猪源乳酸菌的分离、鉴定及其生物学特性

【目的】将分离自猪肠道粘膜、食糜和粪便的乳酸菌,通过产乳酸能力、生长性能、耐酸和耐胆盐性能及抑菌能力评价,筛选适应养猪生产的潜在益生特性的菌株。【方法】共分离获得155株乳酸菌纯菌株,从中筛选出4株产酸能力较强的乳酸菌,结合生理生化试验及细菌16S rRNA测序鉴定其种属,评价候选乳酸菌的生长情况、耐酸、耐胆盐及抑菌特性。【结果】综合变色时间(8 h)、pH值(3.9)和乳酸含量(100 mmol/L),筛选出4株(L45、L47、L63和L79)候选菌株,经鉴定依次为罗伊氏乳杆菌、植物乳杆菌、约氏乳杆菌和粪肠球菌。该4株乳酸菌均可在体外快速生长;L47和L79能够耐受pH 2.5的酸性环境,L47能够耐受0.5%胆盐环境;各乳酸菌上清液与指示菌共培养,发现对E coli K88和沙门氏菌均产生了抑制作用,其中L47上清液对指示菌的抑制作用较强。【结论】L47具有较好的产酸性能与生长性能、可耐受猪胃酸和肠道胆盐环境,对E.coli K88和沙门氏菌具有较好的抑制作用,说明该乳酸菌具有潜在的益生特性。     

Characterization and antioxidant ability of potential probiotic lactic acid bacteria in ogi liquor and lemon juice-ogi liquor

Purpose

Ogi is an indigenous edible fermented cereal slurry but the steep liquor is usually wasted or administered as therapeutic to suppress certain illnesses. The combination of lemon juice and ogi steep liquor (OSL) is known to possess bioactive metabolites.

Method

This study evaluated potential probiotic lactic acid bacteria (LAB) in different OSL (Zea mays, Sorghum bicolor, and Pennisetum glaucum L.) and lemon juice-ogi steep liquor (LJOSL) based on low pH, bile and lysozyme tolerances, hydrophobicity and auto-aggregation, antibiotic, cholesterol removal, exopolysaccharide production, β-galactosidase, and antimicrobial and hemolytic activities using standard methods. Presumptive LAB were sequenced and assayed for radical scavenging using 2,2-diphenyl-1-picrylhydrazyl (DPPH) and lipid peroxidation inhibitory (LPI) tests.

Results

Presumptive LAB counts were higher in maize OSL (0 h:5.09 log CFU/ml) and combined cereal OSL (24–48 h:7.65 and 7.72 log CFU/ml) but decreased in all steep liquors at 72 h, except in millet OSL (7.72 log CFU/ml). A total of 120 LAB isolates were randomly selected. Based on pH and bile tolerances, 14 isolates were comparable to reference strains. All these isolates demonstrated probiotics properties except for three that did not show γ-hemolysis. Sequenced LAB isolates were identified as Lactobacillus plantarum, Lactobacillus fermentum, Pediococcus pentosaceus, and Weissella cibara. DPPH activities of LAB gradually increased during fermentation with the highest activity of DPPH (58.77%) and LPI (57.94%) activity in L. plantarum. Strong correlations were found between DPPH and LPI in all the selected isolates.

Conclusion

The antioxidant property of probiotic LAB in OSL and LJOSL could contribute to its therapeutic nature.

     

Assessment and comparison of probiotic potential of four Lactobacillus species isolated from feces samples of Iranian infants

The probiotic potential of Lactobacillus species isolated from infant feces was investigated. For this study, the antibiotic susceptibility, tolerance in gut‐related conditions, antimicrobial activity, and ability to adhere to a human colorectal adenocarcinoma cell line (Caco‐2 cells) of four common Lactobacillus species (Lactobacillus paracasei [n = 15], Lactobacillus rhamnosus [n = 45], Lactobacillus gasseri [n = 20] and Lactobacillus fermentum [n = 18]) were assessed. Most isolates that which were sensitive to imipenem, ampicillin, gentamycin, erythromycin and tetracycline were selected for other tests. L. gasseri isolates had the greatest sensitivity to gastric and intestinal fluids (<10% viability). L. fermentum (FH5, FH13 and FH18) had the highest adhesion to Caco‐2 cells. The lowest antibacterial activity against pathogenic bacteria was shown by L. gasseri strains in spot tests. Furthermore, non‐adjusted cell‐free culture supernatants with low pH had greater antimicrobial activity, which was related to organic acid. The results showed that some isolates of L. rhamnosus and L. fermentum are suitable for use as a probiotic.     

