Detection of viable but nonculturable Vibrio parahaemolyticus induced by prolonged cold-starvation using propidium monoazide real-time polymerase chain reaction.

08:00 EDT 1st April 2019 | BioPortfolio

Summary of "Detection of viable but nonculturable Vibrio parahaemolyticus induced by prolonged cold-starvation using propidium monoazide real-time polymerase chain reaction."

Viable but nonculturable (VBNC) Vibrio parahaemolyticus cannot be detected by the standard cultivation-based methods. In this study, commonly used viability assessment methods were evaluated for the detection of Vibrio parahaemolyticus in a VBNC state. V. parahaemolyticus cells exposed to nutrient-deficiency at cold temperature were used for epifluorescence microscopy with SYTO and propidium iodide (PI) staining and real-time polymerase chain reaction (qPCR) with propidium monoazide (PMA), and its resuscitative ability was determined by a temperature upshift in freshly prepared artificial sea water (ASW; pH 7) fluids. Viable cells with intact membranes always exceeded 5.0 log CFU ml in ASW microcosms at 4°C. After 80 days, cycle thresholds for V. parahaemolyticus ATCC 27969 were 16.15-16.69. During cold-starvation, PMA qPCR selectively excluded DNAs from heat-killed cells. However, there may be some penetration of PMA into undamaged cells that persisted in ASW for 150 days, as evidenced by their ability to resuscitate from a VBNC state after a temperature upshift (25°C); V. parahaemolyticus ATCC 33844 and V. parahaemolyticus ATCC 27969 were successfully reactivated from a VBNC state in ASW microcosms containing <5% NaCl, following enrichment in ASW medium (pH 7). This article is protected by copyright. All rights reserved.


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Name: Letters in applied microbiology
ISSN: 1472-765X


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