Lactic acid bacteria (LAB) are often used for prevention and treatment of dysbiosis. However, the action of various strains of LAB on metabolism and digestion under these conditions are poorly understood. The purpose of this study was to investigate the influence of probiotic LAB on metabolism, digestion and microbiota in animals with dysbiosis. After administration of ampicillin and metronidazole male Wistar rats, were fed products containing Enterococcus faecium L3 (E.f.), Lactobacillus fermentum Z (L.f.) or milk (control 1). Animals in control group 2 were fed milk, after water instead of antibiotics. Dyspeptic symptoms disappeared after administration of probiotic compared with control 1. At the end of the experiment, an increase in the content of enterococci and lactobacilli in the proximal part of the small intestine was found in the animals treated with E.f. and L.f., respectively. After the introduction of probiotic enterococci, the quantity of lactobacilli and bifidobacteria in the intestines of rats increased, and the content of Klebsiella spp. and Escherichia coli decreased in comparison with the control group 1 and the group fed lactobacilli. The activity of alkaline phosphatase and aspartate transaminase was greater in blood serum of rats with dysbiosis receiving milk and lactobacilli. Intestinal alkaline phosphatase activity increased in the epithelium and chyme in the jejunum of the animals treated with L. f. and in the chyme only in the animals treated with E. f. Thus, the specific effects of different strains of probiotic LAB on the microbiota, and on metabolism and digestion of various nutrients were demonstrated.
Visceral fat accumulation is a major risk factor for the development of obesity-related diseases, including diabetes, hyperlipidemia, hypertension, and arteriosclerosis. Stimulation of lipolytic activity in adipose tissue or inhibition of fat synthesis is one way to prevent these serious diseases. Lactic acid bacteria have an anti-obesity effect, but the mechanisms are unclear. Therefore, we evaluated the effect of the administration of lactic acid bacteria (Lactobacillus gasseri NT) on lipid metabolism and fat synthesis in a mouse high-fat-diet model, focusing on visceral fat. Balb/c mice were fed a 45 kcal% fat diet for 13 weeks with and without a freeze-dried preparation of L. gasseri NT (109 CFU/g). An ex vivo glycerol assay with periovarian fat revealed that L. gasseri NT did not stimulate lipolytic activity. However, L. gasseri NT decreased the mRNA expression of sterol regulatory element-binding protein (SREBP) and its target gene fatty acid synthase (FAS) in the liver and decreased free fatty acid (FFA) in the blood. In conclusion, these findings indicated that administration of L. gasseri NT did not enhance lipid mobilization but can reduce fat synthesis, suggesting its potential for improving obesity-related diseases.
We constructed a deletion mutant of the pyrE gene in Bifidobacterium longum 105-A. A pyrE knockout cassette was cloned into pKKT427, a Bifidobacterium-Escherichia coli shuttle vector, and then introduced into B. longum 105-A by electroporation. The transformants were propagated and spread onto MRS plates containing 5-fluoroorotic acid (5-FOA) and uracil. 5-FOA-resistant mutants were obtained at a frequency of 4.7 × 10−5 integrations per cell. To perform pyrE gene complementation, the pyrE gene was amplified by PCR and used to construct a complementation plasmid (pKKT427-pyrE+). B. longum 105-A ∆pyrE harboring this plasmid could not grow on MRS plates containing 5-FOA, uracil and spectinomycin. We also developed a chemically defined medium (bifidobacterial minimal medium; BMM) containing inorganic salts, glucose, vitamins, isoleucine and tyrosine for positive selection of pyrE transformants. B. longum 105-A ∆pyrE could not grow on BMM agar, but the same strain harboring pKKT427-pyrE+ could. Thus, pyrE can be used as a counterselection marker in B. longum 105-A and potentially other Bifidobacterium species as well. We demonstrated the effectiveness of this system by constructing a knockout mutant of the xynF gene in B. longum 105-A by using the pyrE gene as a counterselection marker. This pyrE-based selection system will contribute to genetic studies of bifidobacteria.
Pyrosequencing-based 16S rRNA profiling has become a common powerful tool to obtain the community structure of gastrointestinal tract microbiota, but it is still hard to process the massive amount of sequence data into microbial composition data, especially at the species level. Here we propose a new approach in combining the quantitative insights into microbial ecology (QIIME), Mothur and ribosomal database project (RDP) programs to efficiently process 454 pyrosequence data to bacterial composition data up to the species level. It was demonstrated to precisely convert batch sequence data of 16S rRNA V6-V8 amplicons obtained from adult Singaporean fecal samples to taxonomically annotated biota data.
The intestinal microbiota compositions of 92 Japanese men were identified following consumption of identical meals for 3 days, and collected feces were analyzed through terminal restriction fragment length polymorphism. The obtained operational taxonomic units and smoking habits of subjects were analyzed by a data mining software. The constructed decision tree was able to identify explicitly the groups of smokers and nonsmokers. In particular, 4 smokers, who smoked 20 cigarettes/day, i.e., heavy smokers, were gathered in the same group of the decision tree and were clearly identified. Related operational taxonomic unit were traced to understand the species of bacteria, but all were found to be uncultured bacteria.