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Incorporation of deoxycytidine into deoxyribonucleic acid deoxycytidylate in Lactobacillus acidophilus R-26.

Lactobacillus acidophilus R-26 a strain deficient in ribonucleotide reductase, was grown with [G-14C]deoxycytidine as the only source of deoxyribose in the medium. Of the radioactivity incorporated into deoxyribonucleic acid, a fifth moved directly into deoxyribonucleic acid deoxycytidylate, without deamination. Furthermore, deoxycytidine and thymidine nucleotides had similar sugar/base ratios, suggesting a direct conversion of deoxycytidine nucleotides to thymidine nucleotides through deamination, without further dilution by glycosyl transfer. Although radioactivity was incorporated into both the sugar and base moieties of deoxyribonucleic acid pyrimidine deoxyribonucleotides, only the sugar moiety of purine deoxyribonucleotides was labeled. Purine deoxyribonucleotides probably were synthesized by glycosyl transfer from [G-14C]deoxycytidine to unlabeled purines, followed by phosphorylation of the deoxynucleotides.

Cytosine Nucleotides↗

Purification and characterization of a bacteriocin produced by Lactobacillus acidophilus IBB 801.

Lactobacillus acidophilus IBB 801 produces a small bacteriocin, designated acidophilin 801, with an estimated molecular mass of less than 6.5 kDa. It displays a narrow inhibitory spectrum (only related lactobacilli but including the Gram-negative pathogenic bacteria Escherichia coli Row and Salmonella panama 1467) with a bactericidal activity. The antimicrobial activity of cell-free culture supernatant fluid was insensitive to catalase but sensitive to proteolytic enzymes such as trypsin, proteinase K and pronase, heat-stable (30 min at 121 degrees C), and maintained in a wide pH range. The proteinaceous compound was isolated from cell-free culture supernatant fluid and purified. Crude bacteriocin was isolated as a floating pellicle after ammonium sulphate precipitation (40% saturation) and partially purified by extraction/precipitation with chloroform/methanol (2/1, v/v). Further purification to homogeneity was performed by reversed phase Fast Performance Liquid Chromatography. The amino acid composition was determined. Amino acid sequencing revealed that the N-terminal end was blocked.

Bacteriocins↗

Isolation, partial characterization and mode of action of acidocin J1229, a bacteriocin produced by Lactobacillus acidophilus JCM 1229.

Lactobacillus acidophilus JCM 1229 produces a heat-stable bacteriocin, designated as acidocin J1229, that has a narrow inhibitory spectrum. Production of acidocin J1229 in MRS broth was pH dependent, with maximum activity detected in broth culture maintained at pH 5.0. Acidocin J1229 was purified by ammonium sulphate precipitation and sequential cation exchange and reversed-phase chromatographies. The sequence of the first 24 amino acid residues of the N terminus of acidocin J1229 was determined. The molecular mass of acidocin J1229 as determined by mass spectrometry was 6301 Da. Acidocin J1229 showed a bactericidal effect but not a bacteriolytic effect on sensitive cells. Acidocin J1229 dissipated the membrane potential and the pH gradient in sensitive cells, which affected such proton motive force-dependent processes as amino acid transport. Acidocin J1229 also caused an efflux of glutamate, previously taken up via a unidirectional ATP-driven transport system. Secondary structure prediction revealed the presence of an amphiphilic alpha-helix region that could form hydrophilic pores. These results suggest that acidocin J1229 is a pore-forming peptide that creates cell membrane channels through the "barrel-stave' mechanism.

Amino Acid Sequence↗

Isolation, partial characterization, and mode of action of Acidocin J1132, a two-component bacteriocin produced by Lactobacillus acidophilus JCM 1132.

Lactobacillus acidophilus JCM 1132 produces a heat-stable, two-component bacteriocin designated acidocin J1132 that has a narrow inhibitory spectrum. Maximum production of acidocin J1132 in MRS broth was detected at pH 5.0. Acidocin J1132 was purified by ammonium sulfate precipitation and sequential cation exchange and reversed-phase chromatographies. Acidocin J1132 activity was associated with two components, termed alpha and beta. On the basis of N-terminal amino acid sequencing and the molecular masses of the alpha and beta components, it is interpreted that the compounds differ by an additional glycine residue in the beta component. Both alpha and beta had inhibitory activity, and an increase in activity by the complementary action of the two components was observed. Acidocin J1132 is bactericidal and dissipates the membrane potential and the pH gradient in sensitive cells, which affect such proton motive force-dependent processes as amino acid transport. Acidocin J1132 also caused efflux of preaccumulated amino acid taken up via a unidirectional ATP-driven transport system. Secondary structure prediction revealed the presence of an amphiphilic alpha-helix region that could form hydrophilic pores. These results suggest that acidocin J1132 is a pore-forming bacteriocin that creates cell membrane channels through the "barrel-stave" mechanism.

