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Serological studies on chemostat-grown cultures of Lactobacillus fermentum and Lactobacillus plantarum.

Lactobacillus fermentum NCTC 6991 and Lactobacillus plantarum NCIB 7220 were grown in a chemostat in the diffusible fraction of complex medium at pH 6.0 with glucose limitation. Organisms grown at different dilution rates (D) were injected into rabbits, and the resultant antisera were examined for reactivity with antigens previously isolated from batch-grown organisms. For L. fermentum, antisera obtained on injecting cells grown at D = 0.5 h-1 contained a significantly higher level of antibody reacting with lipoteichoic acid and a lower level of antibody reacting with wall polysaccharide than did antisera obtained with slower-growing cells (D = 0.05 and 0.033). Antibodies to the cell wall polysaccharide were alpha-D-glucosyl specific and cross-reacted with dextran and alpha-D-glucosyl ribitol teichoic acid from L. plantarum. The immunogenicity of the ribitol teichoic acid and lipoteichoic acid components of L. plantarum was not influenced by injecting organisms grown at different rates. However, chemical and serological studies indicate that growth of L. plantarum in the diffusible fraction of complex medium results in a wall teichoic acid of lower glucose substitution. This apparently influences the specificity of the resultant antibodies so that some sera react much less with glucosyl-substituted lipoteichoic acid and dextran.

Antibody Specificity↗

Functional expression in Lactobacillus plantarum of xylP encoding the isoprimeverose transporter of Lactobacillus pentosus.

The xylP gene of Lactobacillus pentosus, the first gene of the xylPQR operon, was recently found to be involved in isoprimeverose metabolism. By expression of xylP on a multicopy plasmid in Lactobacillus plantarum 80, a strain which lacks active isoprimeverose and D-xylose transport activities, it was shown that xylP encodes a transporter. Functional expression of the XylP transporter was shown by uptake of isoprimeverose in L. plantarum 80 cells, and this transport was driven by the proton motive force generated by malolactic fermentation. XylP was unable to catalyze transport of D-xylose.

Bacterial Proteins↗

Inhibitory effect of Lactobacillus gasseri TMC0356 and Lactobacillus GG on enhanced vascular permeability of nasal mucosa in experimental allergic rhinitis of rats.

The nasal vascular permeability of ovablumin (OVA)-sensitized Brown Norway rats was evaluated by analyzing a brilliant blue concentration in perfusate from the nose after exposure of the nasal mucus to OVA. Oral administration of Lactobacillus GG and L. gasseri TMC0356 significantly inhibited the increase in nasal vascular permeability (P<0.01). The serum IgE of the tested rats also decreased, although the change was not statistically significant. These results indicate that Lactobacillus GG and L. gasseri TMC0356 might alleviate nasal allergic symptoms by suppressing the increase in nasal vascular permeability caused by local inflammation associated with allergic rhnititis.

Animals↗

Lactobacillus casei NY1301 increases the adhesion of Lactobacillus gasseri NY0509 to human intestinal caco-2 cells.

In the presence of Lactobacillus casei NY1301, the adhesion of Lactobacillus gasseri NY0509 to cultured human intestinal Caco-2 cells was significantly increased (P<0.01). In contrast, L. gasseri NY0509 did not affect the adhesion of L. casei NY1301. A heat-stable cell component of L. casei NY1301 was involved in this increase of adhesion. These results suggest that a combination of these strains may have synergistic effects of adhesion to human intestinal mucosa.

Bacterial Adhesion↗

Amino acid sequence of a dodecapeptide from the substrate-binding region of the L-lactate dehydrogenase from Lactobacillus curvatus, Lactobacillus xylosus and Bacillus stearothermophilus.

