Cooperation between Lactococcus lactis and nonstarter lactobacilli in the formation of cheese aroma from amino acids.
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In Gouda and Cheddar type cheeses the amino acid conversion to aroma compounds, which is a major process for aroma formation, is essentially due to lactic acid bacteria (LAB). In order to evaluate the respective role of starter and nonstarter LAB and their interactions in cheese flavor formation, we compared the catabolism of phenylalanine, leucine, and methionine by single strains and strain mixtures of Lactococcus lactis subsp. cremoris NCDO763 and three mesophilic lactobacilli. Amino acid catabolism was studied in vitro at pH 5.5, by using radiolabeled amino acids as tracers. In the presence of alpha-ketoglutarate, which is essential for amino acid transamination, the lactobacillus strains degraded less amino acids than L. lactis subsp. cremoris NCDO763, and produced mainly nonaromatic metabolites. L. lactis subsp. cremoris NCDO763 produced mainly the carboxylic acids, which are important compounds for cheese aroma. However, in the reaction mixture containing glutamate, only two lactobacillus strains degraded amino acids significantly. This was due to their glutamate dehydrogenase (GDH) activity, which produced alpha-ketoglutarate from glutamate. The combination of each of the GDH-positive lactobacilli with L. lactis subsp. cremoris NCDO763 had a beneficial effect on the aroma formation. Lactobacilli initiated the conversion of amino acids by transforming them mainly to keto and hydroxy acids, which subsequently were converted to carboxylic acids by the Lactococcus strain. Therefore, we think that such cooperation between starter L. lactis and GDH-positive lactobacilli can stimulate flavor development in cheese. (+info)
Febrile gastroenteritis after eating on-farm manufactured fresh cheese--an outbreak of listeriosis?
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An outbreak of febrile gastroenteritis affected consumers of on-farm manufactured dairy products from a summer farm in Sweden. Symptoms included diarrhoea, fever, stomach cramps and vomiting in 88, 60, 54 and 21% of cases identified. The median incubation period was 31 h. A cohort study with 33 consumers showed an attack rate of 52% and an association between the total amount of product eaten and illness (P=0.07). Twenty-seven of 32 (84%) stool samples cultured for Listeria monocytogenes tested positive, although there was no association between clinical disease and the isolation of L. monocytogenes. In addition, gene sequences for VTEC and ETEC were detected in 6 and 1 subjects, respectively. Bacteriological analysis of cheese samples revealed heavy contamination with L. monocytogenes and coagulase positive staphylococci in all of them and gene markers for VTEC in one of them. Molecular profiles for L. monocytogenes isolated from dairy products, stool samples and an abscess from 1 patient who developed septic arthritis were identical. Results of both microbiological and epidemiological analyses point to L. monocytogenes as the most likely cause of this outbreak. The finding of markers for VTEC in some humans and cheese samples means that a mixed aetiology at least in some cases cannot be conclusively ruled out. (+info)
A pediocin-producing Lactobacillus plantarum strain inhibits Listeria monocytogenes in a multispecies cheese surface microbial ripening consortium.
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The growth of Listeria monocytogenes WSLC 1364, originating from a cheese-borne outbreak, was examined in the presence and in the absence of a pediocin AcH-producing Lactobacillus plantarum strain on red smear cheese. Nearly complete inhibition was observed at 10(2) CFU of L. monocytogenes per ml of salt brine solution, while contamination with Listeria mutants resistant to pediocin resulted in high cell counts of the pathogen on the cheese surface. The inhibition was due to pediocin AcH added together with the L. plantarum culture to the brine solution but not to bacteriocin production in situ on cheese. Pediocin resistance developed in vitro at different but high frequencies in all 12 L. monocytogenes strains investigated, and a resistant mutant remained stable in a microbial surface ripening consortium over a 4-month production process in the absence of selection pressure. In conclusion, the addition of a L. plantarum culture is a potent measure for combating Listeria in a contaminated production line, but because of the potential development of resistance, it should not be used continuously over a long time in a production line. (+info)
Dietary supplementation with zinc and a growth factor extract derived from bovine cheese whey improves methotrexate-damaged rat intestine.
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BACKGROUND: Oral administration of zinc or bovine whey-derived growth factor extract (WGFE) is known to reduce intestinal permeability and ameliorate methotrexate (MTX)-induced mucositis, respectively. OBJECTIVE: We examined the effects of zinc, WGFE, and zinc plus WGFE on gut damage in MTX-treated rats. DESIGN: Rats (n = 16/group) were fed zinc (1000 mg/kg diet), WGFE (32 mg/kg diet), zinc plus WGFE, or control (10 mg Zn/kg diet) diets for 7 d and then injected subcutaneously with MTX (2.5 mg/kg) for 3 d to induce gut damage. Gut histology and intestinal permeability were assessed. RESULTS: The Zn+WGFE diet was associated with both reduced gut damage on day 5 and enhanced recovery on day 7. The WGFE diet ameliorated gut damage, whereas the Zn and Zn+WGFE diets enhanced repair. Gut metallothionein and tissue zinc concentrations were significantly (P < 0.01) higher with Zn and Zn+WGFE on days 5 and 7 than without zinc supplementation. The Zn and Zn+WGFE diets significantly (P < 0.05) decreased gut permeability on days 3-4 compared with the control diet. Intestinal permeability was significantly (P < 0.05) increased on days 5-6. On days 6-7, only the WGFE diet improved gut permeability (by 80%) compared with the control diet. CONCLUSIONS: Dietary administration of WGFE and a pharmacologic dose of zinc reduced intestinal damage and enhanced recovery, respectively. WGFE also improved gut permeability after MTX-induced bowel damage. In combination, zinc and WGFE hastened repair of gut damage, which may have clinical application in chemotherapy-induced mucositis. (+info)
Purification, characterization, and gene cloning of lysyl aminoeptidase from Streptococcus thermophilus YRC001.
