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PubMed2006

The effect of prebiotics on production of antimicrobial compounds, resistance to growth at low pH and in the presence of bile, and adhesion of probiotic cells to intestinal mucus.

AIMS: Screening of five bile salt-resistant and low pH-tolerant lactic acid bacteria for inhibitory activity against lactic acid bacteria and bacterial strains isolated from the faeces of children with HIV/AIDS. Determining the effect of prebiotics and soy milk-base on cell viability and adhesion of cells to intestinal mucus. METHODS AND RESULTS: Lactobacillus plantarum 423, Lactobacillus casei LHS, Lactobacillus salivarius 241, Lactobacillus curvatus DF 38 and Pediococcus pentosaceus 34 produced the highest level of antimicrobial activity (12,800 AU ml(-1)) when grown in MRS broth supplemented with 2% (m/v) dextrose. Growth in the presence of Raftilose Synergy1, Raftilose L95 and Raftiline GR did not lead to increased levels of antimicrobial activity. Cells grown in the presence of Raftilose Synergy1 took longer to adhere to intestinal mucus, whilst cells grown in the absence of prebiotics showed a linear rate of binding. CONCLUSIONS: A broad range of gram-positive and gram-negative bacteria were inhibited. Dextrose stimulated the production of antimicrobial compounds. Adhesion to intestinal mucus did not increase with the addition of prebiotics. SIGNIFICANCE AND IMPACT OF THE STUDY: The strains may be incorporated in food supplements for HIV/AIDS patients suffering from gastro-intestinal disorders.

Anti-Bacterial Agents↗
PubMed2006

Isolation and characterization of lactic acid bacteria from suan-tsai (fermented mustard), a traditional fermented food in Taiwan.

AIMS: To isolate, characterize, and identify lactic acid bacteria (LAB) in suan-tsai (fermented mustard), a traditional fermented food in Taiwan. METHODS AND RESULTS: Suan-tsai samples were collected at five time points from a fixed fermenting bucket. Fifty cultures were isolated from suan-tsai samples, and isolates were divided into classes by phenotype and then into groups by restriction-fragment length polymorphism analysis and sequencing of 16S ribosomal DNA. Phenotypic and biochemical characteristics identified two different bacterial groups (A and B), and the results showed that Pediococcus pentosaceus was the most abundant LAB during the initial fermentation time. However, the more NaCl-tolerant species Tetragenococcus halophilus took the place of P. pentosaceus and became the most abundant LAB later. All isolates were grown in de Man, Rogosa, and Sharpe (MRS) broth containing 6% NaCl, but T. halophilus could grow only in MRS broth containing 10% NaCl. CONCLUSIONS: These results suggest that the LAB P. pentosaceus and T. halophilus play roles in the fermentation of suan-tsai. SIGNIFICANCE AND IMPACT OF THE STUDY: This is the first report describing the distribution and varieties of LAB that exist in the suan-tsai fermentation process.

Colony Count, Microbial↗
PubMed1994

Simple method for extracting plasmid DNA from lactic acid bacteria.

Rapid screening and large-scale plasmid DNA isolation procedures are described for lactic acid bacteria, using glass beads to break cells. The rapid screening procedure allows one to obtain plasmid DNA pellets in less than 1 h. This method has been successfully tested on various bacteria from the genera Lactococcus, Leuconostoc, Lactobacillus, Pediococcus, Streptococcus, Enterococcus and Propionibacterium. This procedure yields plasmid DNA with minor chromosomal and plasmid DNA-degraded form contaminations.

Bacteriological Techniques↗
PubMed2004

Comparative analysis of conserved genetic markers and adjacent DNA regions identified in beer-spoilage lactic acid bacteria.

AIMS: To conduct an inter-species comparative study on the nucleotide sequences of the conserved DNA regions surrounding ORF5, a genetic marker for differentiating beer-spoilage lactic acid bacteria. METHODS AND RESULTS: The conserved DNA regions surrounding ORF5 were examined by PCR analysis, using three beer-spoilage strains, Lactobacillus brevis ABBC45C, L. paracollinoides LA2T and Pediococcus damnosus ABBC478. As a result, the DNA regions containing ORF1-7, originally found in ABBC45C, appeared to be conserved among the three strains, while the downstream region was not found in L. paracollinoides LA2T and P. damnosus ABBC478. The sequencing analysis of the conserved DNA regions of LA2T and ABBC478 revealed ca 99% nucleotide sequence identities with that of ABBC45C. CONCLUSIONS: The nucleotide sequences of the ca 8.2 kb DNA regions containing ORF1-7 were virtually identical among the three strains belonging to different species. The internal organizations of the ORFs were found to be remarkably similar. SIGNIFICANCE AND IMPACT OF THE STUDY: The level of nucleotide sequence identities suggests the DNA regions surrounding ORF5 were horizontally acquired by these beer-spoilage strains belonging to the three different species of lactic acid bacteria.

