Cloning, sequencing and expression of the gene encoding the cell-envelope-associated proteinase from Lactobacillus paracasei subsp. paracasei NCDO 151.

The gene encoding the cell-envelope-associated proteinase of Lactobacillus paracasei subsp. paracasei NCDO 151 (formerly Lactobacillus casei NCDO 151) was cloned and sequenced. The gene was located on the chromosome and encoded a polypeptide of 1902 amino acids. The proteinase is N-terminally cleaved upon maturation. It shows extensive homology to the Lactococcus lactis subsp. cremoris Wg2 proteinase. Similar to the situation in Lactococcus, a maturation gene was found upstream of the proteinase gene. The cloned proteinase gene was expressed in Lactobacillus plantarum. However, no expression was observed when the gene was cloned in Lactococcus lactis.     

Synthesis of gamma-aminobutyric acid by lactic acid bacteria isolated from a variety of Italian cheeses

The concentrations of gamma-aminobutyric acid (GABA) in 22 Italian cheese varieties that differ in several technological traits markedly varied from 0.26 to 391 mg kg(-1). Presumptive lactic acid bacteria were isolated from each cheese variety (total of 440 isolates) and screened for the capacity to synthesize GABA. Only 61 isolates showed this activity and were identified by partial sequencing of the 16S rRNA gene. Twelve species were found. Lactobacillus paracasei PF6, Lactobacillus delbrueckii subsp. bulgaricus PR1, Lactococcus lactis PU1, Lactobacillus plantarum C48, and Lactobacillus brevis PM17 were the best GABA-producing strains during fermentation of reconstituted skimmed milk. Except for L. plantarum C48, all these strains were isolated from cheeses with the highest concentrations of GABA. A core fragment of glutamate decarboxylase (GAD) DNA was isolated from L. paracasei PF6, L. delbrueckii subsp. bulgaricus PR1, L. lactis PU1, and L. plantarum C48 by using primers based on two highly conserved regions of GAD. A PCR product of ca. 540 bp was found for all the strains. The amino acid sequences deduced from nucleotide sequence analysis showed 98, 99, 90, and 85% identity to GadB of L. plantarum WCFS1 for L. paracasei PF6, L. delbrueckii subsp. bulgaricus PR1, L. lactis PU1, and L. plantarum C48, respectively. Except for L. lactis PU1, the three lactobacillus strains survived and synthesized GABA under simulated gastrointestinal conditions. The findings of this study provide a potential basis for exploiting selected cheese-related lactobacilli to develop health-promoting dairy products enriched in GABA.     

Diversity and dynamics of lactobacilli populations during ripening of RDO Camembert cheese

The diversity and dynamics of Lactobacillus populations in traditional raw milk Camembert cheese were monitored throughout the manufacturing process in 3 dairies. Culture-dependent analysis was carried out on isolates grown on acidified de Man - Rogosa - Sharpe agar and Lactobacillus anaerobic de Man Rogosa Sharpe agar supplemented with vancomycin and bromocresol green media. The isolates were identified by polymerase chain reaction - temperature gradient gel electrophoresis (PCR-TGGE) and (or) species-specific PCR and (or) sequencing, and Lactobacillus paracasei and Lactobacillus plantarum isolates were characterized by pulsed field gel electrophoresis (PFGE). Milk and cheese were subjected to culture-independent analysis by PCR-TGGE. Presumed lactobacilli were detected by plate counts throughout the ripening process. However, molecular analysis of total DNA and DNA of isolates failed to detect Lactobacillus spp. in certain cases. The dominant species in the 3 dairies was L. paracasei. PFGE analysis revealed 21 different profiles among 39 L. paracasei isolates. Lactobacillus plantarum was the second most isolated species, but it occurred nearly exclusively in one dairy. The other species isolated were Lactobacillus parabuchneri, Lactobacillus fermentum, Lactobacillus acidophilus, Lactobacillus helveticus, a Lactobacillus psittaci/delbrueckii subsp. bulgaricus/gallinarum/crispatus group, Lactobacillus rhamnosus, Lactobacillus delbrueckii subsp. bulgaricus, L. delbrueckii subsp. lactis, Lactobacillus brevis, Lactobacillus kefiri, and Lactobacillus perolens. Lactobacilli diversity at the strain level was high. Dynamics varied among dairies, and each cheese exhibited a specific picture of species and strains.     

