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Lactic acid bacteria: hydrophobicity and strength of attachment to meat surfaces.

The hydrophobicity and strength of attachment of several lactic acid bacteria with antimicrobial activity were studied. Hydrophobicity was determined by bacterial adherence to hydrocarbons (BATH; octane or xylene), adhesion to nitrocellulose filters (NCF), salt aggregation test (SAT) and adherence to phenyl-Sepharose beads (PSB). The relative hydrophobicity of lactic acid bacteria depended markedly on the method used. No correlation between either SAT or BATH (octane) and strength of attachment (Sr value) existed. However, a significant relationship between strength of attachment and BATH (xylene), NCF and PSB, respectively, was observed, showing the highest correlation coefficient (r = 0.778) for BATH (xylene).

Ammonium Sulfate↗

Variations in tolerance of Listeria monocytogenes to nisin, pediocin PA-1 and bavaricin A.

A collection of 381 Listeria monocytogenes strains was examined for strain variations in nisin and pediocin sensitivity at three different concentrations on Tryptic Soya Agar. Two of the strains were able to grow on agar plates containing 500 IU nisin ml-1. These strains were characterized as having an enhanced nisin tolerance. Twenty strains had normal growth on agar plates containing 1600 AU pediocin ml-1 and higher. These strains were characterized as pediocin-resistant. Another 34 strains, characterized as having enhanced tolerance, exhibited inhibited growth at 1600 AU pediocin ml-1. Bavaricin sensitivity, examined for 22 strains of L. monocytogenes, correlated with the pediocin sensitivity of the strains. It is therefore assumed that there is accordance between the pediocin and bavaricin sensitivity of the remaining strains. Cross-resistance between nisin and pediocin/bavaricin was not found.

Abattoirs↗

Lactobacillus farciminis MD, a newer strain with potential for bacteriocin and antibiotic assay.

A native isolate Lactobacillus farciminis MD isolated from fermenting mushroom exhibited a high degree of sensitivity to the majority of the bacteriocins produced by strains of lactobacilli, leuconostoc and pediococci. Also, the efficacy of Lact. farciminis MD as a sensitive strain for antibiotic assay was established against different antibiotics including ampicillin, cefazoline, chloramphenicol and nitrofurantoin at concentrations of 30 microg each, showing an inhibition zone of 30 mm diameter. The high degree of sensitivity towards bacteriocins and antibiotics provide potential for the exploitation of Lact. farciminis MD in establishing very well-defined bacteriocin producers.

Agaricales↗

Nisin and pediocin production on mussel-processing waste supplemented with glucose and five nitrogen sources.

AIMS: Optimization of bacteriocin production by L. lactis subsp. lactis CECT 539 and Ped. acidilactici NRRL B-5627 on mussel-processing wastes. METHODS AND RESULTS: The concentrations of glucose and five nitrogen sources that optimize nisin and pediocin production were determined using a second order orthogonal factorial design. NH4Cl, glycine and glutamic acid were poor nitrogen sources. Enhanced pediocin and nisin productions were achieved in a medium supplemented with yeast extract or Bacto casitone. CONCLUSIONS: Mussel-processing wastes could be successfully used as a culture medium for bacteriocin production at low production cost. SIGNIFICANCE AND IMPACT OF THE STUDY: Average optimization led to a threefold increase in pediocin production (from 322 to 934 BU ml(-1)) and in nisin production (from 32 to 100 BU ml(-1)) when compared with the unsupplemented medium. For this reason, mussel-processing waste warrants further investigation due to its potential use as a cheap culture medium for upscaling bacteriocin production.

Animals↗

Prevalence and diversity of tetracycline resistant lactic acid bacteria and their tet genes along the process line of fermented dry sausages.

In order to study the prevalence and diversity of tetracycline resistant lactic acid bacteria (Tc(r) LAB) along the process line of two different fermented dry sausage (FDS) types, samples from the raw meat, the meat batter and the fermented end product were analysed quantitatively and qualitatively by using a culture-dependent approach. Both the diversity of the tet genes and their bacterial hosts in the different stages of FDS production were determined. Quantitative analysis showed that all raw meat components of both FDS types (FDS-01 and FDS-08) contained a subpopulation of Tc(r) LAB, and that for FDS-01 no Tc(r) LAB could be recovered from the samples after fermentation. Qualitative analysis of the Tc(r) LAB subpopulation in FDS-08 included identification and typing of Tc(r) LAB isolates by (GTG)5-PCR fingerprinting, plasmid profiling, protein profiling and a characterization of the resistance by PCR detection of tet genes. Two remarks can be made when the results of this analysis for the different samples are compared. (i) The taxonomic diversity of Tc(r) LAB varies along the process line, with a higher diversity in the raw meat (lactococci, lactobacilli, streptococci, and enterococci), and a decrease after fermentation (only lactobacilli). (ii) Also the genetic diversity of the tet genes varies along the process line. Both tet(M) and tet(S) were found in the raw meat, whereas only tet(M) was found after fermentation. A possible relationship was found between the disappearing of species other than lactobacilli and tet(S), because tet(S) was only found in lacotocci, enterococci, and streptococci. These data suggest that fermented dry sausages are among those food products that can serve as vehicles for Tc(r) LAB and that the raw meat already contains a subpopulation of these bacteria. Whether these results reflect the transfer of resistant bacteria or of bacterial resistance genes from animals to man via the food chain is difficult to ascertain and may require a combination of cultivation-dependent and cultivation-independent approaches.