Beer↗
PubMed2004

Isolation and characterization of anti-Salmonella lactic acid bacteria from the porcine gastrointestinal tract.

AIMS: To identify lactic acid bacteria (LAB) of porcine intestinal origin with anti-Salmonella activity. METHODS AND RESULTS: Samples were obtained from pig faeces and caeca and screened for the presence of anti-Salmonella LAB. The 11 most promising isolates were identified as belonging to the genera Lactobacillus and Pediococcus. The LAB exhibited large variation in their ability to survive in simulated gastric juice at pH 1.85. While Lactobacillus johnsonii species survived at levels of 80% for up to 30 min, Lactobacillus pentosus species declined to <0.001% in that time. All isolates tolerated porcine bile at a concentration of 0.3% (w/v), with some isolates capable of growth in the presence of up to 5% (w/v) bile. The ability of the LAB isolates to prevent Salmonella invasion of intestinal epithelial HT-29 cells varied, with reductions of between 30% (Lact. pentosus) and 80% (Lactobacillus murinus spp.) observed. CONCLUSIONS: LAB of porcine origin were observed to survive simulated passage through the GIT and inhibit growth of Salmonella and its invasion of the intestinal epithelium. SIGNIFICANCE AND IMPACT OF THE STUDY: The data demonstrate that some porcine intestinal LAB isolates may offer potential as probiotics for the reduction of Salmonella carriage in pigs.

Animals↗
PubMed2006

Gluten breakdown by lactobacilli and pediococci strains isolated from sourdough.

AIMS: To evaluate the growth and metabolic activity of lactobacilli and pediococci strains in a gluten base medium (GBM), formulated for a proper selection of proteolytic strains to be used in sourdough fermentation. METHODS AND RESULTS: Proteolytic activity by lactic acid bacteria (LAB) was evaluated by SDS-PAGE and by the amino acids released determined by reversed-phase high-performance liquid chromatography. Only 13 LAB (nine lactobacilli and four pediococci), among the 42 evaluated were able to utilize gluten as nitrogen source and to grow in GBM. Pediococcus pentosaceus CRL 797 showed a similar proteolytic activity to lactobacilli strains. In the majority of the cultures, basic amino acid group increased (c. 80% after 12 h incubation) mainly due to the release of ornithine, a flavour precursor of bread. Lysine, a limiting essential amino acid in wheat flour, increased by 150% in cultures of P. pentosaceus CRL 797. CONCLUSIONS: This study allows selecting P. pentosaceus CRL 797 and L. plantarum CRL 759 as potential starter culture for type III sourdough fermentation. It is shown for the first time that pediococci strains isolated from sourdough are proteolytically active on gluten. SIGNIFICANCE AND IMPACT OF THE STUDY: The physiological studies on gluten breakdown by LAB will contribute to the better selection of strains to produce breads with enhanced organoleptic characteristics.

Amino Acids↗
PubMed2006

Diversity of a stable enrichment culture which is useful for silage inoculant and its succession in alfalfa silage.

Alfalfa is a kind of forage that is difficult to ensile for good quality. Therefore, inoculants are always used to enhance the preservation of alfalfa silage. Through continuous restricted subcultivation, a lactic acid bacteria community (Al2) was selected from well-fermented alfalfa silage, which sharply decreased the pH level and produced a large amount of lactic acid. The adding of Al2 to alfalfa at ensiling resulted in a more rapid drop in pH and higher levels of lactic acid, and it also reduced the ammonia-nitrogen content significantly (P < 0.01). Plate isolation, denaturing gradient gel electrophoresis (DGGE) and the construction of a 16S rRNA gene clone library were used to identify the composition diversity of the Al2 community; seven strains were detected in the community, the predominant strain belonging to Lactobacillus plantarum. Samples of alfalfa silages of duration 0, 2, 5, 10, 20 and 30 days were studied with DGGE analysis. The DGGE band patterns of Al2-treated and non inoculated were rather different, and the components of Al2 were the dominant bacteria in Al2-treated silages, especially L. plantarum, while Pediococcus pentosaceaus was predominant in naturally fermented alfalfa silage.

