COMPARAÇÃO DE MÉTODOS DEPENDENTES DE CULTIVO E DE MÉTODO MOLECULAR PARA DETERMINAÇÃO DA DIVERSIDADE BACTERIANA DE KEFIR

Introduction: Kefir is a fermented beverage with high probiotic potential and associated with several health benefits, such as antitumor and anticancer effects. To study it, lactic acid bacteria must be isolated and identified, for which molecular techniques are the most advanced. However, these techniques are also more expensive and require more specialized labor compared to culture-dependent techniques, which have lower discriminatory power. Aims: The objective o this study was to develop in vitro culture techniques that allow the isolation of a greater diversity of lactic acid bacteria from milk and kefir grains, enabling the study of different species with probiotic potential. Materials and methods: Ten selected samples were maintained under constant culture. Different DNA extraction methods were tested on selected lactic acid bacteria samples, and tests were performed using different methods, including PCR-RAPD under varying conditions, to differentiate the bacterial species under study. Results: A protocol for PCR-RAPD and for future DNA extractions involving kefir lactic acid bacteria was established. It was also determined that the culture media would be compared to each other to identify which allows the isolation of a greater diversity of species, rather than aiming to replace culture-independent techniques. Furthermore, the media to be tested were defined. However, it was not possible to perform cultivation in the different media, and the PCR-RAPD technique was not yet optimized and still requires refinement. Final considerations: Although it has not yet been possible to carry out in vitro culture techniques, molecular methods such as PCR-RAPD were explored so that, in the future, sequencing of the microbiome from an original kefir sample may serve as a reference for determining which culture medium is the most selective and allows the isolation of lactic acid bacteria with the greatest diversity.

Keywords: Lactic acid bacteria; Kefir; Microbial diversity; Growth medium.