Design and Evaluation of a Lactobacillus Group-specific Ribosomal RNA-targeted Hybridization Probe and its Application to the Study of Intestinal Microecology in Pigs

The potential importance of anaerobic bacteria belonging to the Lactobacillus group has been well documented. Appropriate methods to rapidly evaluate species diversity and fluctuations in their population levels within the Lactobacillus group are being developed. Molecular tools such as hybridization probes based on rRNA sequences are well suited to these studies. The work reported here was undertaken to test the specificity of an hybridization probe to specifically recognize microorganisms of the Lactobacillus group and assess its usefulness as a quantitative tool to study fluctuations of the Lactobacillus population relative to the total bacterial population in gastrointestinal contents of pigs. We have designed a 25-mer oligonucleotide that targets a region common to and specific for the Lactobacillus group 16S rRNA sequences within the available database. The optimal wash temperature of the probe was experimentally determined to be 54 degrees C. The results obtained using the Lactobacillus group-specific probe (LGP) shows that Lactobacillus populations vary along the different segments of the gastrointestinal tract (GIT). In weaning piglets, the relative Lactobacillus signal intensity obtained constituted 100% of the relative RNA index in the stomach contents as determined by a bacterial domain probe (BDP), and between 90 to 100% in the duodenum. The signal of the Lactobacillus population decreased and reached its minimum in the distal part of the GIT. The same trend was observed in adult pigs, but in the stomach they constituted no more than 30% as determined by the BDP, and were present at lower levels in the other parts of the GIT. These studies document the quantitative importance of the lactobacilli in the stomach and small intestine of pigs. Further studies to investigate the role of lactobacilli in promoting the ecological balance of gut bacteria for probiotic therapy are being undertaken.