Dataset Information

Novel Cholera Toxin Variant and ToxT Regulon in Environmental Vibrio mimicus Isolates: Potential Resources for the Evolution of Vibrio cholerae Hybrid Strains.

ABSTRACT: Atypical El Tor strains of Vibrio cholerae O1 harboring variant ctxB genes of cholera toxin (CT) have gradually become a major cause of recent cholera epidemics. Vibrio mimicus occasionally produces CT, encoded by ctxAB on CTX? genome; toxin-coregulated pilus (TCP), a major intestinal colonization factor; and also the CTX?-specific receptor. This study carried out extensive molecular characterization of CTX? and ToxT regulon in V. mimicus ctx-positive (ctx +) strains (i.e., V. mimicus strains containing ctx) isolated from the Bengal coast. Southern hybridization, PCR, and DNA sequencing of virulence-related genes revealed the presence of an El Tor type CTX prophage (CTXET) carrying a novel ctxAB, tandem copies of environmental type pre-CTX prophage (pre-CTXEnv), and RS1 elements, which were organized as an RS1-CTXET-RS1-pre-CTXEnv-pre-CTXEnv array. Additionally, novel variants of tcpA and toxT, respectively, showing phylogenetic lineage to a clade of V. cholerae non-O1 and to a clade of V. cholerae non-O139, were identified. The V. mimicus strains lacked the RTX (repeat in toxin) and TLC (toxin-linked cryptic) elements and lacked Vibrio seventh-pandemic islands of the El Tor strains but contained five heptamer (TTTTGAT) repeats in ctxAB promoter region similar to those seen with some classical strains of V. cholerae O1. Pulsed-field gel electrophoresis (PFGE) analysis showed that all the ctx + V. mimicus strains were clonally related. However, their in vitro CT production and in vivo toxigenicity characteristics were variable, which could be explainable by differential transcription of virulence genes along with the ToxR regulon. Taken together, our findings strongly suggest that environmental V. mimicus strains act as a potential reservoir of atypical virulence factors, including variant CT and ToxT regulons, and may contribute to the evolution of V. cholerae hybrid strains.IMPORTANCE Natural diversification of CTX? and ctxAB genes certainly influences disease severity and shifting patterns in major etiological agents of cholera, e.g., the overwhelming emergence of hybrid El Tor variants, replacing the prototype El Tor strains of V. cholerae This report, showing the occurrence of CTXET comprising a novel variant of ctxAB in V. mimicus, points out a previously unnoticed evolutionary event that is independent of the evolutionary event associated with the El Tor strains of V. cholerae Identification and cluster analysis of the newly discovered alleles of tcpA and toxT suggest their horizontal transfer from an uncommon clone of V. cholerae The genomic contents of ToxT regulon and of tandemly arranged multiple pre-CTX?Env and of a CTX?ET in V. mimicus probably act as salient raw materials that induce natural recombination among the hallmark virulence genes of hybrid V. cholerae strains. This report provides valuable information to enrich our knowledge on the evolution of new variant CT and ToxT regulons.

SUBMITTER: Neogi SB

PROVIDER: S-EPMC6344616 | biostudies-literature | 2019 Feb

REPOSITORIES: biostudies-literature

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Novel Cholera Toxin Variant and ToxT Regulon in Environmental Vibrio mimicus Isolates: Potential Resources for the Evolution of Vibrio cholerae Hybrid Strains.

Neogi Sucharit Basu SB Chowdhury Nityananda N Awasthi Sharda Prasad SP Asakura Masahiro M Okuno Kentaro K Mahmud Zahid Hayat ZH Islam Mohammad Sirajul MS Hinenoya Atsushi A Nair Gopinath Balakrish GB Yamasaki Shinji S

Applied and environmental microbiology 20190123 3

Atypical El Tor strains of Vibrio cholerae O1 harboring variant ctxB genes of cholera toxin (CT) have gradually become a major cause of recent cholera epidemics. Vibrio mimicus occasionally produces CT, encoded by ctxAB on CTXФ genome; toxin-coregulated pilus (TCP), a major intestinal colonization factor; and also the CTXФ-specific receptor. This study carried out extensive molecular characterization of CTXФ and ToxT regulon in V. mimicusctx-positive (<i ...[more]

