Rapid screening and identification strategy of lactic acid bacteria and yeasts based on Ramanomes technology and its application in fermented food

Abstract

There is an urgent need for quick, sensitive, thorough, and low-cost preliminary screening and rapid identification method for probiotic resource development. Here, we constructed a culture-free, accurate and sensitive “separation-enrichment-detection” rapid evaluation and identification method of probiotics in fermented food based on Ramanomes technology, and established a Ramanome reference of lactic acid bacteria and yeasts by AgNPs nanostructure array (AgNPs NSA) chips with uniform “hot spots” and high signal enhancement ability as SERS substrates. Systematic cluster analysis could clearly distinguish among different genera of lactic acid bacteria and yeast, and could indicate the affinity and difference between lactic acid bacteria of the same genus and yeast of the same genus. The recognition accuracy of convolutive neural networks was 100 %, and the recognition sensitivity was less than 10 CFU/mL. We constructed a probiotic isolation and enrichment method by velocity gradient centrifugation and density gradient differential centrifugation, and the average recoveries of lactobacilli and yeasts were greater than 98 % in the practical application, and the accuracy of the verified classification was 100 %. In conclusion, this study has established a preliminary screening strategy of “screening before cultivating” for lactic acid bacteria and yeasts, which breaks through the principle limitation of the current traditional paradigm of “cultivating before screening”. It can greatly improve the preliminary screening rate of probiotics in fermented foods, and provide a good technical support for the mining of probiotic resources from environmental or complex matrix samples.