Ribotyping of lactobacilli isolated from spoiled beer

Twenty-nine Lactobacillus strains contaminating beers in different Czech breweries as well as representative type strains obtained from the Czech Collection of Microorganisms were characterized using ribotyping with EcoRI and a probe made complementary to 16S and 23S rRNA genes. Biochemical test results assigned the 29 strains to the species L. brevis, L. plantarum, L. buchneri and L. paracasei subsp. paracasei. Ribotyping separated L. brevis, L. plantarum and L. paracasei subsp. paracasei strains into species-specific ribogroups in full correspondence with biotyping; L. buchneri strains were split into two ribogroups. Characteristic band patterns for each species and even typical bands of certain sizes were observed.     

Cell-wall-bound proteinase of Lactobacillus delbrueckii subsp. lactis ACA-DC 178: characterization and specificity for beta-casein

Lactobacillus delbrueckii subsp. lactis ACA-DC 178, which was isolated from Greek Kasseri cheese, produces a cell-wall-bound proteinase. The proteinase was removed from the cell envelope by washing the cells with a Ca2+-free buffer. The crude proteinase extract shows its highest activity at pH 6.0 and 40 degrees C. It is inhibited by phenylmethylsulfonyl fluoride, showing that the enzyme is a serine-type proteinase. Considering the substrate specificity, the enzyme is similar to the lactococcal PI-type proteinases, since it hydrolyzes beta-casein mainly and alpha- and kappa-caseins to a much lesser extent. The cell-wall-bound proteinase from L. delbrueckii subsp. lactis ACA-DC 178 liberates four main peptides from beta-casein, which have been identified.     

The proteolytic system of Lactobacillus sanfrancisco CB1: purification and characterization of a proteinase, a dipeptidase, and an aminopeptidase.

A cell envelope 57-kDa proteinase, a cytoplasmic 65-kDa dipeptidase, and a 75-kDa aminopeptidase were purified from Lactobacillus sanfrancisco CB1 sourdough lactic acid bacterium by sequential fast protein liquid chromatography steps. All of the enzymes are monomers. The proteinase was most active at pH 7.0 and 40 degrees C, while aminopeptidase and dipeptidase had optima at pH 7.5 and 30 to 35 degrees C. Relatively high activities were observed at the pH and temperature of the sourdough fermentation. The proteinase is a serine enzyme. Urea-polyacrylamide gel electrophoresis of digest of alpha s1- and beta-caseins showed differences in the pattern of peptides released by the purified proteinase and those produced by crude preparations of the cell envelope proteinases of Lactobacillus delbrueckii subsp. bulgaricus B397 and Lactococcus lactis subsp. lactis SK11. Reversed-phase fast protein liquid chromatography of gliadin digests showed a more-complex peptide pattern produced by the proteinase of Lactobacillus sanfrancisco CB1. The dipeptidase is a metalloenzyme with high affinity for dipeptides containing hydrophobic amino acids but had no activity on tripeptides or larger peptides. The aminopeptidase was also inhibited by metal-chelating agents, and showed a broad N-terminal hydrolytic activity including di- and tripeptides. Km values of 0.70 and 0.44 mM were determined for the dipeptidase on Leu-Leu and the aminopeptidase on Leu-p-nitroanilide, respectively.     

Dominant lactic acid bacteria and their technological properties isolated from the Himalayan ethnic fermented milk products

Ethnic people of the Himalayan regions of India, Nepal, Bhutan and China consume a variety of indigenous fermented milk products made from cows milk as well as yaks milk. These lesser-known ethnic fermented foods are dahi, mohi, chhurpi, somar, philu and shyow. The population of lactic acid bacteria (LAB) ranged from 10(7) to 10(8) cfu/g in these Himalayan milk products. A total of 128 isolates of LAB were isolated from 58 samples of ethnic fermented milk products collected from different places of India, Nepal and Bhutan. Based on phenotypic characterization including API sugar test, the dominant lactic acid bacteria were identified as Lactobacillus bifermentans, Lactobacillus paracasei subsp. pseudoplantarum, Lactobacillus kefir, Lactobacillus hilgardii, Lactobacillus alimentarius, Lactobacillus paracasei subsp. paracasei, Lactobacillus plantarum, Lactococcus lactis subsp. lactis, Lactococcus lactis subsp. cremoris and Enterococcus faecium. LAB produced a wide spectrum of enzymes and showed high galactosidase, leucine-arylamidase and phosphatase activities. They showed antagonistic properties against selected Gram-negative bacteria. None of the strains produced bacteriocin and biogenic amines under the test conditions used. Most strains of LAB coagulated skim milk with a moderate drop in pH. Some strains of LAB showed a high degree of hydrophobicity, suggesting these strains may have useful adhesive potential. This paper is the first report on functional lactic acid bacterial composition in some lesser-known ethnic fermented milk products of the Himalayas.     

