Multiplex PCR identification and culture-independent quantification of Bacillus licheniformis by qPCR using specific DNA markers.

Multiplex PCR identification and culture-independent quantification of Bacillus licheniformis by qPCR using specific DNA markers. Food Microbiol. 2018 Sep;74:1-10 Authors: Almeida E, Serra CR, Albuquerque P, Guerreiro I, Teles AO, Enes P, Tavares F Abstract Probiotics benefits in fish farming have been usually inferred appraising the effects observed on the host and not through the direct assessment of probiotic dynamics in the host gut microbiota. To overcome this gap, quantitative PCR (qPCR) can be a powerful approach to study the bacterial dynamics in fish gut microbiota. The presented work proposes four B. licheniformis-specific DNA markers and details a qPCR method to track putative probiotics B. licheniformis on fish gut. The four B. licheniformis-specific DNA markers - BL5B (hypothetical protein BL00303), BL8A (serA2), BL13C (rfaB) and BL18A (ligD) - were selected and validated by PCR and multiplex-PCR with 20 B. licheniformis isolates and a broad range of non-target bacteria. To assess the dynamics of B. licheniformis in the digesta of farmed fish, a qPCR was validated using markers BL8A and BL18A and calibration curves obtained for both markers with digesta samples spiked with B. licheniformis cells showed a high correlation (R2 > 0.99) over 6 log units (CFU/reaction), and a limit of detection (LOD) as low as 247 CFUs/reaction. Furthermore, the consistent qPCR repeatability and reproducibility underline the sp...
Source: Food Microbiology - Category: Food Science Authors: Tags: Food Microbiol Source Type: research