Survival, induction and resuscitation of vibrio cholerae from the viable but non-culturable state in the Southern Caribbean sea / Supervivencia, inducción y resucitación de Vibrio cholerae del estado viable no cultivable en el sur del Mar Caribe

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.

El agente causal del cólera, Vibrio cholerae, puede entrar a un estado viable no cultivable (VNC) en respuesta a condiciones desfavorables. El objetivo de este estudio fue evaluar la supervivencia in situ de V. cholerae en un ambiente acuático al sur del Mar Caribe y su inducción y resucitación del estado VBNC. V. cholerae no-O1, no-O139 fue inoculado en cámaras de difusión ubicadas en el Refugio de Fauna Cuare, Venezuela, y monitoreado para contaje de colonias, células totales y viables. En 119 días de exposición al ambiente, el contaje de colonias fue < 10 UFC/mL y una fracción de la población bacteriana entró al estado VBNC. Adicionalmente, la viabilidad disminuyó dos órdenes de magnitud y ocurrieron cambios morfológicos de células bacilares a cocoides. Entre las variables del ambiente acuático, la salinidad presentó correlación negativa con el contaje de colonias. Los estudios de resucitación mostraron recuperación significativa de la cultivabilidad celular con adición de sobrenadantes de cultivos en crecimiento activo (p < 0.05). Estos resultados sugieren que V. cholerae puede persistir en estado VBNC en este ambiente de Caribe y revertir a una forma cultivable bajo condiciones favorables. El estado VBNC podría representar un paso crítico en la transmisión del cólera en áreas susceptibles.

Efecto de medios de cultivo preparados con agua de mar sobre indicadores sanitarios en aguas marinas de balnearios de Chichiriviche, estado Falcón, Venezuela / Effect of culture media prepared with sea water over health indicators in sea water of the Chichiriviche resort, Falcon State, Venezuela

Debido a su uso por humanos y para preservar la salud pública, los ambientes marinos recreacionales, deberían cumplir requisitos de calidad fisico-química y microbiológica. Sin embargo, actividades antropogénicas, tales como agricultura, ganadería, industriales, turísticas y domésticas, pueden descargar aguas servidas en playas marinas. La metodología vigente para evaluar la carga bacteriana de aguas recreacionales, no pareciera satisfactoria, particularmente en la detección de microorganismos patógenos debido a la presencia, en aguas marinas, de bacterias viables pero no cultivables (VBNC). Este estudio evaluó la carga bacteriana de aerobios mesófilos, coliformes totales/fecales y enterococos, en playas de Chichiriviche, Falcón, Venezuela y el efecto de concentraciones de agua marina sobre las densidades bacterianas. Las determinaciones se efectuaron en agar nutritivo, Mac Conkey y KF, preparados con agua destilada (MAD) o con agua de mar 10% v/v, (MAM). La carga bacteriana se incrementó entre 20 y 47 veces cuando los medios de cultivo se suplementaron con agua de mar. Las diferencias en las cargas bacterianas entre MAD y MAM, para aerobios mesófilos, coliformes totales y fecales fueron estadísticamente significativas (p≤0,05). El oxígeno disuelto y pH, en ambos medios presentaron valores permisibles.

Due to their use by humans and to preserve public health, recreational marine environments should fulfill physico-chemical and microbiological quality prerequisites. Nevertheless, anthropogenic activities such as agriculture, cattle raising, industrial, touristic and domestic, can discharge contaminated water at marine beaches. The present methods for evaluating the bacterial load of recreational waters does not appear satisfactory, especially concerning the detection of pathogenic microorganisms due to the presence, in marine waters, of viable but not cultivable bacteria (VNCB). This study evaluated the bacterial load of mesophilic aerobials, total and fecal coliforms, and enterococci at the Chichiriviche beaches, Falcon State, Venezuela, and the effect of marine water concentrations over the bacterial densities. The determinations were done in nutrient, Mac Conkey and KF agar prepared with distilled water (MAD) or with sea water (MSW). The bacterial load increased between 27 and 47 times when the culture media were supplement with sea water. The differences of the bacterial loads between MAD and MSW for mesophilic aerobials, total and fecal coliforms were statistically significant (p≤0.05). Dissolved oxygen and pH presented permissible values in both media.

Lessons from cholera & Vibrio cholerae.

Cholera is an acute form of diarrhoeal disease that plagued human civilization over the centuries. The sudden and explosive onset of the disease in the form of an outbreak or epidemic, coupled with high mortality and morbidity rates, had a tragic impact on the personal as well as social life of people living in the affected areas. The enormity of human sufferings led clinicians and scientists to carry out extensive research on cholera and Vibrio cholerae (the causative bacterium of the disease) leading to major discoveries that opened up novel areas of research or new disciplines in biomedical sciences. An attempt is made here to summarize some of these breakthroughs and outline their significance in broader perspectives. Finally, the possible impact of the global socio-political scenario on the spread of cholera epidemics (pandemicity of cholera) is briefly discussed.

Advance in Technique on the Development of Uncultured Microorganism / 微生物学通报

Viable but non-culturable state in bacterial cells as received uncultured microorganism has be- come the fundamental research issues in medical microbiology, epidemiology, general microbial ecology and sanitation quarantine since it was put forward in 1982. When in this state, bacterium are no longer able to grow and form colonies on conventional culture media, but maintain their patheogenicity, may become latent infection escaping detection to threaten enviroment and human security. With the development of modern molecular biology technique and metagenomics, it provide new research method and opportunity for uncul- tured microorganism in the environment. Recently, with the application of the metagenome technique, ge- netic fingerprinting technical etc, it becomes more and more popular. Simultaneously, along with this state become maturity in the laboratory. It provides a new research route for development and exploitation the uncultured microorganism.

PROGRESS IN RESEARCH OF MICROORGANISMS OF NATURAL ENVIRONMENTS IN THE VIABLE BUT NON-CULTURABLE STATE / 微生物学通报

At the beginning of the 1980s, a concept of viable but non-culturable(VBNC) was suggested. VBNC is a survival strategy adopted by microorganisms when they are exposed to environmental stress. This article try to make a summary of research of the conditions of VBNC formation, recovery of culturability and methods of VBNC cells detection. In addition, introduces the first growth factor of microorganisms-Rpf.