Diversity and mechanisms of alkali tolerance in lactobacilli

We determined the maximum pH that allows growth (pHmax) for 34 strains of lactobacilli. High alkali tolerance was exhibited by strains of Lactobacillus casei, L. paracasei subsp. tolerans, L. paracasei subsp. paracasei, L. curvatus, L. pentosus, and L. plantarum that originated from plant material, with pHmax values between 8.5 and 8.9. Among these, L. casei NRIC 1917 and L. paracasei subsp. tolerans NRIC 1940 showed the highest pHmax, at 8.9. Digestive tract isolates of L. gasseri, L. johnsonii, L. reuteri, L. salivarius subsp. salicinius, and L. salivarius subsp. salivarius exhibited moderate alkali tolerance, with pHmax values between 8.1 and 8.5. Dairy isolates of L. delbrueckii subsp. bulgaricus, L. delbrueckii subsp. lactis, and L. helveticus exhibited no alkali tolerance, with pHmax values between 6.7 and 7.1. Measurement of the internal pH of representative strains revealed the formation of transmembrane proton gradients (DeltapH) in a reversed direction (i.e., acidic interior) at alkaline external-pH ranges, regardless of their degrees of alkali tolerance. Thus, the reversed DeltapH did not determine alkali tolerance diversity. However, the DeltapH contributed to alkali tolerance, as the pHmax values of several strains decreased with the addition of nigericin, which dissipates DeltapH. Although neutral external-pH values resulted in the highest glycolysis activity in the presence of nigericin regardless of alkali tolerance, substantial glucose utilization was still detected in the alkali-tolerant strains, even in a pH range of between 8.0 and 8.5, at which the remaining strains lost most activity. Therefore, the alkali tolerance of glycolysis reactions contributes greatly to the determination of alkali tolerance diversity.     

Isolation,identification and characterization of <Emphasis Type="Italic">Pediococcus pentosaceus</Emphasis> LB44 and <Emphasis Type="Italic">Weissella confusa</Emphasis> LM85 for the presence of bacteriocin-like inhibitory substances (BLIS)

Lactic acid bacteria (LAB) were isolated from different soil samples such as dairy, garden, rhizospheric soil and sediments of lake. Thirty nine strains of bacteria were isolated using direct spreading method but only one showed the characteristics of LAB. Whereas, using accumulation with incubation method, fifty seven strains were found to be LAB out of sixty eight strains of bacteria isolated. Therefore, accumulation with incubation method was found to be more effective and faster. On the basis of higher growth and antimicrobial activity, two potential isolates LB44 and LM85 were selected for the further characterization. Strain LB44 and LM85 were identified using 16S rDNA amplification and sequencing as Pediococcus pentosaceus LB44 and Weissella confusa LM85, respectively. Cell free supernatant (CFS) of these strains showed antimicrobial activity, stability at higher temperatures, acidic pH and in the presence of organic solvents, detergents and surfactants. Antimicrobial activity was not affected by treatment with catalase and lipase but reduced after treatment with proteinase K reveals the presence of bacteriocin-like inhibitory substance in CFS. CFS inhibited the growth of other Gram-positive and Gram-negative bacteria such as Micrococcus luteus, Lactobacillus delbrueckii, L. curvatus, L. plantarum, L. acidophilus, Enterobacter cloaceae, Lactococcus lactis subsp. cremoris, L. lactis subsp. lactis, Bacillus subtilis, B. licheniformis, Salmonella typhi and Escherichia coli.     

