Using of Real Time PCR with Propidium Mono Azide for Detection of Viable but Non Culturable Enterobacteriaceae Isolated from Raw Milk Samples
Alaa Eldin H. Mustapha, Maha M. Mansour.
Abstract
A total of 260 samples of raw milk were gathered from a variety of farms, street vendors, residences, and dairy stores in the governorate of EL-Menoufia. 220 (84.6%) of the 260 samples had Enterobacteriaceae. At species level, Klebsiella and E. coli were most predominant at 40.4% and 32.6% respectively, followed by 21.2% Citrobacter freundii and 7.3% Proteus while Salmonella showed the lowest 1.9%. Using SYBER green real-time PCR, thirty uncultured samples were subjected to rpoB genotypic identification for viable Enterobacteriaceae detection, and all thirty (100%) of the samples showed the gene. By using SYBER green real-time PCR after PMA treatment, 10 (33.3%) of these 30 culture-negative samples showed the presence of VBNC enterobacterial isolates.
Key words: Propidium Monoazide PCR, real-time PCR, and viable but non-cultivable Enterobacteriaceae
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