Amino Acid Sequence↗

Properties of alpha-glucosidase from Lactobacillus acidophilus NCTC 1723.

Lactobacillus acidophilus NCTC 1723 produced intracellular and extracellular alpha-glucosidase (alpha-D-glucoside glucohydrolase, EC 3.2.1.20). The alpha-glucosidase was partially purified by ammonium sulphate fractionation and DEAE-cellulose column chromatography, and attained a 10.3-fold purification. The Km for alpha-PNPG was 2.9 mM and the Vmax for alpha-PNPG hydrolysis was 6.45 mumole ml-1 min-1. The enzyme was stable only at pH 6.0-7.5 while incubated at 25 degrees C. At pH 6.5, a 100% activity was retained at 15 degrees-37 degrees C. However, the enzyme was easily destroyed at 50 degrees C. The pH optimum for stability of the enzyme at low temperature (2 degrees C) was between 5 and 6. It was found that addition of Mn++, Ba++ and EDTA, to the medium stimulated alpha-glucosidase activity, while the presence of Hg++, Cu++, Co++, Ni++, Zn++, L-histidine, arabitol, erythritol, sorbitol and glycerol inhibited enzyme activity. Although isomaltase activity was found in the partially purified alpha-glucosidase, it was not known whether this activity was an intrinsic capability of the enzyme. Transglucosylase and weak glucoamylase activities were also found to associate with the partially purified alpha-glucosidase. Since only the alpha-1,6 linked isomaltose was detected as the transferase product, it was thought that the alpha-glucosidase was capable of glucosyl transfer via alpha-1,6-glucosidic bonds.

Glucan 1,4-alpha-Glucosidase↗

[Therapy of Helicobacter pylori infection using Lactobacillus acidophilus].

INTRODUCTION: In vitro experiments with Lactobacillus acidophilus have revealed its inhibitory effect on Helicobacter pylori and its application in treatment of Helicobacter pylori positive gastritis was examined. MATERIAL AND METHODS: 15 patients have undergone gastroscopy with biopsy and by histopathological examination. Helicobacter pylori positive gastritis was detected. During a two-month period these patients took acidophilus milk (3 x 250 ml a day) prepared according to a special protocol and which contained 4 x 10(9)-1 x 10(10) live cells of Lactobacillus acidophilus at the moment of preparation. Lactobacillus acidophilus strain NAS, gained from lyophilized preparation Bio-Nate (Natren Inc. USA) was used as a test organism. Control gastroscopy was performed 2 months later. RESULTS: 14 patients have completed the examination. All of them were satisfied with the taste of acidophilus milk and could stand it well, whereas in 6 out of 14 Helicobacter pylori was eradicated. DISCUSSION: Helicobacter pylori eradication therapy is recommended in all cases of Helicobacter pylori positive gastritis associated with peptic ulcer, as well as in absence of ulcer when subjective difficulties occur. Antibiotic therapy is often unsuccessful and most often associated with risks of significant adverse effects, being the consequence of intestinal microflora disorders. The aim of using Lactobacillus acidophilus in the therapy is to reduce risks of adverse effects. In our study, by using acidophilus milk only, without other therapy, eradication of Helicobacter pylori was achieved in 6 out of 14 patients. All patients could stand the therapy well and were satisfied with the taste of the preparation. The number of examinees was small in regard to making conclusions, but the results are encouraging and show that apart from established in vitro effect. Lactobacillus acidophilus has a potential in vivo effect.

Adult↗

Detection of plasmid deoxyribonucleic acid in an isolate of Lactobacillus acidophilus.

Eight strains of Lactobacillus acidophilus were examined for the presence of plasmid deoxyribonucleic acid, and one, a pig intestinal isolate, showed the presence of a 13.7- and a 6.3-megadalton plasmid. This is the first reported evidence for plasmid deoxyribonucleic acid in Lactobacillus acidophilus. The functions of these plasmids are presently unknown.

Animals↗

Lactobacillus acidophilus, Bifidobacterium lactis and Lactobacillus F19 prevent antibiotic-associated ecological disturbances of Bacteroides fragilis in the intestine.

