RAPD and REA for Characterization of Lactobacillus plantarum strains, and Enterococcus spp. with Special Reference to Enterococcus spp. from Meat and Humans

Sammanfattning: Restriction Endonuclease Analysis (REA) was used to characterize strains of Lactobacillus plantarum, Lb. pentosus, and Lb. reuteri. This method was shown to be a powerful tool for separation at the species and strain level. RAPD analysis of raw cell abstracts was used to characterize strains of Lb. plantarum. This method was shown to be useful for separation of most strains, and the results were obtained within one working day. The species distribution and antibiotic resistance profile of 277 Enterococcus strains from Swedish and Danish retailed pork and chicken were investigated. Enterococcus faecium was the most common species in Swedish and Danish chicken, followed by E. faecalis. E. faecalis was the dominating species in Danish and Swedish pork. The number of unidentified Enterococcus isolates, was highest in Swedish pork, and lowest in Swedish chicken. Fourteen percent of the Enterococcus isolates from Danish pork were resistant to one or more of the tested antibiotics. The values for Danish chicken, Swedish pork, and Swedish chicken were 55%, 9%, and 73%, respectively. The most common antibiotic resistance in Danish pork and Swedish chicken was tetracycline. Erythromycin resistance was the most common in Danish chicken. In the Swedish pork, only five resistant isolates were found, resistant to erythromycin, norfloxacin, penicillin V, tetracycline, and trimethoprim/sulfa, respectively. Vancomycin resistant strains, harboring the vanA gene, were found only in Danish meat, and with one exception, only in chicken. One vancomycin resistant isolate from Danish chicken was identified as E. durans, one as E. hirae, one as belonging to the E. faecium group (E. faecium, E. durans, E. hirae, and E. mundtii), and the isolate from Danish pork could not be identified to the species level. The rest were identified as E. faecium. The use of RAPD analysis, partial 16S rDNA sequence analysis including the variable regions V4 and V9, and Enterococcus specific 16S rDNA hybridization analysis in the identification and classification of enterococci from clinical isolates was compared. 16S rDNA sequencing was more reproducible than RAPD, whereas RAPD was easier to perform, and had a somewhat higher discriminatory power. RAPD was used to identify clinically relevant species of Enterococcus. It was shown to be a useful way to rapidly identify the two clinically most important species of Enterococcus, i.e. E. faecalis and E. faecium, and species specific profiles were also obtained for other Enterococcus species. REA was used to characterize Enterococcus faecium strains from chicken, pork, and humans, together with probiotic E. faecium strains. Except for two probiotic strains, all tested strains were clearly separated using REA. Enterococcus faecium strains cluster according to their source of isolation, indicating that strains from animals do not spread to humans, and that different strains are adapted to their respective source. The number of isolates included in the study was rather limited, however, so the existence of such a spread cannot be ruled out.