Comet Assay↗
PubMed2000

Acetaldehyde metabolism by wine lactic acid bacteria.

Acetaldehyde is a volatile flavor compound present in many fermented foods and is important in the production of red and white wines. Nine strains of the genera Lactobacillus and Oenococcus were able to metabolize acetaldehyde in a resting cell system, whereas two Pediococcus strains were not. Acetic acid and ethanol were produced from its degradation. A Lactobacillus and an Oenococcus were able to degrade SO(2)-bound acetaldehyde, as well. A coincubation of resting cells of Saccharomyces bayanus Première Cuvée and Oenococcus oeni Lo111 showed that strain Lo111 metabolized acetaldehyde produced by the yeast. The ability of malolactic bacteria to degrade free and SO(2)-bound acetaldehyde has implications for sensory and color qualities and the use of SO(2) in wine.

Acetaldehyde↗
PubMed2005

Extremely low electrical current generated by porcine small intestine smooth muscle alters bacterial autolysin production.

The effect of extremely low electrical currents, identical to those generated by the gut smooth muscle, on bacterial autolysin production in vitro was tested in the present study. When stimulated with these electrical currents, the bacteria Pediococcus pentosaceus 16:1 produced groups of peptidoglycan hydrolases that differed from those produced by the unstimulated (control) bacteria. The autolysins synthesized by the P. pentosaceus 16:1 under extremely low electrical currents were effective against peptidoglycans from the cell walls of various lactic acid bacteria strains, whereas the autolysins from the control bacteria acted exclusively against P. pentosaceus 16:1 cell walls. Thus, it can be predicted that in vivo the electrical currents generated by the intestinal smooth muscles, which can be recorded as the myoelectrical migrating complexes, could regulate lactic acid bacteria strain growth in the gut.

Animals↗
PubMed1972

Cross-resistance and collateral susceptibility to antifolic antimalarial compounds.

Series of strains of Streptococcus faecium ATCC 8043, Lactobacillus casei ATCC 7469, and Pediococcus cerevisiae ATCC 8081 with increasing resistance to the active antifolate antimalarial drugs chlorguanide triazine (CGT), pyrimethamine (PM), and trimethoprim (TMP) were isolated. These mutant strains, stable for at least 3 to 5 years, were examined for cross-resistance and collateral susceptibility to the above compounds and to methotrexate (MTX). Generally, they exhibited cross-resistance to all four compounds, but resistance of a strain to one compound did not predict accurately its resistance to another drug. Unexpectedly, L. casei resistant to CGT exhibited collateral susceptibility to MTX, TMP, and PM varying from 5- to 20-fold. P. cerevisiae developed resistance to CGT readily but maintained its susceptibility to PM and TMP after prolonged exposure to these compounds. Resistance to these antimalarial antifolates was accompanied by only low-grade cross-resistance to MTX, a representative antileukemic antifolate agent.

Antimalarials↗
PubMed2006

Biodiversity of bacterial ecosystems in traditional Egyptian Domiati cheese.

Bacterial biodiversity occurring in traditional Egyptian soft Domiati cheese was studied by PCR-temporal temperature gel electrophoresis (TTGE) and PCR-denaturing gradient gel electrophoresis (DGGE). Bands were identified using a reference species database (J.-C. Ogier et al., Appl. Environ. Microbiol. 70:5628-5643, 2004); de novo bands having nonidentified migration patterns were identified by DNA sequencing. Results reveal a novel bacterial profile and extensive bacterial biodiversity in Domiati cheeses, as reflected by the numerous bands present in TTGE and DGGE patterns. The dominant lactic acid bacteria (LAB) identified were as follows: Leuconostoc mesenteroides, Lactococcus garvieae, Aerococcus viridans, Lactobacillus versmoldensis, Pediococcus inopinatus, and Lactococcus lactis. Frequent non-LAB species included numerous coagulase-negative staphylococci, Vibrio spp., Kocuria rhizophila, Kocuria kristinae, Kocuria halotolerans, Arthrobacter spp./Brachybacterium tyrofermentans. This is the first time that the majority of these species has been identified in Domiati cheese. Nearly all the dominant and frequent bacterial species are salt tolerant, and several correspond to known marine bacteria. As Domiati cheese contains 5.4 to 9.5% NaCl, we suggest that these bacteria are likely to have an important role in the ripening process. This first systematic study of the microbial composition of Domiati cheeses reveals great biodiversity and evokes a role for marine bacteria in determining cheese type.

Biodiversity↗
PubMed1985

Lactate metabolism by pediococci isolated from cheese.

