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Survival, induction and resuscitation of Vibrio cholerae from the viable but non-culturable state in the Southern Caribbean Sea.

Fernández-Delgado M, García-Amado MA, Contreras M, Incani RN, Chirinos H, Rojas H, Suárez P - Rev. Inst. Med. Trop. Sao Paulo (2015 Jan-Feb)

Bottom Line: At 119 days of exposure to the environment, the colony count was < 10 CFU/mL and a portion of the bacterial population entered the VBNC state.Additionally, the viability decreased two orders of magnitude and morphological changes occurred from rod to coccoid cells.Among the aquatic environmental variables, the salinity had negative correlation with the colony counts in the dry season.

View Article: PubMed Central - PubMed

Affiliation: Departamento de Biología de Organismos, Universidad Simón Bolívar, Caracas, Venezuela.

ABSTRACT
The causative agent of cholera, Vibrio cholerae, can enter into a viable but non-culturable (VBNC) state in response to unfavorable conditions. The aim of this study was to evaluate the in situ survival of V. cholerae in an aquatic environment of the Southern Caribbean Sea, and its induction and resuscitation from the VBNC state. V. cholerae non-O1, non-O139 was inoculated into diffusion chambers placed at the Cuare Wildlife Refuge, Venezuela, and monitored for plate, total and viable cells counts. At 119 days of exposure to the environment, the colony count was < 10 CFU/mL and a portion of the bacterial population entered the VBNC state. Additionally, the viability decreased two orders of magnitude and morphological changes occurred from rod to coccoid cells. Among the aquatic environmental variables, the salinity had negative correlation with the colony counts in the dry season. Resuscitation studies showed significant recovery of cell cultivability with spent media addition (p < 0.05). These results suggest that V. cholerae can persist in the VBNC state in this Caribbean environment and revert to a cultivable form under favorable conditions. The VBNC state might represent a critical step in cholera transmission in susceptible areas.

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Related in: MedlinePlus

In situ survival of Vibrio cholerae onexposure to natural conditions of the Cuare Wildlife Refuge. Data are mean ± SEvalues of triplicate samples. Total cell counts (), viability cell counts (), and culturable counts ().
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f01: In situ survival of Vibrio cholerae onexposure to natural conditions of the Cuare Wildlife Refuge. Data are mean ± SEvalues of triplicate samples. Total cell counts (), viability cell counts (), and culturable counts ().

Mentions: V. cholerae showed declining recoverability on exposure to the aquaticenvironment and to nutrient depletion conditions. A large population of thismicroorganism progressively became non-culturable over a period of 119 days when thetiter of culturable or colony counts decreased four orders of magnitude (from 2 x105 to 1 x 101 CFU/mL), and the number of live cells withmembrane integrity was 5.2 x 105 cells/mL. Regardless of whether the cellscould be grown on agar, they could be seen under the microscope by direct total countwithin 106-107 cells/mL. A great difference between colony countsand total cell counts was observed since time zero of the study. Increases in the periodof V. cholerae exposure to the natural environment (up to 119 days)resulted in a progressive enhancement of non-culturable cells (Fig. 1). From these data, three subpopulations of cells could beinferred at the end of the survival studies: culturable (0.00013%), VBNC (6.80%), andnonviable (93.20%). Morphological changes and decreased size of bacterial cells wereobserved since the first days of incubation in the natural environment, comprising alarge population of coccoid forms. These results indicate physiological changes duringthe prolonged exposure of V. cholerae cells to the aquatic environmentwhich could promote the bacterial survival but decrease the recovery of stressed cellson BHI agar. Preliminary in situ survival studies and microcosmexperiments of V. cholerae non-O1, non-O139 in ASW at 17 °C wereperformed with similar viability and cultivability results (data not shown).


Survival, induction and resuscitation of Vibrio cholerae from the viable but non-culturable state in the Southern Caribbean Sea.

Fernández-Delgado M, García-Amado MA, Contreras M, Incani RN, Chirinos H, Rojas H, Suárez P - Rev. Inst. Med. Trop. Sao Paulo (2015 Jan-Feb)

In situ survival of Vibrio cholerae onexposure to natural conditions of the Cuare Wildlife Refuge. Data are mean ± SEvalues of triplicate samples. Total cell counts (), viability cell counts (), and culturable counts ().
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4325519&req=5

f01: In situ survival of Vibrio cholerae onexposure to natural conditions of the Cuare Wildlife Refuge. Data are mean ± SEvalues of triplicate samples. Total cell counts (), viability cell counts (), and culturable counts ().
Mentions: V. cholerae showed declining recoverability on exposure to the aquaticenvironment and to nutrient depletion conditions. A large population of thismicroorganism progressively became non-culturable over a period of 119 days when thetiter of culturable or colony counts decreased four orders of magnitude (from 2 x105 to 1 x 101 CFU/mL), and the number of live cells withmembrane integrity was 5.2 x 105 cells/mL. Regardless of whether the cellscould be grown on agar, they could be seen under the microscope by direct total countwithin 106-107 cells/mL. A great difference between colony countsand total cell counts was observed since time zero of the study. Increases in the periodof V. cholerae exposure to the natural environment (up to 119 days)resulted in a progressive enhancement of non-culturable cells (Fig. 1). From these data, three subpopulations of cells could beinferred at the end of the survival studies: culturable (0.00013%), VBNC (6.80%), andnonviable (93.20%). Morphological changes and decreased size of bacterial cells wereobserved since the first days of incubation in the natural environment, comprising alarge population of coccoid forms. These results indicate physiological changes duringthe prolonged exposure of V. cholerae cells to the aquatic environmentwhich could promote the bacterial survival but decrease the recovery of stressed cellson BHI agar. Preliminary in situ survival studies and microcosmexperiments of V. cholerae non-O1, non-O139 in ASW at 17 °C wereperformed with similar viability and cultivability results (data not shown).

Bottom Line: At 119 days of exposure to the environment, the colony count was < 10 CFU/mL and a portion of the bacterial population entered the VBNC state.Additionally, the viability decreased two orders of magnitude and morphological changes occurred from rod to coccoid cells.Among the aquatic environmental variables, the salinity had negative correlation with the colony counts in the dry season.

View Article: PubMed Central - PubMed

Affiliation: Departamento de Biología de Organismos, Universidad Simón Bolívar, Caracas, Venezuela.

ABSTRACT
The causative agent of cholera, Vibrio cholerae, can enter into a viable but non-culturable (VBNC) state in response to unfavorable conditions. The aim of this study was to evaluate the in situ survival of V. cholerae in an aquatic environment of the Southern Caribbean Sea, and its induction and resuscitation from the VBNC state. V. cholerae non-O1, non-O139 was inoculated into diffusion chambers placed at the Cuare Wildlife Refuge, Venezuela, and monitored for plate, total and viable cells counts. At 119 days of exposure to the environment, the colony count was < 10 CFU/mL and a portion of the bacterial population entered the VBNC state. Additionally, the viability decreased two orders of magnitude and morphological changes occurred from rod to coccoid cells. Among the aquatic environmental variables, the salinity had negative correlation with the colony counts in the dry season. Resuscitation studies showed significant recovery of cell cultivability with spent media addition (p < 0.05). These results suggest that V. cholerae can persist in the VBNC state in this Caribbean environment and revert to a cultivable form under favorable conditions. The VBNC state might represent a critical step in cholera transmission in susceptible areas.

Show MeSH
Related in: MedlinePlus