Animals↗

Development of a fast DNA-DNA hybridization method based on melting profiles in microplates.

DNA-DNA hybridization is still the "gold standard" for the genotypic delineation of bacterial species. However, it is not widely used because traditional DNA-DNA hybridization techniques are rather time-consuming and not easy to perform in routine laboratories. In the present study, DNA of reference strains was digested with Sau3A, ligated with linker oligonucleotides S1/2 and in vitro amplified. The amplified DNA fragments were immobilized on MaxiSorb 96-well plates. DNA isolated from target strains was also digested with Sau3A, ligated with linker oligonuleotides P1/2 and in vitro amplified in the presence of digoxygenin modified dUTP. The labeled amplificate was hybridized to the immobilized reference DNA under isothermal conditions. Thermal denaturation curves of the DNA-DNA hybrids were obtained by using washing solutions of increasing stringency. Remaining hybrids were colorimetrically detected with anti-digoxygenin-horseradish peroxidase anti-bodies. The new method was validated with strains of the genus Pedioccocus for which DNA-DNA similarities have also been determined by the filter hybridization method. In addition, DNA-DNA hybridizations were performed with genotypically defined Enterobacter species.

Bacteria↗

An improved procedure for the assay of available lysine and methionine in feedstuffs using Tetrahymena pyriformis W.

1. A study was made of the assay of available lysine and available methionine using Tetrahymena pyriformis W, as applied to a variety of protein-rich feedstuffs. 2. Results were strongly influenced by the conditions under which the analyses were done. With most of the test samples predigestion with papain caused a large increase in the values obtained. 3. An improved assay procedure was developed which included enzymic predigestion of the test samples, "all-in" sterilization of the medium constituents, and measurement of growth from extinction values. It gave results for available lysine that were broadly similar to those obtained in growth tests with rats, and in chemical tests by the method of Carpenter (1960). Values for available methionine agreed closely with those obtained in microbiological tests with Streptococcus zymogenes.

Animal Feed↗

Comparison of chemical and microbiological methods in the estimation of methionine in cowpea (Vigna unguiculata) seeds.

1. Meals were prepared from the seeds of fifteen varieties of cowpea (Vigna unguiculata), one of lima bean (Phaseolus lunatus) and one of yam bean (Sphenostylis stenocarpa), and their methionine content was determined by six different methods. 2. Total methionine content was determined by two chemical methods (ion-exchange chromatography and a colorimetric procedure) and by two microbiological methods. The 'available' methionine content was determined by microbiological assay with Streptococcus zymogenes. 3. All the different methods for total methionine determination gave similar results, with much the same high extent of precision. 4. The value for 'available' methionine content were similar to or marginally higher than the corresponding microbiological assay value for total methionine content. There was no indication that the methionine in any of the test samples was not completely available.

Biological Assay↗

Environmental stress response in wine lactic acid bacteria: beyond Bacillus subtilis.

Lactic acid bacteria (LAB) constitute a heterogeneous group of bacteria that are traditionally used to produce fermented foods. The industrialization of food transformations has increased the economical importance of LAB, as they play a crucial role in the development of the organoleptic and hygienic quality of fermented products. However, the strains selected for industrial purposes, should tolerate adverse conditions encountered in industrial processes, either during starter handling and storage (freeze-drying, freezing, or spray-drying) or during food processing in which abiotic stresses such as heat, cold, acidity, and high concentration of NaCl or ethanol are common. Wine LAB have to deal with several stresses including an acidic pH, a high alcoholic content, non optimal growth temperatures, and growth-inhibitory compounds such as fatty acids and tannins, originated from yeast and bacteria metabolism. Wine LAB have developed several mechanisms to escape or to tolerate wine conditions. They carry out a malolactic fermentation in this stressful environment. In addition to the regulation of the expression of specific genes, bacteria have evolved adaptive networks to face the challenges of a changing environment and to survive under conditions of stress. The so called Global Regulatory Systems control the simultaneous expression of a large number of genes in response to a variety of environmental stress factors. CIRCE sequences able to bind the HrcA repressor, sigma(B) dependent promoters and CtsR regulatory elements have been observed in several genes identified from wine LAB. Improved knowledge of regulators and a better understanding of LAB stress responses could constitute a basis of comparison with the well known model microorganisms, Escherichia coli and Bacillus subtilis. Moreover, it can provide an important insight into improving current industrial starter strains.