The amino acid sequence of dodecapeptides from the substrate-binding region of 3 bacterial L-lactate dehydrogenases (Lactobacillus xylosus, Lactobacillus curvatus and Bacillus stearothermophilus) were determined. They show a very high homology to the sequences of the corresponding known animal enzymes. There is, however, an essential difference between the sequences of pro- and eucaryotic enzymes: the Asn residue in position 166, common to all eucaryotes, is replaced by serine in lactobacilli and by isoleucine in Bacillus stearothermophilus. The cysteine residue in position 165, formerly considered as essential, seems to be restricted to the vertebrates, while all so far investigated invertebrates and bacteria have threonine at this position.

Amino Acid Sequence↗

The effect of sodium acetate on the growth yield, the production of L- and D-lactic acid, and the activity of some enzymes of the glycolytic pathway of Lactobacillus sakei NRIC 1071(T) and Lactobacillus plantarum NRIC 1067(T).

The effect of sodium acetate was studied on the change of the growth yield, the production of L- and D-lactic acid, and the activity of lactate dehydrogenases (LDHs; L-lactate dehydrogenase [EC 1.1.1.27, L-LDH] plus D-lactate dehydrogenase [EC 1.1.1.28, D-LDH]), fructose-1, 6-bisphosphate aldolase [EC 4.1.2.13, FBP-aldolase], and phosphofructokinase [EC 2.7.1.11, PFK] of Lactobacillus sakei NRIC 1071(T) and Lactobacillus plantarum NRIC 1067(T). The growth yield of L. sakei NRIC 1071(T) was increased 1.6 times in the presence of sodium acetate compared with its absence. The activity of LDHs in L. sakei NRIC 1071(T) and L. plantarum NRIC 1067(T) was retained longer under the addition of sodium acetate in the reaction mixture. As a result, these strains produced much more lactic acid in the presence of sodium acetate compared with its absence. Furthermore, the activity of L-LDH in L. sakei NRIC 1071(T) cultivated in the presence of sodium acetate increased three times or more compared with the activity of the cells cultivated in its absence. Consequently, the type of stereoisomers of lactic acid produced by L. sakei shifted from the DL-type to the L-type because the ratio of L-lactic acid to D-lactic acid produced became larger with the addition of sodium acetate to culture media. This phenomenon was not observed in L. plantarum NRIC 1067(T). Further, the participation of lactate racemase is discussed from the viewpoint of the production of D-lactic acid by L. sakei.

Fructose-Bisphosphatase↗

Viability of probiotic (Bifidobacterium, Lactobacillus acidophilus and Lactobacillus casei) and nonprobiotic microflora in Argentinian Fresco cheese.

We evaluated the suitability of Argentinian Fresco cheese as a food carrier of probiotic cultures. We used cultures of Bifidobacterium bifidum (two strains), Bifidobacterium longum (two strains), Bifidobacterium sp. (one strain), Lactobacillus acidophilus (two strains), and Lactobacillus casei (two strains) in different combinations, as probiotic adjuncts. Probiotic, lactic starter (Lactococcus lactis and Streptococcus thermophilus), and contaminant (coliforms, yeasts, and molds) organisms were counted at 0, 30, and 60 d of refrigerated storage. Furthermore, the acid resistance of probiotic and starter bacteria was determined from hydrochloric solutions (pH 2 and 3) of Fresco cheese. The results showed that nine different combinations of bifidobacteria and L. acidophilus had a satisfactory viability (count decreases in 60 d <1 log order) in the cheese. Both combinations of bifidobacteria and L. casei cultures assayed also showed a satisfactory survival (counts decreased <1 log order for bifidobacteria but no decrease was detected for L. casei). On the other hand, the three combinations of bifidobacteria, L. acidophilus, and L. casei tested adapted well to the Fresco cheese environment. When a cheese homogenate at pH 3 was used to partially simulate the acidic conditions in the stomach, the probiotic cultures had an excellent ability to remain viable up to 3 h. At pH 2, the cell viability was more affected; B. bifidum was the most resistant organism. This study showed that the Argentinian Fresco cheese could be used as an adequate carrier of probiotic bacteria.

Bifidobacterium↗

Effect of Lactobacillus buchneri, Lactobacillus fermentum, Leuconostoc mesenteroides inoculants, or a chemical additive on the fermentation, aerobic stability, and nutritive value of crimped wheat grains.

