Comparison and evaluation of molecular methods used for identification and discrimination of lactic acid bacteria

Abstract

Lactobacillus and Bifidobacterium strains are present in a great variety of habitats, including fermented products, probiotic concoctions and the human colon. Some species are so closely related that it is difficult to distinguish them by microbiological techniques. Nevertheless, discrimination of isolates is an important issue in respect of application, and molecular methods such as restriction fragment length polymorphism (RFLP), random amplification of polymorphic DNA (RAPD) or species-specific polymerase chain reaction (PCR) might help in resolving this problem. In this study, PCR, RFLP and sequencing were applied to identify lactobacilli and bifidobacteria originating from various sources and the DSMZ strain collection. The microbiological composition of foods was analysed by molecular methods. Using species-specific PCR primers, three restriction enzymes (AluI, HhaI and RsaI) and sequencing, three Bifidobacterium and six Lactobacillus reference strains could be distinguished and four additional lactobacilli of food origin were identified. A combination of three molecular methods resulted in successful discrimination of nine reference strains and four isolates of food origin. Since these methods are not always accurate owing to their high genetic homogeneity, it is advisable to use more than one method for the identification of L. casei and closely related species.