Pediococcus pentosaceus is commonly found among the adventitious microflora of Cheddar cheese. When this organism was incubated with L-(+)-lactate under anaerobic conditions, L-(+)-lactate was rapidly converted to D-(-)-lactate until racemic (DL) lactate was present. Under aerobic conditions this initial reaction was followed by a slower reaction resulting in the use of both lactate isomers and in the production of acetate and CO2. With intact cells the lactate oxidation system had an optimum pH of 5 to 6, depending on the initial lactate concentration. Cells grown anaerobically possessed lactate-oxidizing activity which increased two- to fourfold as sugar was exhausted from the medium. Aerobic growth further increased specific activities. Cheddar cheese was made with the deliberate addition of P. pentosaceus. When the resulting cheese was grated to expose a large surface area to O2, lactate was converted to acetate at a rate which depended on the density of pediococci in the cheese. The lactate oxidation system remained active in cheese which had been ripened for 6 months.

Acetates↗
PubMed1991

Bacteriocin-mediated inhibition of Clostridium botulinum spores by lactic acid bacteria at refrigeration and abuse temperatures.

The bacteriocinogenicity of Lactococcus lactis ATCC 11454, Pediococcus pentosaceus ATCC 43200, P. pentosaceus ATCC 43201, Lactobacillus plantarum BN, L. plantarum LB592, L. plantarum LB75, and Lactobacillus acidophilus N2 against Clostridium botulinum spores at 4, 10, 15, and 35 degrees C was investigated by modified deferred and simultaneous antagonism methods. All the strains, except L. acidophilus N2, produced inhibition zones on lawns of C. botulinum spores at 30 degrees C. L. plantarum BN, L. lactis ATCC 11454, and P. pentosaceus ATCC 43200 and 43201 were bacteriocinogenic at 4, 10, and 15 degrees C. Supplementation of brain heart infusion agar with 0 to 5% NaCl increased the radii of inhibition zones during simultaneous antagonism assays. Detectable bacteriocin activities were extracted from freeze-thawed agar cultures of L. plantarum BN and L. lactis ATCC 11454 which had been grown at 4 and 10 degrees C. These results suggest that low levels of L. plantarum BN or L. lactis ATCC 11454, in the presence of 3 or 4% NaCl, could be formulated into minimally processed refrigerated food products for protection against possible botulism hazards.

Bacteriocins↗
PubMed1992

Enhanced control of Listeria monocytogenes by in situ-produced pediocin during dry fermented sausage production.

To determine whether pediocin is produced and has effective antilisterial activity during food fermentation, six sausage fermentation trials were conducted with antibiotic-resistant, pediocin-producing (Bac+) Pediococcus acidilactici PAC 1.0 (Strr Rifr) and an isogenic pediocin-negative (Bac-) derivative used as a control. Meat was inoculated (ca. 10(5) CFU/g) with a composite of five Listeria monocytogenes strains, each electrotransformed with pGK12 (Cmr Emr). P. acidilactici and L. monocytogenes populations were selectively enumerated by plating on media with antibiotics. This study indicated that the dry sausage fermentation process can reduce L. monocytogenes populations. Effective inactivation of L. monocytogenes was observed when the pH at the end of the fermentation portion of the process was less than 4.9. Pediocin was responsible for part of the antilisterial activity during the fermentation in each of the six trials. Furthermore, inhibition of L. monocytogenes during drying was enhanced in the presence of pediocin in the three trials in which L. monocytogenes could be detected throughout the drying process. Thus, pediocin production contributed to an increase in safety during both the fermentation and drying portions of sausage manufacturing.

Animals↗
PubMed1998

Isolation and characterization of pediocin AcH chimeric protein mutants with altered bactericidal activity.

A collection of pediocin AcH amino acid substitution mutants was generated by PCR random mutagenesis of DNA encoding the bacteriocin. Mutants were isolated by cloning mutagenized DNA into an Escherichia coli malE plasmid that directs the secretion of maltose binding protein-pediocin AcH chimeric proteins and by screening transformant colonies for bactericidal activity against Lactobacillus plantarum NCDO955 (K. W. Miller, R. Schamber, Y. Chen, and B. Ray, 1998. Appl. Environ. Microbiol. 64:14-20, 1998). In all, 17 substitution mutants were isolated at 14 of the 44 amino acids of pediocin AcH. Seven mutants (N5K, C9R, C14S, C14Y, G37E, G37R, and C44W) were completely inactive against the pediocin AcH-sensitive strains L. plantarum NCDO955, Listeria innocua Lin11, Enterococcus faecalis M1, Pediococcus acidilactici LB42, and Leuconostoc mesenteroides Ly. A C24S substitution mutant constructed by other means also was inactive against these bacteria. Nine other mutants (K1N, W18R, I26T, M31T, A34D, N41K, H42L, K43N, and K43E) retained from <1% to approximately 60% of wild-type activity when assayed against L. innocua Lin11. One mutant, K11E, displayed approximately 2. 8-fold-higher activity against this indicator. About one half of the mutations mapped to amino acids that are conserved in the pediocin-like family of bacteriocins. All four cysteines were found to be required for activity, although only C9 and C14 are conserved among pediocin-like bacteriocins. Several basic amino acids as well as nonpolar amino acids located within the hydrophobic C-terminal region also were found to be important. The mutations are discussed in the context of structural models that have been proposed for the bacteriocin.