The preservation of crimped wheat grains by three bacterial inoculants or a chemical additive was compared. Crimped wheat grain [56.8 g dry matter (DM)/kg] was conserved in 1.75-kg plastic bag, mini-silos without treatment, with 4L/tonne of Crimpstore (CS; an additive containing a mixture of ammonium formate, propionate, ethyl benzoate, and benzoate, SAS Kelvin Cave, Ltd., UK) or 1 x 10(5) cfu/g of each of three inoculant additives containing Lactobacillus fermentum (A), Leuconostoc mesenteroides (B), and Lactobacillus buchneri (C). Six replicates were conserved per treatment. Ensiling DM losses, chemical composition, fermentation characteristics, and aerobic stability were measured in the silages after 68 d of ensiling. All the silages were well fermented and remained stable for 84 h after aeration. Subsequently, the rate of deterioration was slowest in crimped grains treated with CS treatment, followed by those treated with inoculant C, while those treated with inoculant A deteriorated most rapidly. Residual water-soluble carbohydrate concentration was higher in crimped grains treated with CS than those treated with the inoculants. Ammonia nitrogen concentrations were lowest in CS-treated crimped grains, followed by inoculants C and A. DM losses were greater in CS-treated crimped grains than in crimped grains treated with inoculants A and C. In vivo digestibility was also measured in Texel-cross lambs fed a grass silage basal diet supplemented with the additive-treated crimped grains or a conventional, lamb finisher concentrate. Dry matter intake and digestibility were unaffected by treatment. In conclusion, bacterial inoculants containing L. buchneri are promising preservatives for crimped wheat grains.

Animals↗

Production of free conjugated linoleic acid by Lactobacillus acidophilus and Lactobacillus casei of human intestinal origin.

A gas chromatographic procedure was used for analysis of conjugated linoleic acid (CLA) isomers cis-9, trans-11-octadecadienoic; trans-10, cis-12 octadecadienoic; and trans-9, trans-11-octadecadienoic (c9t11, t10c12, t9t11) produced by lactobacilli. Four different cultures, two strains each of Lactobacillus acidophilus and Lactobacillus casei were tested for their ability to produce CLA from free linoleic acid in MRS broth supplemented with linoleic acid. Different concentrations of linoleic acid (0, 0.05, 0.1, 0.2 and 0.5 mg/ml) were added to MRS broth, inoculated with the lactobacilli, and incubated at 37 degrees C. Viable counts and amounts of individual isomers of CLA (c9t11, t10c12, t9t11) were measured at 0, 24, 48, and 72 h. All the cultures were able to produce free CLA in media supplemented with linoleic acid. Maximum production of CLA (80.14 to 131.63 microg/ml) was observed at 24 h of incubation in broth containing 0.02% of free linoleic acid. No significant (P > 0.05) increases in total CLA levels were observed after 24 h of incubation. The ability of the cultures to produce CLA in skim milk supplemented with 0.02% free linoleic acid also was studied. In this medium, the total amounts of free CLA after 24 h of incubation ranged from 54.31 to 116.53 microg/ml. The use of lactic acid bacteria able to form free CLA in cultured dairy products may have potential health or nutritional benefits. Free CLA in the products likely would be more readily available for absorption from the digestive tract than if it were incorporated into the cells of the starter culture.

Animals↗

The effect of Lactobacillus buchneri and Lactobacillus plantarum on the fermentation, aerobic stability, and ruminal degradability of low dry matter corn and sorghum silages.

