Analysis and identification of Lactobacillus strains using species-specific fluorescent probes in an array format

Several strains belonging to Lactobacillus species were isolated from Ricotta forte, a typical Apulian cheese. PCR fragments of 450 bp werre obtained using primers designed from conserved regions of Tuf gene invovled in protein biosynthesis. PCR products were spotted onto nylon coated slides and hybridised with cyanine-3 labelled oligonucleotide probes specific for L. feremntum, L. sakei, L plantarum, L. rhamnosus and L casei Tuf Genes. All isolated and type strains were correctly identified although species-specific probes showed difficulty to differentiate L. casei and L. zeae. To improve the accuracy of the method 3 new specific probes were designed having single base divergence form the sequences and novel structure with pairing in the stem-loop region. In this work the fluorescent probes were able to identify closely relatedspecies without the need to sequence the PCR products. These resutls show this technolgoy could be exploited for the screening of amplified PCR products form natural isolates.