Binding of Lactobacillus reuteri to fibronectin immobilized on glass beads.

Human fibronectin was immobilized on glass beads. The beads were used to evaluate binding of Lactobacillus reuteri to fibronectin. Organisms bound to the glass beads were detected using fluorescence microscopy after treatment with acridine orange. This binding was confirmed and quantified with the use of [3H]-labelled organisms. Three strains of Lactobacillus reuteri, three strains of Lactobacillus acidophilus and one strain of Lactobacillus fermentum were tested for binding capacity. L. reuteri strain 1063 exhibited a strong binding to the immobilized fibronectin, and L. acidophilus 1754 showed a slight binding. The binding of L. reuteri to the fibronectin was mediated by a protein as judged by the absence of binding after treatment of the bacteria with proteolytic enzymes. Treatment of the bacteria with urea, SDS and heat (80 degrees C) also reduced binding. Treatment of the bacterial cells prior to the assay with fibronectin interfered with binding. Albumin did not show this interaction.

Utilization of Glycerol as a Hydrogen Acceptor by Lactobacillus reuteri: Purification of 1,3-Propanediol:NAD Oxidoreductase.

Lactobacillus reuteri utilizes exogenously added glycerol as a hydrogen acceptor during carbohydrate fermentations, resulting in higher growth rates and cell yields than those obtained during growth on carbohydrates alone. Glycerol is first converted to 3-hydroxypropionaldehyde by a coenzyme B(12)-dependent glycerol dehydratase and then reduced to 1,3-propanediol by an NAD -dependent oxidoreductase. The latter enzyme was purified and determined to have a molecular weight of 180,000; it is predicted to exist as a tetramer of identical 42,000-molecular-weight subunits.

Tn917 transposition in Lactobacillus plantarum using the highly temperature-sensitive plasmid pTV1Ts as a vector.

pTV1Ts, a temperature-sensitive plasmid coding for chloramphenicol (Cm) resistance and carrying the macrolide-lincosamide-steptogramin B (MLS) resistance transposon Tn917, was introduced into strains of Lactobacillus plantarum by electroporation. After two passages in broth medium selecting for MLS resistance at 40 degrees C and subsequent plating on solid medium, two strains, L. plantarum NC4Ts1 and L. plantarum NC7Ts5, lost chloramphenicol resistance but retained MLS resistance, indicative of Tn917 transposition into host DNA. Analysis of DNA from MLSrCms isolates from both strains revealed Tn917 insertions into resident plasmids. Restriction analysis of plasmid DNA from four MLSrCms isolates from NC7Ts5 indicated four different insertion sites.

Identification and cloning of a plasmid-encoded erythromycin resistance determinant from Lactobacillus reuteri.

Plasmid analysis, plasmid curing, cloning, and hybridization experiments were used to study four Lactobacillus reuteri strains showing high resistance to erythromycin. Plasmid curing with acriflavine resulted in a loss of erythromycin resistance in a frequency of 1-10%. For three of the strains this was accompanied by a loss of a 6.9-MDa plasmid, which was shown to be identical for the different strains and designated pLUL631. The erythromycin (erm) gene was located on a 5.5-MDa plasmid in the fourth strain. A restriction map of pLUL631 was constructed and the location of the erm gene on the plasmid was identified by cloning in Escherichia coli. By using a Streptococcus lactis-E. coli shuttle vector, the erm gene was also transformed to S. lactis and expressed. The erm gene from L. reuteri was shown to be related to the erm gene from pIP501 (Streptococcus agalactiae) by DNA-DNA hybridization.