Bacillus subtilis↗

Pediocin PA-1, a wide-spectrum bacteriocin from lactic acid bacteria.

Pediocin PA-1 is a broad-spectrum lactic acid bacteria bacteriocin that shows a particularly strong activity against Listeria monocytogenes, a foodborne pathogen of special concern among the food industries. This antimicrobial peptide is the most extensively studied class Ila (or pediocin family) bacteriocin, and it has been sufficiently well characterized to be used as a food biopreservative. This review focuses on the progress that have been made in the elucidation of its structure, mode of action, and biosynthesis, and includes an overview of its applications in food systems. The aspects that need further research are also addressed. In the future, protein engineering, genetic engineering and/or chemical synthesis may lead to the development of new antimicrobial peptides with improved properties, based on some features of the pediocin PA-1 molecule.

Amino Acid Sequence↗

Regulation of the glutamate racemase of Escherichia coli investigated by site-directed mutagenesis.

The biosynthesis of D-glutamic acid, one of the essential components of bacterial cell-wall peptidoglycan, is catalyzed by a glutamate racemase in Escherichia coli. While the other reported glutamate racemases from various (essentially gram-positive) bacterial species did not require any specific activator, the E. coli enzyme absolutely requires the presence of the peptidoglycan precursor UDP-N-acetylmuramyl-L-alanine to catalyze the interconversion of glutamic acid isomers. A comparison of the amino acid sequences of these different enzymes was made to identify amino acid residues from the E. coli enzyme that are involved in the catalysis or binding to the activator. Site-directed mutagenesis experiments are described that demonstrate the participation of cysteines 96 and 208 in the two-base reaction mechanism of the enzyme. The construction of N- or C-terminal-truncated enzymes is also described. The attractive hypothesis that the characteristic N-terminal amino acid extension (20 residues) of the E. coli enzyme could be involved in its activation by the nucleotide precursor is disproved by these experiments.

Amino Acid Isomerases↗

Experimental production of elevated serum folate in dogs with intestinal blind loops. II. Nature of bacterially produced folate coenzymes in blind loop fluid.

Jejunal and ileal blind loops were constructed in separate groups of dogs and the folate coenzymes present in these loops were investigated. Although only those dogs with high blind loops had appreciably elevated folate levels active for Lactobacillus casei but not Streptococcus faecalis, every blind loop appeared to contain four folate compounds--5-formyltetrahydrofolic acid, 5-methyltetrahydrofolic acid, tetrahydrofolic acid and a compound tentatively identified as 5-formyltetrahydrofolic triglutamate. Why increased levels of 5-methyltetrahydrofolate alone are present in the serum of dogs with a high blind loop is discussed.

Animals↗

Real-time PCR detection of lactic acid bacteria in cecal contents of eimeria tenella-lnfected broilers fed soybean oligosaccharides and soluble soybean polysaccharides.

This experiment was conducted to test whether dietary soybean meal oligosaccharides (SMO) and water-soluble polysaccharides (SMP) can assist broiler chickens in resisting Eimeria tenella, and to determine the survival of lactic acid bacteria in cecal contents postinfection. All birds received a soybean meal-free diet. The 6 experimental treatments were as follows: positive (COR) and negative (COW) control groups, 2 groups fed diets containing either 1% SMO or 0.5% SMP from 1 to 11 d of age; a vaccinated group (VAC), and an anticoccidial medicated group (ANT). Chickens of all treatments except COW were orally infected with 1000 sporulated oocysts of E. tenella on d 15. Fecal oocyst shedding was monitored per treatment group between d 5 and 13 postinfection. Lactic acid bacteria (LAB) in cecal contents were evaluated by a real-time PCR technique on d 7 postinfection. The results showed that the SMO and SMP groups had a lower number of oocysts per gram of feces during the monitoring period than the COR group. Threshold cycles were 22.21, 27.68, 13.99, 14.92, 12.97, and 14.85, for COW, COR, SMO, SMP, VAC, and ANT groups, respectively; specific PCR products were confirmed by the results of melting curve analysis and agarose gel electrophoresis. The results suggest that these LAB communities were promoted by SMO and SMP and have a competitive exclusion function when broiler chickens are infected with E. tenella.

Animal Feed↗

Tetragenococcus muriaticus sp. nov., a new moderately halophilic lactic acid bacterium isolated from fermented squid liver sauce.

A total of 11 strains of moderately halophilic histamine-producing bacteria isolated from fermented squid liver sauce were studied phenotypically, genotypically, and phylogenetically. These strains are considered members of the genus Tetragenococcus based on their physiological, morphological, and chemotaxonomic characteristics. A 16S rRNA gene sequence analysis showed that these strains clustered with, but were separate from, Tetragenococcus halophilus. The results of DNA-DNA hybridization experiments indicated that the new isolates represent a new Tetragenococcus species, for which we propose the name Tetragenococcus muriaticus; strain X-1 (= JCM 10006) is the type strain of this species.

Animals↗