Identification of Bulgarian Sourdough Microbiota by Metagenomic Approach Using Three Commercially Available DNA Extraction Protocols
Ivelina Vassileva1, Vesselin Baev2, Galina Yahubyan2, Elena Apostolova-Kuzova2, Angel Angelov3 and Miglena Koprinarova1,4*
1Institute of Molecular Biology “Acad. Roumen Tsanev”, Bulgarian Academy of Sciences, Acad. G. Bonchev Str. bl. 21, 1113 Sofia, Bulgaria
2Department of Plant Physiology and Molecular Biology, Tzar Assen 24, University of Plovdiv, 4000 Plovdiv, Bulgaria
3Department of Biotechnology, University of Food Technologies, 26 Maritza Blvd., 4002 Plovdiv, Bulgaria
4Department of Catering and Nutrition, University of Food Technologies, 26 Maritza Blvd., 4002 Plovdiv, Bulgaria
Received: 24 May 2022
Accepted: 6 February 2023
sourdough; DNA extraction methods; V1-V316S rRNA; next-generation sequencing; metagenomics; microbiota
Research background. Sourdoughs are spontaneously formed, complex microbial ecosystems of various lactic acid bacteria (LAB) and yeast which, by producing specific metabolites, determine the quality of the baked products. In order to design and control sourdoughs with preferred nutritional characteristics it is crucial that the LAB diversity of the product of interest be elucidated.
Experimental approach. Using the opportunities of next-generation sequencing (NGS), of the 16S rRNA of V1-V3 hypervariable gene region, we studied the microbial ecosystem of a whole grain sourdough made of Triticum monococcum, originating from Southwestern Bulgaria. Since the DNA extraction method is considered crucial for the accuracy of the sequencing results, as it can introduce significant differences in the analyzed microbiota, we used three different commercial kits for DNA isolation and analyzed their impact on the observed bacterial diversity.
Results and conclusions. All three DNA extraction kits provided bacterial DNA which passed quality control and was successfully sequenced on Illumina MiSeq platform. The results received from the different DNA protocols showed variations in the microbial profiles. Alpha diversity indices (ACE, Chao, Shannon, Simpson) were also different among the three groups of results. Nevertheless, a strong dominance of phylum Firmicutes, class Bacilli, order Lactobacillales, represented mostly by family Lactobacillaceae, genus Lactobacillus (relative abundance of 63.11–82.28 %) and family Leuconostocaceae, genus Weissella (relative abundance of 3.67–36.31 %) were observed. Lactiplantibacillus plantarum and Levilactobacillus brevis with relative abundance of 16.15–31.24 % and 6.21−16.29 % respectively, were the two dominant species identified in all three DNA isolates.
Novelty and scientific contribution. The presented results give insight into the taxonomic composition of bacterial community of a specific Bulgarian sourdough. Having in mind that the sourdough is a difficult matrix for DNA isolation, on the one hand, and that there is no standardized DNA extraction protocol for this matrix, on the other hand, this pilot study aimed to give a small contribution to the future establishment and validation of such a protocol, which will allow accurate assessment of the specific microbiota of sourdough samples.