Viili中益生菌的分离及其发酵性质研究

目的 利用传统方法和分子生物学方法从viili中筛菌并对其特性进行研究.方法 采用MRS、YPD和脱脂乳营养琼脂3种平板从viili中分离出15株单菌,利用V3区通用引物和乳杆菌特异性引物对各单菌的PCR产物进行变性梯度凝胶电泳(denaturing gradient gel electrophoresis,DGGE)分析,将其归为5种不同菌.结果 16S DNA测序表明,5种单菌分别为Lactobacillus plantarum、Streptococcus thermophilus、Lactobacillus paracasei、Bacillus cereus和Lactobacillus delbrueckii subsp. bulgaricus.混合发酵实验表明:从viili中筛出的5种菌除Bacillus cereus外均有凝乳现象,其中Lactobacillus delbrueckii凝乳能力强,可在5 h内凝乳;Lactobacillus paracasei、Bacillus cereus和Lactobacillus plantarum组合产生的EPS量最多,高达186.71 mg/L,而Streptococcus thermophilus EPS产量仅为33.56 mg/L.结论 传统方法与分子生物学方法DGGE相结合,可以快速准确判断细菌种类;筛选菌特性研究结果表明,细菌之间的相互作用导致凝乳时间和EPS产量发生变化,其复合作用有待于进一步研究.     

Phenotypic and genotypic characteristics of lactic acid bacteria isolated from sour congee in Inner Mongolia of China

Sour congee is a popular food in the western regions of Inner Mongolia in China. It has a complex microbial population, which contributes to its unique flavor and functional properties. In this study, we chose 28 sour congee samples that were collected from different areas in Inner Mongolia for analysis of the microbial community of lactic acid bacteria (LAB) by classical biochemical tests, 16S rRNA gene sequencing, multiplex PCR assay of recA gene and restriction fragment length polymorphism (RFLP) analysis of the tuf gene (encoding elongation factor Tu). The results revealed that all the isolates were identified as Lactobacillus (L.) paracasei (38 strains), L. fermentum (28 strains), L. plantarum (7 strains), L. brevis (4 strains), L. reuteri (2 strains), L. amylolyticus (1 strain), Enterococcus (E.) faecalis (3 strains), E. italicus (2 strains) or Lactococcus lactis subsp. lactis (1 strain). The predominant LAB were L. casei and L. fermentum in sour congee samples. The diversity of LAB derived from sour congee could offer useful information for further research on sour congee, and the results demonstrated that the combination of tuf gene and RFLP patterns can be considered as a useful tool for differentiation of the L. casei group.     

Induction of systemic and mucosal immune response and decrease in Streptococcus pneumoniae colonization by nasal inoculation of mice with recombinant lactic acid bacteria expressing pneumococcal surface antigen A

Mucosal epithelia constitute the first barriers to be overcome by pathogens during infection. The induction of protective IgA in this location is important for the prevention of infection and can be achieved through different mucosal immunization strategies. Lactic acid bacteria have been tested in the last few years as live vectors for the delivery of antigens at mucosal sites, with promising results. In this work, Streptococcus pneumoniae PsaA antigen was expressed in different species of lactic acid bacteria, such as Lactococcus lactis, Lactobacillus casei, Lactobacillus plantarum, and Lactobacillus helveticus. After nasal inoculation of C57Bl/6 mice, their ability to induce both systemic (IgG in serum) and mucosal (IgA in saliva, nasal and bronchial washes) anti-PsaA antibodies was determined. Immunization with L. lactis MG1363 induced very low levels of IgA and IgG, possibly by the low amount of PsaA expressed in this strain and its short persistence in the nasal mucosa. All three lactobacilli persisted in the nasal mucosa for 3 days and produced a similar amount of PsaA protein (150-250 ng per 10(9) CFU). However, L. plantarum NCDO1193 and L. helveticus ATCC15009 elicited the highest antibody response (IgA and IgG). Vaccination with recombinant lactobacilli but not with recombinant L. lactis led to a decrease in S. pneumoniae recovery from nasal mucosa upon a colonization challenge. Our results confirm that certain Lactobacillus strains have intrinsic properties that make them suitable candidates for mucosal vaccination experiments.     

