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Microbiota Analysis Using Two-step PCR and Next-generation 16S rRNA Gene Sequencing
Published on: October 15, 2019
Analysis of the bacterial diversity in Moroccan Jben cheese using TTGE, DGGE, and 16S rRNA sequencing
Safae Azzouz1,2, Soumaya Ahadaf1,2, Said Zantar2
1Abdelmalek Essaâdi University, Tetouan. Faculty of Science and Technology, Biotechnology and Biomolecular Engineering Research Team, B.P.416, Tangier, Morocco.
Abstract:
This research investigated the physicochemical, microbiological, and bacterial diversity of Jben cheese, a popular artisanal variety in Morocco. The bacterial diversity was explored using culture-independent methods, including temporal temperature gel electrophoresis (TTGE), denaturing gradient gel electrophoresis (DGGE), and high-throughput sequencing (HTS). Significant intra-sample differences were observed for most physicochemical parameters within each milk type, while inter-sample differences occurred between cow and goat cheeses for dry matter and ash. Jben cheese exhibited distinct characteristics, with low pH values of 3.96, 4.16, and 4.18 for cow, goat, and mixed cheeses, respectively. Goat cheeses had higher fat (49.23 g/100 g), ash (1.91 g/100 g), and dry matter (36.39 g/100 g) than cow cheeses. All cheeses displayed high microbial counts, with a notable prevalence of the lactic acid bacteria (LAB) group, averaging 8.80 ± 0.92 log CFU/g. Jben cheese also displayed high contamination levels with total coliforms, faecal coliforms, yeast, and molds. Fatty acid profiling revealed fraudulent practices in Jben cheese marketing, with cow or mixed cheeses sold as goat cheese, as proven by low capric acid concentration. HTS analysis of Jben cheese identified ten genera and twenty-four species, highlighting Lactococcus lactis as predominant. TTGE and DGGE confirmed the presence of L. lactis but failed to provide the detailed profile achieved through HTS analysis. HTS has been demonstrated to be more reliable, whereas TTGE/DGGE methods, though informative, were more time-consuming and less reliable. Despite limitations, the combined use of TTGE, DGGE, and HTS provided a comprehensive view of indigenous bacterial communities in Jben cheese, identifying L. lactis as the main species.
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