Quorum-sensing autoinducers resuscitate dormant Vibrio cholerae in environmental water samples

Abstract

Cholera epidemics have long been known to spread through water contaminated with human fecal material containing the toxigenic bacterium Vibrio cholerae. However, detection of V. cholerae in water is complicated by the existence of a dormant state in which the organism remains viable, but resists cultivation on routine bacteriological media. Growth in the mammalian intestine has been reported to trigger "resuscitation" of such dormant cells, and these studies have prompted the search for resuscitation factors. Although some positive reports have emerged from these investigations, the precise molecular signals that activate dormant V. cholerae have remained elusive. Quorum-sensing autoinducers are small molecules that ordinarily regulate bacterial gene expression in response to cell density or interspecies bacterial interactions. We have found that isolation of pathogenic clones of V. cholerae from surface waters in Bangladesh is dramatically improved by using enrichment media containing autoinducers either expressed from cloned synthase genes or prepared by chemical synthesis. These results may contribute to averting future disasters by providing a strategy for early detection of V. cholerae in surface waters that have been contaminated with the stools of cholera patients or asymptomatic infected human carriers.biofilm formation | CVEC | transmissibility T he natural habitats of the species Vibrio cholerae are estuarine or fresh water aquatic environments (1-3). In cholera endemic areas such as Bangladesh, viable V. cholerae can be readily detected in water during seasonal cholera epidemics; however, as disease incidence decreases, the isolation of viable V. cholerae becomes dramatically more difficult perhaps in part due to the influence of lytic bacteriophages (4, 5). However, during the interepidemic period, the organism can also occasionally be found in a viable but dormant state, which has been alternately referred to as the viable but nonculturable (VBNC) cells (1), conditionally viable environmental cells (CVEC) (6), or active but nonculturable (ABNC) (7). Recently we have documented the existence of CVEC in surface waters of Bangladesh by using fluorescent antibody based microscopy, which revealed clumps of V. cholerae O1 cells in water, which were often negative for V. cholerae O1 by conventional culture. To detect possible presence of relatively small number of culturable cells in such water we also used enrichment and selection approaches that depend on antibiotic resistance profiles displayed by the strains that have caused the preceding cholera epidemic. This approach referred to as antibiotic selection technique (AST) (8) allowed enhanced detection of V. cholerae by suppressing growth of other environmental bacteria that would otherwise mask the small number of V. cholerae colonies.In our previous studies, CVEC were found to be organized as aggregates of cells embedded in extracellular material, presumably Vibrio extracellular polysaccharide (VPS) (6). The genes responsible for VPS production are c...

Interspecific Quorum Sensing Mediates the Resuscitation of Viable but Nonculturable Vibrios

“…A previous study demonstrated that artificially supplementing VBNC cells with quorum sensing autoinducers can resuscitate dormant Vibrio cholerae cells to improve their detectability (20). Here we provide evidence that interspecific quorum sensing autoinducers have the capability of resuscitating dormant bacterial populations both in vivo and in vitro.…”

“…This method of validating resuscitation has been utilized by others (18,20,34) to demonstrate that the observed increase in culturability is indeed due to resuscitation rather than regrowth of a small culturable population. Statistical analysis.…”

“…This indicated that preexisting and naturally present VBNC populations within these oysters resuscitated in response to the added metabolically active bacteria. In a related study, environmental water samples were enriched with the quorum sensing molecules CAI-1 and AI-2 to enhance the culturability of Vibrio cholerae (20). These findings led us to test the hypothesis that the increase in the culturability of Vibrio spp.…”

Staying in Shape: the Impact of Cell Shape on Bacterial Survival in Diverse Environments

“…Thus, a major open question remains as to the conditions and mechanisms of VBNC resuscitation. Recently, two studies suggested that in vibrios, quorum-sensing autoinducer molecules promote the efficiency and kinetics of resuscitation (150,151). A further understanding of resuscitation will allow assessment of the specific properties of VBNC and resuscitated forms.…”

In vitro and in vivo antimicrobial efficacy of natural plant-derived compounds against Vibrio cholerae of O1 El Tor Inaba serotype

“…[17][18][19][20][21][22] To examine, if honokiol and magnolol treatment interfere with biofilm formation, V. cholerae cultures were treated with various concentrations of baicalein, honokiol, magnolol, quercetin, or resveratrol. Honokiol-or magnolol-treated V. cholerae cultures showed significantly reduced biofilm formation, while baicalein, quercetin, or resveratrol did not inhibit biofilm formation (Fig.…”