OBJECTIVE: The objective of this study was to compare the effect of clindamycin on the intestinal microflora in subjects ingesting yogurt with added probiotic microorganisms with the microflora in subjects ingesting placebo yogurt. MATERIALS AND METHODS: Twenty-four healthy subjects were included in the study. All subjects received 150 mg clindamycin four times daily for 7 days and 250 ml yogurt twice daily for 14 days. Faecal samples were collected before, during and after administration of clindamycin. RESULTS: In the aerobic intestinal microflora, the numbers of enterococci increased after treatment in both groups, whereas other Gram-positive microorganisms decreased. In both groups, the numbers of Escherichia coli also decreased, whereas there was a concomitant increase in numbers of other Gram-negative bacilli. In the anaerobic microflora in subjects receiving yogurt with added microorganisms, the numbers of lactobacilli and bacteroides remained at the same levels throughout the study, whereas the numbers decreased in the placebo group. Other anaerobic bacteria decreased in both groups. The minimum inhibitory concentration of clindamycin against strains of bacteroides increased in both groups during the study. CONCLUSIONS: The probiotic microorganisms evaluated in this study prevented ecological disturbances in the numbers of intestinal Bacteroides fragilis group species during clindamycin administration.

Adult↗

Enhancement of immune response in mice fed with Streptococcus thermophilus and Lactobacillus acidophilus.

Swiss mice, fed for 8 consecutive d with 50 micrograms/d of viable cultures of Lactobacillus acidophilus and Streptococcus thermophilus, showed significant variation in their immune system. In order to study this phenomenon assays for macrophage and lymphocyte function were carried out. Both lactic acid bacteria enhanced significantly the enzymatic and phagocytic activity of peritoneal macrophages as checked against the controls and also accelerated the phagocytic function of the reticuloendothelial system as revealed by the carbon clearance test. On the 2nd d (100 micrograms), L. acidophilus reached a peak of K = .271, which remained high. Streptococcus thermophilus was effective only on the 2nd d and then decreased. The lymphocytic activity studied by immunoglobulin secreting cells was assayed by Jerne's method of plaque-forming cells (PFC). This activity also was increased by the two microorganisms. Streptococcus thermophilus proved more effective than L. acidophilus. Lactobacillus acidophilus and S. thermophilus activated macrophages and lymphocytes and produced the same increase in the immune response of mice whether administered orally or intraperitoneally.

Administration, Oral↗

Betaine Transport Imparts Osmotolerance on a Strain of Lactobacillus acidophilus.

Unlike most Lactobacillus acidophilus strains, a specific strain, L. acidophilus IFO 3532, was found to grow in rich medium containing 1 M sodium acetate, KCl, or NaCl. This strain could also grow with up to 1.8 M NaCl or 3 M nonelectrolytes (fructose, xylose, or sorbitol) added. Thus, this strain was tolerant to osmotic pressures up to 2.8 osM. A search for an intracellular solute which conferred osmoprotection led to the identification of glycine betaine (betaine). Betaine was accumulated to high concentrations in cells growing in MRS medium supplemented with 1 M KCl or NaCl. Uptake of [C]betaine by L. acidophilus 3532 cells suspended in buffer was stimulated by increasing the medium osmotic pressure with 1 M KCl or NaCl. The accumulated betaine was not metabolized further; transport was relatively specific for betaine and was dependent on an energy source. Other lactobacilli, more osmosensitive than strain 3532, including L. acidophilus strain E4356, L. bulgaricus 8144, and L. delbrueckii 9649, showed lower betaine transport rates in response to an osmotic challenge than L. acidophilus 3532. Experiments with chloramphenicol-treated L. acidophilus 3532 cells indicated that the transport system was not induced but appeared to be activated by an increase in osmotic pressure.

Journal Article↗

Enhanced clearance of Candida albicans from the oral cavities of mice following oral administration of Lactobacillus acidophilus.

Orally administered live Lactobacillus acidophilus was assessed for its capacity to enhance clearance from the oral cavity of DBA/2 mice shown previously to be 'infection prone'. L. acidophilus fed to DBA/2 mice significantly shortened the duration of colonization of the oral cavity compared to controls. Enhanced clearance of Candida albicans correlated with both early mRNA gene expression for interleukin (IL)-4 and interferon (IFN)-gamma and expression of their secreted products in cultures of cervical lymph nodes stimulated with Candida antigen. In addition rapid clearance correlated with higher levels of IFN-gamma and nitric oxide in saliva. Delayed clearance, less pronounced levels of the cytokine response, saliva IFN-gamma and nitric oxide, and later mRNA expression for IL-4 and IFN-gamma relative to feeding with the L. acidophilus isolate were noted in mice fed a different Lactobacillus isolate (L. fermentum). These observations indicate significant variations in individual isolates to activate the common mucosal system.

Administration, Oral↗

Isolation and partial amino acid sequence of bacteriocins produced by Lactobacillus acidophilus.