The effect of Lactobacillus buchneri, alone or in combination with Lactobacillus plantarum, on the fermentation, aerobic stability, and ruminal degradability of low dry matter corn and sorghum silages was studied under laboratory conditions. The inoculants were applied at 1 x 10(6) cfu/g. Silages with no additives served as control. After treatment, the chopped forages were ensiled in 1.5-L anaerobic jars. Three jars per treatment were sampled on d 2, 4, 8, 15, and 90. After 90 d of storage, the silages were subjected to an aerobic stability test lasting 5 d, in which CO2 production, as well as chemical and microbiological parameters, was measured to determine the extent of aerobic deterioration. At the end of the ensiling period (d 90), the L. buchneri- and L. buchneri + L. plantarum-inoculated silages had significantly higher levels of acetic acid than the control and L. plantarum-inoculated silages. Therefore, yeast activity was impaired in the L. buchneri- and L. buchneri + L. plantarum-inoculated silages. As a result, L. buchneri, alone or in combination with L. plantarum, improved aerobic stability of the low dry matter corn and sorghum silages. The combination of L. buchneri and L. plantarum reduced ammonia N concentrations and fermentation losses in the silages compared with L. buchneri alone. However, L. buchneri, L. plantarum, and a combination of L. buchneri + L. plantarum did not effect in situ rumen dry matter, organic matters, or neutral detergent fiber degradability of the silages. The L. buchneri was very effective in protecting the low dry matter corn and sorghum silages exposed to air under laboratory conditions. The use of L. buchneri, alone or in combination with L. plantarum, as a silage inoculant can improve the aerobic stability of low dry matter corn and sorghum silages by inhibition of yeast activity.

Acetates↗

Microbial counts, fermentation products, and aerobic stability of whole crop corn and a total mixed ration ensiled with and without inoculation of Lactobacillus casei or Lactobacillus buchneri.

Whole crop corn (DM 29.2%) and a total mixed ration (TMR, DM 56.8%) containing wet brewers grains, alfalfa hay, dried beet pulp, cracked corn, soybean meal, and molasses at a ratio of 5:1:1:1:1:1 on fresh weight basis, were ensiled with and without Lactobacillus casei or Lactobacillus buchneri in laboratory silos. The effects of inoculation on microbial counts, fermentation products, and aerobic stability were determined after 10 and 60 d. Untreated corn silage was well preserved with high lactic acid content, whereas large numbers of remaining yeasts resulted in low stability on exposure to air. Inoculation with L. casei suppressed heterolactic fermentation, but no improvements were found in aerobic stability. The addition of L. buchneri markedly enhanced the aerobic stability, while not affecting the DM loss and NH3-N production. Large amounts of ethanol were found when the TMR was ensiled, and the content of ethanol overwhelmed that of lactic acid in untreated silage. This fermentation was related to high yeast populations and accounted for a large loss of DM found in the initial 10 d. The ethanol production decreased when inoculated with L. casei and L. buchneri, but the effects diminished at 60 d of ensiling. Inoculation with L. buchneri lowered the yeasts in TMR silage from the beginning of storage; however, the populations decreased to undetectable levels when stored for 60 d, regardless of inoculation. No heating was observed in TMR silage during aerobic deterioration test for 7 d. This stability was achieved even when a high population of yeasts remained and was not affected by either inoculation or ensiling period. The results indicate that inoculation with L. buchneri can inhibit yeast growth and improve aerobic stability of corn and TMR silage; however, high stability of TMR silage can be obtained even when no treatments were made and high population (>10(5) cfu/g) of yeasts were detected.

Aerobiosis↗

Probiotic effects of feeding heat-killed Lactobacillus acidophilus and Lactobacillus casei to Candida albicans-colonized immunodeficient mice.

