Differentiation of Lactobacillus Species by Molecular Typing

Abstract

A total of 64 type, reference, clinical, health food, and stock isolates of microaerophilic Lactobacillus species were examined by restriction fragment length polymorphisms. Of particular interest were members of six of the eight species most commonly recovered from the vaginas of healthy premenopausal women, namely, Lactobacillus jensenii, L. casei, L. rhamnosus, L. acidophilus, L. plantarum, and L. fermentum. Six main groupings were identified on the basis of ribotyping. This technique was able to classify fresh isolates to the species level. In the case of the ribotype A grouping for L. rhamnosus, differences between strains were evident by chromosome typing (chromotyping). Many isolates did not possess plasmids. Six L. rhamnosus strains isolated from four different health food products appeared to be identical to L. rhamnosus ATCC 21052. The molecular typing system is useful for identifying and differentiating Lactobacillus isolates. Studies of strains of potential importance to the urogenital flora should include molecular characterization as a means of comparing genetic traits with those of strains whose characteristics associated with colonization and antagonism against pathogens have been defined.

FIG. 1

Schematic representation of the species-specific ribotypes obtained after digestion of total DNA from Lactobacillus isolates with BclI and hybridization with ³²P-labelled rRNA from E. coli. Lanes 1 to 64 represent strains, and groups A to G represent ribotypes described in Table 1.

FIG. 2

(A) Chromotyping. Agarose gel electrophoresis of total DNA after digestion with BclI. (B) Ribotyping. Autoradiogram of Southern blot of BclI-digested DNA after hybridization with ³²P-labelled rRNA from E. coli. Lanes 1 to 8, health food strains of Lactobacillus (C3-B, C4-B, C7-B, US2-B, US2-C, US3-B, US4, and US6-C); lane 9, L. fermentum ATCC 11739; lane 10, L. fermentum ATCC 8289; lane 11, clinical strain of Lactobacillus (vaginal isolate w); lane 12, L. acidophilus ATCC 4356^T; lane 13, clinical strain of Lactobacillus (strain 36).

FIG. 3

Autoradiogram of Southern blot of BclI-digested DNA after hybridization with ³²P-labelled rRNA from E. coli. Lanes 1 to 12 represent ribotype groups A to F. Lane 1, L. rhamnosus ATCC 7469^T; lane 2, L. casei subsp. casei ATCC 393; lane 3, L. paracasei subsp. paracasei ATCC 25302^T; lane 4, L. casei subsp. casei ATCC 4007; lane 5, L. paracasei subsp. paracasei ATCC 25303; lane 6, L. casei 8; lane 7, L. plantarum ATCC 14917^T; lane 8, L. plantarum ATCC 11974; lane 9, L. plantarum ATCC 14431; lane 10, L. fermentum B-54; lane 11, L. acidophilus ATCC 4356^T; lane 12, L. acidophilus T-13.