PMID: 30446560

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Project description:Coordinate expression of many virulence genes in the human pathogen Vibrio cholerae is controlled by the ToxR, TcpP, and ToxT proteins. These proteins function in a regulatory cascade in which ToxR and TcpP, two inner membrane proteins, are required to activate toxT and ToxT is the direct activator of virulence gene expression. ToxT-activated genes include those whose products are required for the biogenesis of cholera toxin (CTX) and the toxin-coregulated pilus, the major subunit of which is TcpA. This work examined control of toxT transcription. We tested a model whereby activation of toxT by ToxR and TcpP is required to prime an autoregulatory loop in which ToxT-dependent transcription of the tcpA promoter reads through a proposed terminator between the tcpF and toxT genes to result in continued ToxT production. Primer extension analysis of RNA from wild-type classical strain O395 showed that there are two products encoding toxT, one of which is longer than the other by 105 bp. Deletion of the toxT promoter (toxTDeltapro) resulted in the abolishment of toxT transcription, as predicted. Deletion of the tcpA promoter (tcpADeltapro) had no effect on subsequent detection of the smaller toxT primer extension product, but the larger toxT product was not detected, indicating that this product may be the result of transcription from the tcpA promoter and not of initiation directly upstream of toxT. Neither mutant strain produced detectable TcpA, but the CTX levels of the strains were different. The toxTDeltapro strain produced little detectable CTX, while the tcpADeltapro strain produced CTX levels intermediate between those of the wild-type and toxTDeltapro strains. Dependence of toxT transcription on TcpP and TcpH was confirmed by analyzing RNAs from strains carrying deletions in the genes encoding these regulators. The tcpP defect resulted in undetectable toxT transcription, whereas the tcpH mutation led to a diminishing of toxT RNA but not complete abolishment. Taken together, these results suggest that toxT transcription is dependent on two different promoters; one is directly upstream and is activated in part by TcpP and TcpH, and the other is much further upstream and is activated by ToxT.

Project description:BackgroundVirulence determinants are crucial to the risk assessment of pathogens in an environment. This study investigated the presence of eleven key virulence-associated genes in Vibrio cholerae (n = 111) and Vibrio mimicus (n = 22) and eight virulence determinants in Vibrio alginolyticus (n = 65) and Vibrio parahaemolyticus (n = 17) isolated from six important water resources in Eastern Cape, South Africa, using PCR techniques. The multiple virulence gene indexes (MVGI) for sampling sites and isolates as well as hotspots for potential vibriosis outbreaks among sampling sites were determined statistically based on the comparison of MVGI.ResultThe PCR assay showed that all the V. cholerae isolates belong to non-O1/non-O139 serogroups. Of the isolates, Vibrio Cholera (84%), V. mimicus (73%), V. alginolyticus (91%) and V. parahaemolyticus (100%) isolates harboured at least one of the virulence-associated genes investigated. The virulence gene combinations detected in isolates varied at sampling site and across sites. Typical virulence-associated determinants of V. cholerae were detected in V. mimicus while that of V. parahaemolyticus were detected in V. alginolyticus. The isolates with the highest MVGI were recovered from three estuaries (Sunday river, Swartkopps river, buffalo river) and a freshwater resource (Lashinton river). The cumulative MVGI for V. cholerae, V. mimicus, V. alginolyticus and V. parahaemolyticus isolates were 0.34, 0.20, 0.45, and 0.40 respectively. The targeted Vibrio spp. in increasing order of the public health risk posed in our study areas based on the MVGI is V. alginolyticus > V. parahaemolyticus > V. cholerae > V. mimicus. Five (sites SR, PA5, PA6, EL4 and EL6) out of the seventeen sampling sites were detected as the hotspots for potential cholera-like infection and vibriosis outbreaks.ConclusionsOur findings suggest that humans having contact with water resources in our study areas are exposed to potential public health risks owing to the detection of virulent determinants in human pathogenic Vibrio spp. recovered from the water resources. The study affirms the relevancy of environmental Vibrio species to the epidemiology of vibriosis, cholera and cholera-like infections. Hence we suggest a monitoring program for human pathogenic Vibrio spp. in the environment most especially surface water that humans have contact with regularly.

Dataset Information

Novel Cholera Toxin Variant and ToxT Regulon in Environmental Vibrio mimicus Isolates: Potential Resources for the Evolution of Vibrio cholerae Hybrid Strains.

Publications

Novel Cholera Toxin Variant and ToxT Regulon in Environmental <i>Vibrio mimicus</i> Isolates: Potential Resources for the Evolution of <i>Vibrio cholerae</i> Hybrid Strains.

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