Bacteriocins produced by Lactobacillus acidophilus JCM 1023, JCM 1028, JCM 1021, JCM 1229, and JCM 5342 were active against closely related lactobacilli. These bacteriocins were purified and partial sequenced. Bacteriocin activities of L. acidophilus JCM 1023 and JCM 1028 were associated with two components. On the basis of N-terminal amino acid sequencing and the molecular masses, it is interpreted that these two-component bacteriocins are identical to acidocin J1132, a bacteriocin from L. acidophilus JCM 1132 [Tahara et al., Appl. Environ. Microbiol., 62, 892-897 (1996)]. Other bacteriocins were single-peptide bacteriocins.

Amino Acid Sequence↗

Use of a continuous-culture biofilm system to study the antimicrobial susceptibilities of Gardnerella vaginalis and Lactobacillus acidophilus.

Gardnerella vaginalis and Lactobacillus acidophilus have been shown to grow to high titers in a simple biofilm system. This system was used in the present investigation to compare the biofilm-eradicating concentrations (BECs) of amoxicillin, clindamycin, erythromycin, and metronidazole to standard tube MIC and minimum bactericidal concentration (MBC) results. With the lactobacillus, the BEC/tube MBC ratio was at least 16:1, while for G. vaginalis the ratio varied from 2:1 to 512:1. The simple continuous-culture system used in the present investigation is ideal for investigating the BEC for bacteria involved in complex ecological situations such as bacterial vaginosis and may be useful for the identification of the most effective and selective antibiotic therapy.

Amoxicillin↗

Degradation of thymidine by Lactobacillus acidophilus.

Whole cells of Lactobacillus acidophilus are capable of degrading thymidine to thymine, suggesting the presence of thymidine phosphorylase (or thymidine hydrolase). This activity was also demonstrated in cell-free extracts.

Arsenic↗

Bile salt hydrolase activity of three strains of Lactobacillus acidophilus.

Three strains of Lactobacillus acidophilus, two from human intestinal origin (016 and L1) and one from porcine intestinal origin (ATCC 43121), were tested for their bile salt deconjugation activity. The L. acidophilus ATCC 43121 had more deconjugating activity of both sodium glycocholate and sodium taurocholate at pH 6.5 than did either L. acidophilus 016 or L1. The activity of intracellular bile salt hydrolase found in strain ATCC 43121 was 14-fold higher than that in either of the other two strains. The optimum pH for deconjugation of sodium glycocholate was between 4 and 5.5 for all three strains. For deconjugation of sodium taurocholate, the optimum pH was between 3.5 and 4.5 for strains L1 and ATCC 43121 and was between pH 5 and 6 for strain O16. The molecular mass of the enzyme in all three strains of L. acidophilus was estimated to be 126 kDa by Sephadex G-200 gel filtration. All three strains exhibited more bile salt hydrolase activity towards sodium glycocholate than towards sodium taurocholate.

Amidohydrolases↗

Specific antigens of Lactobacillus acidophilus.

Antigens specific for Lactobacillus acidophilus were investigated by double immunodiffusion in agar-gel. Antigenic materials were extracted from whole bacteria and some walls with cold trichloroacetic acid. Antisera were prepared by intravenous injection into rabbits of suspensions of whole organisms in solutions of bovine serum albumin, which had been heated and then washed. Four specific antigens were found as precipitinogens and denoted as antigens 11, 12, 13 and 14. Of 43 strains of L. acidophilus studied, 33 strains possessed antigen 11, six strains antigen 12, two strains antigen 13 and two strains antigen 14. Sugar compositions of wall preparations were analysed in an attempt to characterize the determinants of antigens 11 and 12. The walls contained glucose, galactose, hexosamine and sometimes glycerol, but no rhamnose was found. It was considered that alpha-glucopyranose was the major component of the determinant of antigen 11 since trehalose and maltose significantly inhibited the reaction between antibody 11 and its antigen; the determinant of antigen 12 was not clarified.

Antigen-Antibody Reactions↗

Relationship between bile tolerance and the presence of a ruthenium red staining layer on strains of Lactobacillus acidophilus.

Seventeen strains of Lactobacillus acidophilus were evaluated to determine the relationship between bile tolerance and the presence of an outer polysaccharide layer exterior to the cell wall when viewed by transmission electron microscopy. Bile tolerance is necessary for survival of lactobacilli in the intestinal tract, and the polysaccharide layer may be responsible for adherence to human intestinal tissue. These two factors may be the basis for use of L. acidophilus as a dietary adjunct. Ten strains exhibited a ruthenium red-stained outer polysaccharide layer. Three of the 10 strains had extremely dense layers, which may indicate stronger adherence properties. Seven strains did not contain a ruthenium red-stained outer layer; however, six strains that did not have the stained layer were resistant to 1.0% bile concentration. Fourteen strains were tolerant to 1% bile, one strain was tolerant to 6% bile, and two strains were sensitive to bile. No relationship between bile tolerance and the presence of the ruthenium red-stained outer polysaccharide layer was apparent.

Bacterial Adhesion↗