Complete glutathione system in probiotic <Emphasis Type="Italic">Lactobacillus fermentum</Emphasis> ME-3

There is much information about glutathione (GSH) in eukaryotic cells, but relatively little is known about GSH in prokaryotes. Without GSH and glutathione redox cycle lactic acid bacteria (LAB) cannot protect themselves against reactive oxygen species. Previously we have shown the presence of GSH in Lactobacillus fermentum ME-3 (DSM14241). Results of this study show that probiotic L. fermentum ME-3 contains both glutathione peroxidase and glutathione reductase. We also present that L. fermentum ME-3 can transport GSH from environment and synthesize GSH. This means that it is characterized by a complete glutathione system: synthesis, uptake and redox turnover ability that makes L. fermentum ME-3 a perfect protector against oxidative stress. To our best knowledge studies on existence of the complete glutathione system in probiotic LAB strains are still absent and glutathione synthesis in them has not been demonstrated.     

In Vitro Assessment of Probiotic Potential of Saccharomyces Cerevisiae DABRP5 Isolated from Bollo Batter,a Traditional Goan Fermented Food

Bollo is a traditional Goan fermented food in which coarse wheat/wheat and finger millet is leavened with toddy. We here isolated 42 yeast strains from Bollo batter. Initial screening of the isolates with probiotic properties yielded four yeast isolates (DABRP1, DABRP2, DABRP5 and DABRP12). These isolates exhibited tolerance to high bile salt concentration and acidic pH and resistance to various antibiotics, which indicated their probiotic nature. All these yeast isolates were identified as Saccharomyces cerevisiae through D1D2-LSU-rDNA sequencing. These yeast isolates also showed higher percent hydrophobicity towards chloroform followed by n-hexadecane and o-xylene indicating their mucosal surface-adhesive property. To evaluate the safety of the isolates for them to be called as generally recognized as safe, the pathogenic behavior of the isolates determined through the in vitro hemolysis assay and evaluation of DNase and gelatinase activities. None of the isolates exhibited hemolysis or produced DNase or gelatinase and thus were considered potentially safe. In terms of beneficial effects, the most potent isolate S. cerevisiae DABRP5 showed antibacterial activity against the test pathogens. It also showed excellent antioxidant activity with DPPH free radical scavenging activity of 68.85 ± 0.69%, anti-inflammatory activity with 60.39 ± 0.34% inhibition of protein denaturation, and antidiabetic activity with 71.75 ± 0.45% inhibition of α-amylase activity. The isolate produced α-amylase, lipase, and β-galactosidase. The probiotic potential of the isolate S. cerevisiae DABRP5 was similar to that of the reference strain (Saccharomyces boulardii CNCM I-745) used in this study. The results thus indicate that yeast isolates from Bollo batter have probiotic potential.

     

Identification and functional properties of dominant lactic acid bacteria isolated at different stages of solid state fermentation of cassava during traditional <Emphasis Type="Italic">gari</Emphasis> production

Culture-based technique was used to study the population dynamics of the bacteria and determine the dominant lactic acid bacteria (LAB) during cassava fermentation. LAB was consistently isolated from the fermented mash with an initial viable count of 6.00 log c.f.u. g⁻¹ observed at 12 h. The aerobic viable count of amylolytic lactic acid bacteria (ALAB) was higher than other group of LAB throughout the fermentation up to 96 h with the highest viable count of 8.08 log c.f.u. g⁻¹. Combination of phenotypic parameters and 16S rDNA gene sequencing identified the dominant group of LAB as Lactobacillus plantarum, L. fermentum and Leuconostoc mesenteroides while the pulse field gel electrophoresis determined that the strains were genotypically heterogeneous. The sugar fermentation profile of the isolates showed that indigestible sugars such as raffinose and stachyose can be fermented by the strains. Information was also generated about the functional properties of the strains. Only strain L. plantarum 9st0 isolate at 0 h of the fermentation produced bacteriocin with antagonism against closely related indicator strains. Quantitatively, the highest amylase activity was produced by strain L. plantarum 7st12, while appreciable amylase was also produced by L. fermentum 1st96. The result of this work showed that selection of mixed starter cultures of bacteriocin- and amylase-producing L. plantarum and L. fermentum will be highly relevant as starter cultures during the intermediate and large scale gari production.     

Screening and molecular identification of potential probiotic lactic acid bacteria in effluents generated during ogi production

