Reducing bias in complex microbial community analysis in shrimp based on propidium monoazide combined with PCR-DGGE

Abstract

Microbial community plays a prominent role in food quality and safety, which was regarded as the major cause for food spoilage and foodborne disease. Pre-PCR propidium monoazide (PMA) treatment is an effective approach to exclude DNA amplification signal from dead cells, and application of this method in the analysis of bacterial community might help us better understand the diversity and distribution of viable bacteria in food. In this study, the effects of PMA treatment on bacterial community characteristics in shrimp were assessed by polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) fingerprinting technique. The results showed that the microbial diversity was dramatically influenced by PMA treatment. During storage at 4 °C, Shannon–Wiener index and PCoA analysis indicated there were significant differences between bacterial diversity in samples treated with and without PMA. At the level of species, compared with samples without PMA treatment, Vibrio cholera, Exiguobacterium acetylicum, Aeromonas enteropelogenes, Eubacterium multiforme, Vibrio mimicus in PMA-treated samples disappeared during storage. When samples stored at 25 °C, PMA treatment has little influence on banding patterns in DGGE profiles. In conclusion, this study provides an effective tool for precisely monitoring the diversity of viable bacteria in food samples, especially at low temperature.