We propose a new method that allows accurate discrimination of Lactobacillus helveticus from other closely related homofermentative lactobacilli, especially Lactobacillus gallinarum. This method is based on the amplification by PCR of two peptidoglycan hydrolytic genes, Lhv_0190 and Lhv_0191. These genes are ubiquitous and show high homology at the intra-species level. The PCR method gave two specific PCR products, of 542 and 747 bp, for 25 L. helveticus strains coming from various sources. For L. gallinarum, two amplicons were obtained, the specific 542 bp amplicon and another one with a size greater than 1,500 bp. No specific PCR products were obtained for 12 other closely related species of lactobacilli, including the L. acidophilus complex, L. delbrueckii, and L. ultunensis. The developed PCR method provided rapid, precise, and easy identification of L. helveticus. Moreover, it enabled differentiation between the two closely phylogenetically related species L. helveticus and L. gallinarum.

• MeSH
• DNA bakterií analýza   genetika   MeSH
• fylogeneze MeSH
• genetické markery genetika   MeSH
• Lactobacillus enzymologie   genetika   MeSH
• N-acetylmuramoyl-L-alaninamidasa genetika   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• DNA bakterií MeSH
• genetické markery MeSH
• N-acetylmuramoyl-L-alaninamidasa MeSH

Eleven type strains and 24 Lactobacillus isolates, preliminarily classified to the species due to phenotypic features, were investigated. Standard methods of identification with species-specific PCRs and typing with PFGE (with ApaI, NotI and SmaI restriction enzymes) allowed us to distinguish 16 unique strains belonging to 5 species (L. acidophilus, L. delbrueckii ssp. bulgaricus, L. plantarum, L. rhamnosus, L. salivarius). Alternative approach with 16S-23S rDNA ARDRA identification (with merely two restrictases, BsuRI and TaqI) and PCR-based typing (RAPD with two random- and rep-PCR with (GTG)(5) primers) showed to be more discriminative, i.e. 21 unique strains were classified in the same species as above. As a result, 7 out of 24 phenotypically species-assigned isolates were reclassified. The alternative procedure of rapid identification and typing of Lactobacillus isolates appeared to be equally effective and shortened from 1 week to 2-3 d (in comparison to the standard methods).

• MeSH
• časové faktory MeSH
• DNA bakterií analýza   genetika   MeSH
• DNA primery MeSH
• druhová specificita MeSH
• elektroforéza v polyakrylamidovém gelu MeSH
• fenotyp MeSH
• Lactobacillus klasifikace   genetika   izolace a purifikace   MeSH
• polymerázová řetězová reakce metody   MeSH
• restrikční mapování metody   MeSH
• RNA ribozomální 16S genetika   MeSH
• RNA ribozomální 23S genetika   MeSH
• technika náhodné amplifikace polymorfní DNA metody   MeSH
• techniky typizace bakterií metody   MeSH
• Publikační typ
• časopisecké články MeSH
• hodnotící studie MeSH
• Názvy látek
• DNA bakterií MeSH
• DNA primery MeSH
• RNA ribozomální 16S MeSH
• RNA ribozomální 23S MeSH