Cloning and heterologous expression of Lactobacillus reuteri uroporphyrinogen III synthase/methyltransferase gene (cobA/hemD): preliminary characterization.

PURPOSE OF WORK: To clone, express and characterize uroporphyrinogen III synthase/methyltransferase gene (cobA/hemD) from Lactobacillus reuteri. Some strains of Lb. reuteri produce cobalamin (vitamin B(12)). Cobalamin biosynthesis relies on the sequential action of more than 25 enzymes in a complex metabolic pathway. We have cloned, expressed and characterized the gene in Lb. reuteri that codes for the S-adenosy L: -methionine uroprophyrinogen III methyltransferase/synthase (CobA/HemD), a key bifunctional enzyme in the biosynthesis of cobalamin and other tetrapyrrols.

Effect of fermented soy product on the fecal microbiota of rats fed on a beef-based animal diet.

BACKGROUND: The aim of this study was to determine whether the consumption of soy product fermented with Enterococcus faecium CRL 183, would modify the fecal microbiota of rats fed a diet containing red meat. The rats were placed in groups, distinguished by their diets. For 60 days, group I was given a standard casein-based rodent feed and groups II-VI, the beef-based feed. From the 30th day, groups III-VI also ingested the following products: group III, E. faecium-fermented soy product; group IV, pure suspension of E. faecium; group V, sterilized fermented soy product; and group VI, unfermented soy product. RESULTS: Rats that ingested fermented soy product showed a slight increase in the numbers of lactobacilli (0.45 log CFU g(-1)), as did the casein-based diet group (0.47 log CFU g(-1)). The fermented soy product did not cause any reduction in the number of enterobacteria or clostridia, but promoted a slight fall in the viable count of Bacteroides spp. (2.80 +/- 0.20 to 2.34 +/- 0.07 log CFU g(-1)). CONCLUSIONS: The results indicate that the ingestion of this fermented soy product did not lead to significant changes in the fecal microbiota of the rats fed on a beef-based diet.

Effects of isoflavone-supplemented soy yogurt on lipid parameters and atherosclerosis development in hypercholesterolemic rabbits: a randomized double-blind study.

BACKGROUND: There is increasing interest in natural treatments to control dyslipidemia and reduce the risk of cardiovascular disease. Previous studies have demonstrated the beneficial effects of soy yogurt fermented with Enterococcus faecium CRL 183 and of dietary isoflavones on the lipid profile. The purpose of the present study was to investigate the effects of isoflavone-supplemented soy yogurt, fermented with E. faecium CRL183, on lipid parameters and atherosclerosis development in rabbits with induced hypercholesterolemia. METHODS: Forty-eight rabbits were randomly assigned to eight groups fed on the following diets for 60 days: C - control; IY - isoflavone-supplemented soy yogurt; H - hypercholesterolemic (1.0% cholesterol wt/wt diet); HY - hypercholesterolemic plus soy yogurt; HIY - hypercholesterolemic plus isoflavone-supplemented soy yogurt; HP - hypercholesterolemic plus placebo; HI - hypercholesterolemic plus isoflavone and HE - hypercholesterolemic plus pure culture of E. faecium CRL 183. Serum lipids and autoantibodies against oxLDL (oxLDL Ab) were analyzed on days 0, 30 and 60 of the treatment and the atherosclerotic lesions were quantified at the end of the experiment. RESULTS: Soy yogurt, soy yogurt supplemented with isoflavones and placebo promoted significant reductions in total cholesterol level (38.1%, 27.0% and 26.6%, respectively). Significant increases in serum HDL-C concentration relative to group H were detected in animals that ingested soy yogurt, with or without the isoflavone supplement (55.2%), E. faecium culture (43.3%) or placebo (35.8%). Intake of soy yogurt and soy yogurt supplemented with isoflavones prevented the rise of oxLDL Ab during the study period. The extent of atherosclerosis in the thoracic and abdominal aortas was reduced in the HIY, HY and HP groups. However, when the whole aorta was analyzed, animals treated with soy yogurt supplemented with isoflavones exhibited the greatest reduction (51.4%, P < 0.05) in atherosclerotic lesion area, compared to group H. CONCLUSION: Soy yogurt could be consumed as an alternative means of reducing the risk of cardiovascular disease by improving the lipid profile and inhibiting oxLDL Ab formation. Our findings also suggest that isoflavone supplementation may enhance the antiatherosclerotic effect of soy yogurt.

Pseudovitamin B(12) is the corrinoid produced by Lactobacillus reuteri CRL1098 under anaerobic conditions.

We have reported previously on the ability of Lactobacillus reuteri to produce a compound with vitamin B(12) activity. Here we report on the chemical characterisation of this corrinoid-like molecule. High performance liquid chromatography coupled to an ultraviolet diode array detector, mass spectrometry and nuclear magnetic resonance spectroscopy has enabled us to identify the compound as Coalpha-[alpha-(7-adenyl)]-Cobeta-cyanocobamide or pseudovitamin B(12). This molecule differs from cobalamin in the alpha-ligand, where it has adenine instead of 5,6-dimethylbenzimidazole bound in a alpha-glycosidic linkage to C-1 of ribose. L. reuteri is the first lactic acid bacterium in which the production of a cobalamin-like molecule has been identified and the first microorganism reported to produce exclusively pseudo-B(12).

Lactobacillus reuteri CRL1098 produces cobalamin.

We found that Lactobacillus reuteri CRL1098, a lactic acid bacterium isolated from sourdough, is able to produce cobalamin. The sugar-glycerol cofermentation in vitamin B(12)-free medium showed that this strain was able to reduce glycerol through a well-known cobalamin-dependent reaction with the formation of 1,3-propanediol as a final product. The cell extract of L. reuteri corrected the coenzyme B12 requirement of Lactobacillus delbrueckii subsp. lactis ATCC 7830 and allowed the growth of Salmonella enterica serovar Typhimurium (metE cbiB) and Escherichia coli (metE) in minimal medium. Preliminary genetic studies of cobalamin biosynthesis genes from L. reuteri allowed the identification of cob genes which encode the CobA, CbiJ, and CbiK enzymes involved in the cobalamin pathway. The cobamide produced by L. reuteri, isolated in its cyanide form by using reverse-phase high-pressure liquid chromatography, showed a UV-visible spectrum identical to that of standard cyanocobalamin (vitamin B12).

Microflora of Tafí Cheese: Changes During Manufacture and Maturation.

Changes in microbial flora during the manufacture and maturation of Argentine Tafí cheese was examined. The microbial flora was found to be predominantly mesophilic streptococci, leuconostocs and homofermentative lactobacilli. Streptococcus lactis predominated on the surface and in the internal portion of the cheese after pressing. Streptococcus cremoris reached its maximum population after salting. The main microbial types during maturation were Lactobacillus casei and Lactobacillus plantarum . Pathogenic microorganisms were not detected in any of the samples analyzed.