preferential habitation of warm brackish aquatic environments.\n                  <\/jats:p>\n                <\/jats:sec>","DOI":"10.64898\/2026.09.27.754871","type":"posted-content","created":{"date-parts":[[2026,9,29]],"date-time":"2026-09-29T17:15:19Z","timestamp":1790702119000},"source":"Crossref","is-referenced-by-count":0,"title":["The Estuary Effect: Variations in Temperature and Salinity Alter\n                  <i>msh<\/i>\n                  Promoter Activity in\n                  <i>Vibrio cholerae<\/i>"],"prefix":"10.64898","author":[{"given":"Debajjyoti","family":"Basu","sequence":"first","affiliation":[{"name":"School of Biological Sciences, Illinois State University, Normal, IL, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Gursewak","family":"Bains","sequence":"additional","affiliation":[{"name":"School of Biological Sciences, Illinois State University, Normal, IL, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ben","family":"Ross","sequence":"additional","affiliation":[{"name":"School of Biological Sciences, Illinois State University, Normal, IL, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Andrew","family":"Michael","sequence":"additional","affiliation":[{"name":"School of Biological Sciences, Illinois State University, Normal, IL, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Joseph","family":"Alexander","sequence":"additional","affiliation":[{"name":"School of Biological Sciences, Illinois State University, Normal, IL, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Anindita","family":"Saha","sequence":"additional","affiliation":[{"name":"School of Biological Sciences, Illinois State University, Normal, IL, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Kyle A.","family":"Floyd","sequence":"additional","affiliation":[{"name":"School of Biological Sciences, Illinois State University, Normal, IL, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"54368","reference":[{"key":"2026100304001102000_2026.09.27.754871v1.1","doi-asserted-by":"publisher","DOI":"10.1128\/CMR.8.1.48"},{"key":"2026100304001102000_2026.09.27.754871v1.2","doi-asserted-by":"publisher","DOI":"10.1128\/MMBR.62.4.1301-1314.1998"},{"key":"2026100304001102000_2026.09.27.754871v1.3","first-page":"1","article-title":"Updated global burden of cholera in endemic countries","volume":"9","year":"2015","journal-title":"PLoS Neglected Tropical Diseases"},{"key":"2026100304001102000_2026.09.27.754871v1.4","doi-asserted-by":"publisher","DOI":"10.1175\/JCLI-D-20-0366.1"},{"key":"2026100304001102000_2026.09.27.754871v1.5","doi-asserted-by":"publisher","DOI":"10.1098\/rstb.2018.0002"},{"key":"2026100304001102000_2026.09.27.754871v1.6","unstructured":"S. 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