Viable but Non-Culturable Vibrio cholerae and Related Pathogens in the Environment: Implications for Release of Genetically Engineered Microorganisms

Abstract

In a series of microcosm and field studies, we have observed that over time V. cholerae and related human pathogens enter a viable but non-culturable state. Direct viable counts by epifluorescent microscopy are consistently higher than corresponding plate counts. Thus, the assumption that microorganisms, including pathogens, “die-off” or “decay” in the marine environment must be re-evaluated, since stressed or nutrient starved cells are unable to grow and be enumerated by standard plate count methods. Furthermore, animal passage reveals pathogenicity persisting for such “non-viable”cells. Indirect immunofluorescent microscopy offers a more sensitive detection system for environmental sampling for human pathogens and therefore, a more valid estimation of population size. One implication of these findings for the release of genetically engineered organisms is that highly specific methods of detection and monitoring are required, with direct detection by fluorescent antibody the most reliable at the present time.