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We purified and characterized an aminopeptidase from Streptococcus thermophilus YRC001 to obtain an enzyme for the application of reducing bitter-defect in cheese manufacturing. The purified enzyme was a monomer, and its molecular mass was estimated to be 90-100 kDa. It had a broad substrate specificity, and mostly hydrolyzed lysyl and leucyl peptides. The optimal temperature and pH for the enzyme were 35 degrees C and pH 6.5, respectively. EDTA, o-phenanthroline, and p-chloromercuribenzoate inhibited its activity, therefore it was considered to be a metallopeptidase. The purified enzyme efficiently reduced the bitterness of a trypsin digest of reconstituted skim milk. Therefore, we cloned a gene for the enzyme from YRC001. The nucleotide sequence of a 2,940-bp XbaI fragment containing the gene was analyzed. The gene encoded 849 amino acids, and the calculated molecular mass for the mature enzyme (initial methionine is removed) was 96,434. The deduced amino acid sequence showed high homology with the known bacterial lysyl aminopeptidase (aminopeptidase N). (+info)
Bacterial community structure and location in Stilton cheese.
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The microbial diversity occurring in Stilton cheese was evaluated by 16S ribosomal DNA analysis with PCR-denaturing gradient gel electrophoresis. DNA templates for PCR experiments were directly extracted from the cheese as well as bulk cells harvested from a variety of viable-count media. The variable V3 and V4-V5 regions of the 16S genes were analyzed. Closest relatives of Lactococcus lactis, Enterococcus faecalis, Lactobacillus plantarum, Lactobacillus curvatus, Leuconostoc mesenteroides, Staphylococcus equorum, and Staphylococcus sp. were identified by sequencing of the DGGE fragments. Fluorescently labeled oligonucleotide probes were developed to detect Lactococcus lactis, Lactobacillus plantarum, and Leuconostoc mesenteroides in fluorescence in situ hybridization (FISH) experiments, and their specificity for the species occurring in the community of Stilton cheese was checked in FISH experiments carried out with reference cultures. The combined use of these probes and the bacterial probe Eub338 in FISH experiments on Stilton cheese sections allowed the assessment of the spatial distribution of the different microbial species in the dairy matrix. Microbial colonies of bacteria showed a differential location in the different parts of the cheese examined: the core, the veins, and the crust. Lactococci were found in the internal part of the veins as mixed colonies and as single colonies within the core. Lactobacillus plantarum was detected only underneath the surface, while Leuconostoc microcolonies were homogeneously distributed in all parts observed. The combined molecular approach is shown to be useful to simultaneously describe the structure and location of the bacterial flora in cheese. The differential distribution of species found suggests specific ecological reasons for the establishment of sites of actual microbial growth in the cheese, with implications of significance in understanding the ecology of food systems and with the aim of achieving optimization of the fermentation technologies as well as preservation of traditional products. (+info)
Generation of food-grade lactococcal starters which produce the lantibiotics lacticin 3147 and lacticin 481.
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Transconjugant lactococcal starters which produce both lantibiotics lacticin 3147 and lacticin 481 were generated via conjugation of large bacteriocin-encoding plasmids. A representative of one of the resultant strains proved more effective at killing Lactobacillus fermentum and inhibiting the growth of Listeria monocytogenes LO28H than either of the single bacteriocin-producing parental strains, demonstrating the potential of these transconjugants as protection cultures for food safety applications. (+info)
Comparative study using type strains and clinical and food isolates to examine hemolytic activity and occurrence of the cyl operon in enterococci.
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The hemolytic ability, the presence of cyl genes, and the diagnostic accuracy of cytolysin molecular detection were investigated in the genus Enterococcus by using 164 strains from 20 different species (26 reference strains, 42 clinical isolates from human and veterinary origin, and 96 isolates from ewe cheese and milk). Hemolysis was assayed with sheep and horse erythrocytes and under aerobic or anaerobic conditions. Screening of cytolysin genes (cylL(L), cylL(S), cylM, cylB, and cylA) was performed with new specific primers and the anaerobic assay of beta-hemolysis was used as the "gold standard" for the evaluation of cyl gene-based PCRs. Since beta-hemolysis and cyl genes were found in 10 and 14 species, respectively, the hemolytic ability seems to be spread throughout the genus ENTEROCOCCUS: Beta-hemolysis was observed in 6 of 26 (23%) reference strains, 14 of 42 (33%) clinical isolates, and 6 of 96 (6%) food isolates. The presence of cyl genes was detected in 15 of 26 (58%) reference strains, 37 of 42 (88%) clinical isolates, and 67 of 96 (70%) food isolates. These data indicate a virulence potential in food isolates, reinforcing the need of their safety assessment. Analysis of phenotypic-genotypic congruence suggests a divergent sequence evolution of cyl genes and the effect of environmental factors in the regulation of cytolysin expression. Evaluation of the diagnostic accuracy of cytolysin molecular detection points to cylL(L)-based PCR and cylL(L)L(S)MBA-based PCR as the most reliable approaches. Nevertheless, the low sensitivity (46%) and gene variability indicated by our study strongly recommend the phenotypic assay for the assessment of hemolytic ability in enterococci, followed by the molecular screening of cyl genes in nonhemolytic strains to evaluate their virulence potential. (+info)