Control of Listeria monocytogenes in a biofilm by competitive-exclusion microorganisms

Biofilms from drains in food processing facilities with a recent history of no detectable Listeria monocytogenes in floor drains were cultured for microorganisms producing antilisterial metabolites. A total of 413 microbial isolates were obtained from 12 drain biofilm samples and were assayed at 15 and 37 degrees C for activities that were bactericidal or inhibitory to L. monocytogenes, by two agar plate assays. Twenty-one of 257 bacterial isolates and 3 of 156 yeast isolates had antilisterial activity. All 24 isolates which produced metabolites inhibitory to L. monocytogenes were assayed for antilisterial activity in coinoculated broth cultures containing tryptic soy broth with yeast extract (TSB-YE). A five-strain mixture of 10(3) CFU of L. monocytogenes/ml and 10(5) CFU of the candidate competitive-exclusion microorganism/ml was combined in TSB-YE and incubated at 37 degrees C for 24 h, 15 degrees C for 14 days, 8 degrees C for 21 days, and 4 degrees C for 28 days. Substantial inhibition of L. monocytogenes growth (4 to 5 log CFU/ml) was observed for nine bacterial isolates at 37 degrees C, two at 15 and 8 degrees C, and three at 4 degrees C. The inhibitory isolates were identified as Enterococcus durans (six isolates), Lactococcus lactis subsp. lactis (two isolates), and Lactobacillus plantarum (one isolate). The anti-L. monocytogenes activity of these isolates was evaluated in biofilms of L. monocytogenes on stainless steel coupons at 37, 15, 8, and 4 degrees C. Results revealed that two isolates (E. durans strain 152 and L. lactis subsp. lactis strain C-1-92) were highly inhibitory to L. monocytogenes (growth inhibition of >5 log(10) CFU of L. monocytogenes/cm(2)). These two bacterial isolates appear to be excellent competitive-exclusion candidates to control L. monocytogenes in biofilms at environmental temperatures of 4 to 37 degrees C.     

Bioconversion of oleuropein to hydroxytyrosol by lactic acid bacteria

The aim of this work is to study the conversion of oleuropein-a polyphenol present in olives and olive oil by-products-into hydroxytyrosol, a polyphenol with antioxidant and antibacterial properties. The hydrolysis reaction is performed by lactic acid bacteria. Six bacterial strains (Lactobacillus plantarum 6907, Lactobacillus paracasei 9192, Lactobacillus casei, Bifidobacterium lactis BO, Enterococcus faecium 32, Lactobacillus LAFTI 10) were tested under aerobic and anaerobic conditions. The oleuropein degradation and hydroxytyrosol formation were monitored by HPLC. Results showed that oleuropein could be successfully converted into hydroxytyrosol. The most effective strain was Lactobacillus plantarum 6907, with a reaction yield of hydroxytyrosol of about 30 %. Different reaction mechanisms were observed for different microorganisms; a different yield was observed for Lactobacillus paracasei 9192 under aerobic or anaerobic conditions and an intermediate metabolite (oleuropein aglycone) was detected for Lactobacillus paracasei 9192 and Lactobacillus plantarum 6907 only. This study could have significant applications, as this reaction can be used to increase the value of olive oil by-products and/or to improve the taste of unripe olives.     

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.     

Probiotic properties of Lactobacillus strains isolated from the feces of breast-fed infants and Taiwanese pickled cabbage

This study assessed potential probiotic Lactobacillus strains isolated from the feces of breast-fed infants and from Taiwanese pickled cabbage for their possible use in probiotic fermented foods by evaluating their (i) in vitro adhesive ability, resistance to biotic stress, resistance to pathogenic bacteria, and production of β-galactosidase; (ii) milk technological properties; and (iii) in vivo adhesive ability, intestinal survival and microbial changes during and after treatment. Five Lactobacillus isolates identified as Lactobacillus reuteri F03, Lactobacillus paracasei F08, Lactobacillus rhamnosus F14, Lactobacillus plantarum C06, and Lactobacillus acidophilus C11 that showed resistance to gastric juice and bile salts were selected for further evaluation of their probiotic properties. All the strains demonstrated the ability to adhere to Caco-2 cells, particularly, strain L. plantarum C06 and L. reuteri F03 showed satisfactory abilities, which were similar to that of the reference strain L. rhamnosus GG. The strains L. paracasei F08 and L. acidophilus C11 had the highest β-galactosidase activity. Most of the strains were resistant to aminoglycosides and vancomycin but sensitive to ampicillin, erythromycin, and penicillin. All the 5 strains elicited antibacterial activity against both Gram-positive (Bacillus cereus, Listeria monocytogenes and Staphylococcus aureus) and -negative (Escherichia coli and Salmonella enterica) pathogens. Moreover, the strains L. reuteri F03, L. paracasei F08, and L. plantarum C06 could grow rapidly in milk without nutrient supplementation and reached 10? cfu/mL after 24 h of fermentation at 37 °C. The viable cell counts of the 3 strains remained above 10? cfu/mL after 21 d of storage at 4 °C. In the animal feeding trial, the number of intestinal lactobacilli increased significantly after administration of milk fermented with the 3 strains, and the counts of fecal coliforms and Clostridium perfringens were markedly reduced. Lactobacillus strains could also survive in the ileal intestinal tissue of the treated rats. Technologically interesting Lactobacillus isolates may be used in the future as probiotic starter cultures for manufacturing novel fermented foods.     