Probiotic bacteria can protect immunodeficient mice from orogastric candidiasis but cause some pathology of their own. Severely immunodeficient patients may be at risk if fed viable probiotics, so this study evaluated the probiotic potential of nonviable probiotic bacteria to protect immunodeficient mice from Candida albicans infections. Heat-killed probiotic bacteria were fed to gnotobiotic bg/bg-nu/nu and bg/bg-nu/+ mice to ascertain if they could protect the mice from mucosal and systemic candidiasis. Both heat-killed Lactobacillus acidophilus (HKLA) and heat-killed Lactobacillus casei (HKLC), in comparison to control mice not fed the probiotic bacteria but challenged (oral) with C. albicans, suppressed the severity of orogastric candidiasis in bg/bg-nu/nu mice at 2 weeks after colonization with C. albicans, inhibited disseminated candidiasis in C. albicans-colonized bg/bg-nu/+ mice at 4 weeks after colonization, and suppressed the number of viable C. albicans in the alimentary tract. HKLA, but not HKLC, treatment inhibited disseminated candidiasis in bg/bg-nu/nu mice at 2 weeks after oral challenge and enhanced the proliferative responses of splenocytes from C. albicans-colonized bg/bg-nu/+ mice to C. albicans antigens. Neither HKLA nor HKLC were able to prolong the survival of gnotobiotic bg/bg-nu/nu mice after oral challenge with C. albicans. These results demonstrate that heat-killed lactobacilli can induce some (limited) protection (probiotic effect) against candidiasis in mice.

Animals↗

The consumption of two new probiotic strains, Lactobacillus gasseri CECT 5714 and Lactobacillus coryniformis CECT 5711, boosts the immune system of healthy humans.

Orally ingested probiotic bacteria are able to modulate the immune system. However, differences exist in the immunomodulatory effects of different probiotic strains. Moreover, different regulatory effects, which depend on the health status of the consumer, have been identified. This work describes a randomized, double-blind, placebo-controlled human clinical trial to investigate the immune effects on healthy people of a fermented product containing two new probiotic strains, Lactobacillus gasseri CECT 5714 and Lactobacillus coryniformis CECT 5711, which was compared with another fermented product, a standard yogurt. Consumption of either the new product or yogurt increased the proportion of phagocytic cells, including monocytes and neutrophils, as well as their phagocytic activity. However, combination of the product containing the strains L. gasseri CECT 5714 and L. coryniformis CECT 5711 also induced an increase in the proportion of natural killer (NK) cells and in IgA concentrations. The effects were higher after two weeks of treatment than after 4 weeks, which suggests regulation of the immune system. In addition, the new product enhanced immunity in the participants to a greater extent than did the control standard yogurt.

Adult↗

Systemic augmentation of the immune response in mice by feeding fermented milks with Lactobacillus casei and Lactobacillus acidophilus.

This study investigates the effect of feeding fermented milks with Lactobacillus casei, Lactobacillus acidophilus and a mixture of both micro-organisms on the specific and non-specific host defence mechanisms in Swiss mice. Animals fed with fermented milk for 8 days (100 micrograms/day) showed an increase in both phagocytic and lymphocytic activity. This activation of the immune system began on the 3rd day, reached a maximum on the 5th, and decreased slightly on the 8th day of feeding. In the 8-day treated mice, boosted with a single dose (100 micrograms) on the 11th day, the immune response increased further. The feeding with fermented milk produced neither hepatomegaly nor splenomegaly. These results suggest that L. casei and L. acidophilus, associated with intestinal mucosae, can influence the level of activation of the immune system. The possible clinical application of fermented milks as immunopotentiators is also discussed.

Animals↗

[Lactose digestion by milk fermented with Lactobacillus acidophilus and Lactobacillus casei of human origin].

The aim of the present study was to determine whether a fermented milk containing Lactobacillus casei and Lactobacillus acidophilus strains isolated from human subjects (CFM) and developed by CERELA (Centro de referencia para Lactobacilos) is better tolerated than regular milk (RM) by subjects with lactase deficiency (< 1 unidad) and lactose intolerance. We studied the digestion of the lactose present in the two milk preparations which were administered to 18 healthy subjects with lactase deficiency and 12 control subjects. Each subject ingested in random sequence on different days, 480 ml CFM, 480 ml RM and 240 ml CFM and 240 ml RM. Breath hydrogen (H2) test was used to measure lactose absorption and orocecal transit time (OCTT). The peak H2 ppm results after 480 ml of CFM were ingested, reached 19.5 +/- 12.1 ppm. The same reading when 480 ml of RM were ingested was 52.6 +/- 31.9 ppm (p < 0.008). The results also showed that 480 ml of CFM delayed OCTT median 111.0 +/- 6.78 min versus 54.0 +/- 5.09 min of RM (p < 0.001) and reduced the development of symptoms (p < 0.08) such as bloating (p < 0.05), borborygmi (p < 0.025) diarrhea (p < 0.05) and abdominal colics (p < 0.05). In spite of the small number of cases studied, it seems justified to conclude that CERELA fermented milk significantly influences lactose digestion and minor development of symptoms in lactase-deficient subjects and lengthens significantly the orocecal transit time compared with regular milk.

