Profile of microbial community of naturally fermented Sumbawa mare’s milk using next-generation sequencing

Abstract

This study aimed to investigate the bacterial and fungal/yeast diversity in naturally fermented Sumbawa mare's milk through a next-generation sequencing approach, and evaluate the quality of fermented mare's milk based on the presence of pathogenic or undesirable microorganisms. Microbial density determined using plate count agar (total aerobic bacteria), de Man Rogosa Sharpe agar (Lactobacillus), M17 agar (Lactococcus) and yeast peptone dextrose agar supplemented with streptomycin 50 ppm (yeast). Nutritional content and acidity level of each fermented milk sample were also evaluated. Genomic DNA was extracted using FastDNA Spin (MPBIO). The total gDNA was further analyzed using illumina high-throughput sequencing (paired-end reads), and the sequence results were analysed using QIIME v.1.9.1 to generate diversity profiles. The difference in nutrient content of mare's milk was thought to affect the density and diversity of microbes that were able to grow. Fermented mare's milk samples from Sumbawa had the highest bacterial diversity compared to samples from Bima and Dompu. However, fermented mare's milk from Dompu had the best quality which was indicated by the absence of bacteria that have the potential to be pathogenic or food spoilage, such as members of the Enterobacteriaceae family (Enterobacter, Klebsiella and Escherichia-Shigella) and Pseudomonas. Genus of Kazachstania and Kluyveromyces, as well as family Dipodascaceae were frequently observed fungi/yeast from Sumbawa fermented mare's milk. The presence of potential pathogenic bacteria warrants special attention in improving the hygiene of manufacturing process.

Diversity analysis, identification, and bioprospecting of Lactic Acid Bacteria (LAB) isolated from Sumbawa horse milk

“…Next-generation sequencing (NGS) using Illumina is the most widely applied technique in metagenomic studies for analyzing microbial diversity and functional analysis profiles (Jatmiko et al 2019). NGS is the most effective approach technology for individual classification to particular taxa and allowed finer characterization of bacterial genomes as well as deeper taxonomic recognition of complex microbiomes (Nimnoi and Pongsilp 2020).…”

“…The hypervariable V3-V4 region in the 16S rRNA gene is a target of PCR amplification that can provide information about the bacterial community profile more accurately than the hypervariable V1-V3 region (Teng et al 2018). L.22PR Lactococcus garvieae JCM 10343 42% ± 0,007 c L.24PR Enterococcus faecalis NBRC 100480 31% ± 0,041 ab Note: Result followed by the same letter show no significant difference in the one way anova test, α = 0,05 Research on the bacterial community in Sumbawa horse milk conducted by Jatmiko et al (2019) also detected Lactobacillales and Bacillales. The abundant families that dominate bacteria in Sumbawa horse milk include Enterococcaceae and Streptococcaceae.…”

Occurrence of Yeasts in White-Brined Cheeses: Methodologies for Identification, Spoilage Potential and Good Manufacturing Practices

“…Currently, the rapidly evolving NGS Illumina technology established cost-effective and accurate DNA sequencing platforms (e.g., MiSeq and NextSeq 550), generating reads of more than 300 bp, i.e., compatible to the fragments recommended for yeast identifications by ITS1-5.8S-ITS2 rDNA (Schoch et al, 2012). Especially for scientific purposes, NGS Illumina technology is being a routinely used method to examine ecology and dynamics of microbiotas in diverse fermented milk products (Jatmiko et al, 2019;Sessou et al, 2019). For example, Ceugniez et al (2017) described the evolution of yeasts during ripening of the Tomme d'Orchies type cheeses (France), in which prevalent species, such as Y. lipolytica, G. geotrichum, Kluyveromyces spp., and Debaryomyces spp., were detected by ITS2 rRNA gene sequencing (Illumina).…”