ATP-Binding Cassette Transporters↗
PubMed2000

Biochemical and genetic characterization of coagulin, a new antilisterial bacteriocin in the pediocin family of bacteriocins, produced by Bacillus coagulans I(4).

A plasmid-linked antimicrobial peptide, named coagulin, produced by Bacillus coagulans I(4) has recently been reported (B. Hyronimus, C. Le Marrec and M. C. Urdaci, J. Appl. Microbiol. 85:42-50, 1998). In the present study, the complete, unambiguous primary amino acid sequence of the peptide was obtained by a combination of both N-terminal sequencing of purified peptide and the complete sequence deduced from the structural gene harbored by plasmid I(4). Data revealed that this peptide of 44 residues has an amino acid sequence similar to that described for pediocins AcH and PA-1, produced by different Pediococcus acidilactici strains and 100% identical. Coagulin and pediocin differed only by a single amino acid at their C terminus. Analysis of the genetic determinants revealed the presence, on the pI(4) DNA, of the entire 3.5-kb operon of four genes described for pediocin AcH and PA-1 production. No extended homology was observed between pSMB74 from P. acidilactici and pI(4) when analyzing the regions upstream and downstream of the operon. An oppositely oriented gene immediately dowstream of the bacteriocin operon specifies a 474-amino-acid protein which shows homology to Mob-Pre (plasmid recombination enzyme) proteins encoded by several small plasmids extracted from gram-positive bacteria. This is the first report of a pediocin-like peptide appearing naturally in a non-lactic acid bacterium genus.

Amino Acid Sequence↗
PubMed2001

Optimization of annealing temperature to reduce bias caused by a primer mismatch in multitemplate PCR.

To reduce PCR bias derived from a primer mismatch, the effect of the annealing temperature on the product ratio was investigated by denaturing gradient gel electrophoresis analysis of PCR products from a mixture of perfect-match and one-mismatch templates. These templates were generated by PCR from Pediococcus acidilactici for one mismatch and Micrococcus luteus for the perfect match. PCRs showed that the bias was reduced at lower temperatures. An environmental sample was also examined.

Base Pair Mismatch↗
PubMed2003

Purification and genetic characterization of plantaricin NC8, a novel coculture-inducible two-peptide bacteriocin from Lactobacillus plantarum NC8.

A new, coculture-inducible two-peptide bacteriocin named plantaricin NC8 (PLNC8) was isolated from Lactobacillus plantarum NC8 cultures which had been induced with Lactococcus lactis MG1363 or Pediococcus pentosaceus FBB63. This bacteriocin consists of two distinct peptides, named alpha and beta, which were separated by C(2)-C(18) reverse-phase chromatography and whose complementary action is necessary for full plantaricin NC8 activity. N-terminal sequencing of both purified peptides showed 28 and 34 amino acids residues for PLNC8 alpha and PLNC8 beta, respectively, which showed no sequence similarity to other known bacteriocins. Mass spectrometry analysis showed molecular masses of 3,587 Da (alpha) and 4,000 Da (beta). The corresponding genes, designated plNC8A and plNC8B, were sequenced, and their nucleotide sequences revealed that both peptides are produced as bacteriocin precursors of 47 and 55 amino acids, respectively, which include N-terminal leader sequences of the double-glycine type. The mature alpha and beta peptides contain 29 and 34 amino acids, respectively. An open reading frame, orfC, which encodes a putative immunity protein was found downstream of plNC8B and overlapping plNC8A. Upstream of the putative -35 region of plNC8B, two direct repeats of 9 bp were identified, which agrees with the consensus sequence and structure of promoters of class II bacteriocin operons whose expression is dependent on an autoinduction mechanism.

Amino Acid Sequence↗