GENOMIC AND CULTURE BASED MICROBIOLOGICAL CHARACTERIZATION OF FERMENTED MILK PRODUCT DAHI AND ITS IMPACT ON PRODUCT QUALITY, NUTRITION AND SAFETY

Abstract

Traditional fermented milk products are characterized by the presence of complex microbial communities. It has been well established that quality, safety and sensorial attributes of these products are related with type and succession of microbiota. Dahi is widely known for unique organoleptic, nutritional and therapeutic attributes. However, the quality of the product is highly variable due to undefined starter cultures and poor control over safety. Therefore, present study was aimed to understand the complex microbial diversity of Dahi and its impact on product quality. Selected bacterial strains were characterized for technological properties and genetic potential. Moreover, impact of selected Dahi strains was analyzed in milk ecosystem in comparison to commercial starter cultures. In first phase of study microbial diversity of Dahi was determined by using culture dependent and metagenomic approach and its effect on physiochemical, nutritional and organoleptic properties of the product was analyzed. Lactic acid bacteria were found to be dominating with up to 7 logs/gram including species of Lactobacillus, Streptococcus and Enterococcus, while fungal community was represented by Geotrichum candidum and other yeasts species with a relative abundance of nearly 3 logs/gram. Similarly, Gram negative count was also nearly 3 logs/gram. Culture independent metagenomic based diversity analysis had shown the presence of 146 bacterial and 56 fungal species. Dominating microbial species include Lactobacillus delbrueckii (53%), Streptococcus thermophilus (28%), Kluyveromyces marxianus (22%) and Torulaspora delbrueckii (14%) on average in representative Dahi samples. Potential pathogens including Gram negatives were also present relatively at low level; Pseudomonas syringe (1.9%), P. cinnamophila (0.8%), Klebsiella pneumoniae, (0.3%), Escherichia coli (3.6 %) and Staphylococcus aureus (0.15%). Microbial and nutritional profiling of Dahi was found to be comparable to many traditional cheeses and fermented milks. Average pH was 3.8 having strong correlation with lactic acid bacteria, however change in pH during storage was correlated with presence of higher fungal count. The nutritional profiling illustrated the nutraceutical importance of the product. The average fat and protein (crude) content was 3.29 and 4.82 respectively while ash content was up to 0.78% containing significant concentration of essential minerals including iron (Fe), magnesium (Mg) calcium (Ca), potassium (K) zinc xvi (Zn) and sodium (Na). Detailed lipid analysis revealed the higher HDL to LDL ratio. The organoleptic properties had shown the richness of flavoring compounds that was also confirmed by consumer’s acceptability. In second phase of study it was found that isolated lactic acid bacteria by culturing in previous phase; were mostly thermophilic having higher rate of acid production and variable salt tolerance. All lactic acid bacteria shown strong proteolytic and lipolytic activities and few also showed biogenic amine production at ambient temperature. Moreover, de-acidification ability of various G. candidum strains was analysed out of these G. candidum QAUGc01 showed optimum results. G. candidum QAUGc01 also showed remarkable antimicrobial potential against several pathogens. The consistent presence of Enterococcus spp. provoked the genome sequencing of E. mundtii QAUEM2808. Genomic analysis revealed that this bacterium had Mundaticin and Enterocin genes, while no putative genes for pathogenicity, virulence and antibiotic resistance were present which excludes the safety concerns about enterococci in Dahi. In last phase of the study milk fermentation performance of Streptococcus thermophilus QauSt1 and Lactobacillus bulgaricus QauLb01 co-culture was evaluated. Comparatively, physiochemical, nutritional and sensorial properties of fermented milk product were equally good to a commercial starter culture fermentation. In another experiment, use of G. candidum QAUGc01 was found to have a positive influence on the stability and quality of the fermented milk due to less variation in product pH, increase in solid content and decrease in syneresis. In conclusion, the present study highlighted high microbial diversity of Dahi. It also demonstrates a strong correlation of microbial diversity with physiochemical, nutritional and sensorial attributes of the product.