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.     

Preliminary characterization of lactic acid bacteria isolated from Zlatar cheese

AIMS: Isolation, characterization and identification of lactic acid bacteria (LAB) from artisanal Zlatar cheese during the ripening process and selection of strains with good technological characteristics. METHODS AND RESULTS: Characterization of LAB was performed based on morphological, physiological and biochemical assays, as well as, by determining proteolytic activity and plasmid profile. rep-polymerase chain reaction (PCR) analysis and 16S rDNA sequencing were used for the identification of LAB. PCR analysis was performed with specific primers for detection of the gene encoding nisin production. Strains Lactobacillus paracasei subsp. paracasei, Lactobacillus plantarum, Lactobacillus brevis, Lactococcus lactis subsp. lactis, Enterococcus faecium and Enterococcus faecalis were the main groups present in the Zlatar cheese during ripening. CONCLUSIONS: Temporal changes in the species were observed during the Zlatar cheese ripening. Mesophilic lactobacilli are predominant microflora in Zlatar cheese. SIGNIFICANCE AND IMPACT OF THE STUDY: In this study we determined that Zlatar cheese up to 30 days old could be used as a source of strains for the preparation of potential starter cultures in the process of industrial cheese production. As the Serbian food market is adjusting to European Union regulations, the standardization of Zlatar cheese production by using starter culture(s) based on autochtonous well-characterized LAB will enable the industrial production of this popular cheese in the future.     

Heat resistance of Lactobacillus spp. isolated from Cheddar cheese

Mesophilic Lactobacillus spp. are the dominant organisms in mature Cheddar cheese. The heat resistance of broth grown cultures of Lactobacillus plantarum DPC1919 at temperatures between 50 and 57.5 degrees C, Lact. plantarum DPC2102 at temperatures between 48 and 56 degrees C and Lact. paracasei DPC2103 at temperatures between 50 and 67.5 degrees C was determined. The z-values for Lact. plantarum DPC1919, Lact. Plantarum DPC2102 and Lact. paracasei DPC2103 were 6.7 degrees C, 6.2 degrees C and 5.3 degrees C, respectively. Lactobacillus paracasei DPC2103 showed evidence of injury and recovery, especially at higher temperatures. Milk grown cultures of strains DPC2102 and DPC2103 showed greater heat resistance than broth grown cultures, tailing of the death curves and a nonlinear z-curve. Of the three strains, Lact. paracasei DPC2103 had the potential to survive pasteurization temperatures, whether grown in milk or broth.     

Isolation and identification of lactic acid bacteria with effect of immune protection to Eschericia coli in mice

Lactic acid bacteria were isolated from an alcohol fermentation broth, and the activity as a probiotic was examined using pathogenic E. coli. Thirty-six strains exhibiting good growth were isolated in the medium of concentrated mush which was a residue resulted in the alcohol distillation process. One of these strains, Lactobacillus paracasei subsp. paracasei I-5, could be grown in the medium containing 8 vol% ethanol and at 45 degrees C. The characteristics were different from the type strain, L. paracasei subsp. paracasei NBRC 15889. L. paracasei I-5 showed an excellent growth in the concentrated mush, which just diluted two-fold and adjusted the pH. ICR mice were fed with a standard germ-free feed (CMF) and the strain I-5 (7 x 10(9) cells/day) was orally administrated for 11 days prior to the intraperitoneal challenge with pathogenic E. coli Juhl. After the challenge, mice administrated the strain I-5 exhibited a high survival rate and survival extension days (p < 0.01) compared with the control. The results suggested that the strain might enhance the animal resistance against microbial pathogens. Neonatal diarrhea caused by E. coli is a serious disease in calf breeding. The strain might be practically valuable to prevent diarrhea in calves.