Diversity of Yeasts and Molds by Culture-Dependent and Culture-Independent Methods for Mycobiome Surveillance of Traditionally Prepared Dried Starters for the Production of Indian Alcoholic Beverages

Culturalable and non-culturable microorganisms naturally ferment majority of global fermented foods and beverages. Traditional food fermentation represents an extremely valuable cultural heritage in most regions, and harbors a huge genetic potential of valuable but hitherto undiscovered strains. Holistic approaches for identification and complete profiling of both culturalable and non-culturable microorganisms in global fermented foods are of interest to food microbiologists. The application of culture-independent technique has thrown new light on the diversity of a number of hitherto unknown and non-cultural microorganisms in naturally fermented foods. Functional bacterial groups (“phylotypes”) may be reflected by their mRNA expression in a particular substrate and not by mere DNA-level detection. An attempt has been made to review the microbiology of some fermented foods and alcoholic beverages of the world.

Wine is the product of complex interactions between fungi, yeasts and bacteria that commence in the vineyard and continue throughout the fermentation process until packaging. Although grape cultivar and cultivation provide the foundations of wine flavour, microorganisms, especially yeasts, impact on the subtlety and individuality of the flavour response. Consequently, it is important to identify and understand the ecological interactions that occur between the different microbial groups, species and strains. These interactions encompass yeast-yeast, yeast-filamentous fungi and yeast-bacteria responses. The surface of healthy grapes has a predominance of Aureobasidium pullulans, Metschnikowia, Hanseniaspora (Kloeckera), Cryptococcus and Rhodotorula species depending on stage of maturity. This microflora moderates the growth of spoilage and mycotoxigenic fungi on grapes, the species and strains of yeasts that contribute to alcoholic fermentation, and the bacteria that contribute to malolactic fermentation. Damaged grapes have increased populations of lactic and acetic acid bacteria that impact on yeasts during alcoholic fermentation. Alcoholic fermentation is characterised by the successional growth of various yeast species and strains, where yeast-yeast interactions determine the ecology. Through yeast-bacterial interactions, this ecology can determine progression of the malolactic fermentation, and potential growth of spoilage bacteria in the final product. The mechanisms by which one species/strain impacts on another in grape-wine ecosystems include: production of lytic enzymes, ethanol, sulphur dioxide and killer toxin/bacteriocin like peptides; nutrient depletion including removal of oxygen, and production of carbon dioxide; and release of cell autolytic components. Cell-cell communication through quorum sensing molecules needs investigation.

This work aims to evaluate the antimicrobial potential of ethanolic and water extracts of roselle (Hibiscus sabdariffa), rosemary (Rosmarinus officinalis), clove (Syzygium aromaticum), and thyme (Thymus vulgaris) on some food pathogens and spoilage microorganisms. Agar well diffusion method has been used to determine the antimicrobial activities and minimum inhibitory concentrations (MIC) of different plant extracts against Gram-positive bacteria (Bacillus cereus, Staphylococcus aureus), Gram-negative bacteria (Escherichia coli, Salmonella enteritidis, Vibrio parahaemolyticus, and Pseudomonas aeruginosa), and one fungus (Candida albicans). The extracts exhibited both antibacterial and antifungal activities against tested microorganisms. Ethanolic roselle extract showed significant antibacterial activity (P < 0.05) against all tested bacterial strains, while no inhibitory effect on Candida albicans (CA) was observed. Only the ethanolic extracts of clove and thyme showed antifungal effects against CA with inhibition zones ranging from 25.2 ± 1.4 to 15.8 ± 1.2 mm, respectively. Bacillus cereus (BC) appears to be the most sensitive strain to the aqueous extract of clove with a MIC of 0.315%. To enhance our understanding of antimicrobial activity mechanism of plant extracts, the changes in internal pH (pHint), and membrane potential were measured in Staphylococcus aureus (SA) and Escherichia coli (EC) cells after exposure to the plant extracts. The results indicated that the plant extracts significantly affected the cell membrane of Gram-positive and Gram-negative bacteria, as demonstrated by the decline in pHint as well as cell membrane hyperpolarization. In conclusion, plant extracts are of great value as natural antimicrobials and can use safely as food preservatives.