Study On Classification Of Lactobacillus Species Based On Tuf, Hsp60 And PheS Genes Partial Sequences

All lactobacilli were isolated from traditional home-made fermented dairy samples collected in Tibet, Qinghai, Inner Mongolia, Yunnan and Xinjiang of China and Mongolia. Lactobacilli were preliminary classified and identified into L. plantarum,L. casei and L. helveticus by biochemical tests and 16S rDNA sequences. 234 representative Lactobacilli were chose from all isolates based on different regions and species, which were named L. plantarum group,L. casei group and L. helveticus group in this study. First, the tuf, hsp60 and pheS gene of isolates were amplified and sequenced.Sequences were submitted to the National Center for Biotechnology Information for similarity searches through BLAST and were compared with type strains by multiple alignments with DNAMAN, the result shown that the interspecies homology of the hsp60 sequences among the L. casei group, the L. helveticus group and the L. plantarum group ranged From 86.0% to 98.2%, 86.3% to 96.3% and 93.3% to 95.7%, the intraspecies homology was 98.4% to 100.0%,99.2% to 100.0% and 98.8% to 100%. The similarities of the tuf sequences among L. casei group, the L. helveticus group and the L. plantarum group species ranged from 92.2% to 95.3%,88.1 to 98.8% and 97.4% to 98.3%, the intraspecies homology was 99.3% to 100.0%, 98.4% to 100% and 99.1% to 100.0%. The interspecies similarities of the pheS gene among the L. casei group, the L. helveticus group and the L. plantarum group ranged from 70.5% to 93.3%,82.2% to 90.1% and 82.2% to 95.2%,the intraspecies homology was 99.1% to 100.0%,99.1% to 100.0% and 99.3% to 100.0%.Phylogenies of the tuf,hsp60 and pheS genes were constructed using Mega 4.0 by UPGMA method and compared those trees to 16S rDNA tree of previously study shown that three genes phylogenies were similar to that of 16S rDNA, but the three genes provided better resolution of each Lactobacillus than 16S rDNA gene. A total of 234 strains were clearly identified into L. casei (63), L. plantarum (58) and L.helveticus (113) by comparison of sequences with those of the type strains. The results showed that the partial sequences of the three genes as an alternative molecular tool for the taxonomical analysis of Lactobacillus species and for decreasing the possibility of misidentification.