Adult↗

The genome of the virulent phage Lc-Nu of probiotic Lactobacillus rhamnosus, and comparative genomics with Lactobacillus casei phages.

The complete 36,466-bp genome sequence of the virulent phage Lc-Nu of probiotic Lactobacillus rhamnosus was determined. The linear dsDNA with a GC-content of 44.2% contained 3' single-stranded cohesive ends of 12 nucleotides. A total of 51 putative open reading frames (orfs) were predicted. Lc-Nu showed to be evolutionary closely related to the temperate Lactobacillus casei phages phi AT3 and A2. High DNA homology with phi AT3 was shared over the late transcribed genes, and the highest homology with A2 was within the genetic switch region. The truncated cI-like repressor was the only lysogeny related gene left, which strongly suggested Lc-Nu to be recently evolved from a temperate origin. Three putative methylases and endonucleases were detected from the region of early-transcribed genes. The putative origin of replication within the putative gene orf34 homologous to replisome organizers resembled to that of lambdoid phages. The present study suggested Lc-Nu to be a new candidate for the proposed Sfi21-like species.

Base Composition↗

Cloning and sequencing of plasmid pLC494 isolated from human intestinal Lactobacillus casei: construction of an Escherichia coli-Lactobacillus shuttle vector.

The complete nucleotide sequence of plasmid pLC494 isolated from Lactobacillus casei L-49 was determined. Plasmid pLC494 is an 8846-bp long circular molecule with a G+C content of 41.5%. Two putative open reading frames, ORF4 (282 amino acids) and ORF5 (169 amino acids), were identified as replication proteins A and B that revealed 100 and 99% similarity, respectively, with the replication proteins of plasmid pLA103 from Lactobacillus acidophilus TK8912. Upstream of ORF4 were the four repeat regions (three perfect 22-bp repeats and one imperfect motif), a putative ribosome binding site, a -10 region, and a -35 region. The shuttle vector pJLE4942 (5318 bp) was constructed using repA from pLC494, a multiple cloning site, ColE1 ori, the ori of gram-negative bacteria from vector pUC19, and the chloramphenicol resistance gene from pJIR418 as a selection marker. Transformation of several lactic acid bacteria with the vector pJLE4942 indicated that this vector might be useful as a genetic tool for the intestinal lactobacilli.

Base Sequence↗

Hydrolysis of pork muscle sarcoplasmic proteins by lactobacillus curvatus and lactobacillus sake.

Lactobacillus curvatus CECT 904 and Lactobacillus sake CECT 4808 were selected on the basis of their proteolytic activities against synthetic substrates. Further, the effects of whole cells, cell extracts, and a combination of both enzymatic sources on muscle sarcoplasmic proteins were determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and reverse-phase high-performance liquid chromatography analyses. Strains of both species displayed proteinase activities on five sarcoplasmic proteins. The inoculation of whole cells caused a degradation of peptides, whereas the addition of cell extracts resulted in the generation of both hydrophilic and hydrophobic peptides. This phenomenon was remarkably more pronounced when L. curvatus was involved. Whole cells also consumed a great amount of free amino acids, while the addition of intracellular enzymes contributed to their generation. L. sake accounted for a greater release of free amino acids. In general, cell viability and also proteolytic events were promoted when cell suspensions were provided with cell extracts as an extra source of enzymes.

Journal Article↗