Screening and molecular identification of probiotic lactic acid bacteria (LAB) in effluents generated during the production of ogi, a fermented cereal (maize, millet, and sorghum) were done. LAB were isolated from effluents generated during the first and second fermentation stages in ogi production. Bacterial strains isolated were identified microscopically and phenotypically using standard methods. Probiotic potential properties of the isolated LAB were investigated in terms of their resistance to pH 1.5 and 0.3% bile salt concentration for 4 h. The potential LAB isolates ability to inhibit the growth of pathogenic organisms (Escherichia coli, Staphylococcus aureus, and Salmonella typhimurium) was evaluated in vitro. The pH and LAB count in the effluents ranged from 3.31 to 4.49 and 3.67 to 4.72 log cfu/ml, respectively. A total of 88 LAB isolates were obtained from the effluents and only 10 LAB isolates remained viable at pH 1.5 and 0.3% bile salt. The zones of inhibition of the LAB isolates with probiotic potential ranged from 7.00 to 24.70 mm against test organsisms. Probiotic potential LAB isolates were molecularly identified as Lactobacillus plantarum, Lactobacillus fermentum, Lactobacillus reuteri, Enterococcus faecium, Pediococcus acidilactici, Pediococcus pentosaceus, Enterococcus faecalis, and Lactobacillus brevis. Survival and proliferation of LAB isolates at low pH, 0.3% bile salt condition, and their inhibition against some test pathogens showed that these LAB isolates could be a potential probiotics for research and commercial purposes.     

Comparison of Campylobacter jejuni isolates from human, food, veterinary and environmental sources in Iceland using PFGE, MLST and fla-SVR sequencing

Aims: To evaluate the probiotic properties of strains isolated from smoked salmon and previously identified as bacteriocin producers. Methods and Results: Strains Lactobacillus curvatus ET06, ET30 and ET31, Lactobacillus fermentum ET35, Lactobacillus delbrueckii ET32, Pediococcus acidilactici ET34 and Enterococcus faecium ET05, ET12 and ET88 survived conditions simulating the gastrointestinal tract (GIT) and produced bacteriocins active against several strains of Listeria monocytogenes, but presented very low activity against other lactic acid bacteria (LAB). Cell‐free supernatants containing bacteriocins, added to 3‐h‐old cultures of L. monocytogenes 603, suppressed growth over 12 h. Auto‐aggregation was strain‐specific, and values ranged from 7·2% for ET35 to 12·1% for ET05. Various degrees of co‐aggregation with L. monocytogenes 603, Lactobacillus sakei ATCC 15521 and Enterococcus faecalis ATCC 19443 were observed. Adherence of the bacteriocinogenic strains to Caco‐2 cells was within the range reported for Lactobacillus rhamnosus GG, a well‐known probiotic. The highest levels of hydrophobicity were recorded for Lact. curvatus (61·9–64·6%), Lact. fermentum (78·9%), Lact. delbrueckii (43·7%) and Ped. acidilactici (51·3%), which are higher than the one recorded for Lact. rhamnosus GG (53·3%). These strains were highly sensitive to several antibiotics and affected by several drugs from different generic groups in a strain‐dependent manner. Conclusions: Smoked salmon is a rich source of probiotic LAB. All strains survived conditions simulating the GIT and produced bacteriocins active against various pathogens. Adherence to Caco‐2 cells was within the range reported for Lact. rhamnosus GG, a well‐known probiotic. In addition, the high hydrophobicity readings recorded define the strains as good probiotics. Significance and Impact of the Study: Smoked salmon contains a number of different probiotic LAB and could be marketed as having a potential beneficial effect.     

Effect of acid lactic bacteria isolated from faeces of healthy dogs on growth parameters and aflatoxin B<Subscript>1</Subscript> production by <Emphasis Type="Italic">Aspergillus</Emphasis> species in vitro

The aim of the present study was to evaluate the inhibitory effect of Enterococcus faecium and Lactococcus lactis subsp. lactis isolated from faeces of healthy dogs on (i) lag phase, (ii) growth rate, and (iii) aflatoxin B₁ production by Aspergillus section Flavi on in vitro assays. Thirteen lactic acid bacteria (LAB) isolates were used as antagonist microorganisms. Antagonistic activity was assayed against four potentially aflatoxigenic Aspergillus section Flavi isolates: A. flavus (AF210 and AF281), A. parasiticus (AP245) and A. parasiticus (NRRL 2999). In general, the longest lag phases of Aspergillus isolates were obtained with E. faecium GJ40. Respecting the growth rate, no significant reduction was found in this parameter in the interaction assays with A. flavus and antagonist isolates respecting the control. While in A. parasiticus a significant reduction in growth rate was only observed in the interaction among reference strain and E. faecium MF5 isolate (p < 0.05). In general, AFB₁ production was reduced by most of the LAB isolates assayed, except for E. faecium GJ18, GJ20, MF3 and MF4. This study provides the first data about the antiaflatoxigenic activity of autochthonous LAB isolated from dog faeces.     

圈养大熊猫肠道乳酸菌分离及益生特性

【背景】大熊猫数量稀少、繁殖困难,在其生长过程中极易感染消化系统疾病甚至死亡。【目的】分析圈养大熊猫肠道可培养乳酸菌的群落结构与功能,筛选具有益生特性的乳酸菌菌株,为大熊猫消化系统疾病的预防和肠道微生物研究提供理论参考及菌种资源。【方法】采用3种培养基分离大熊猫粪便中的乳酸菌,通过革兰氏染色镜检和过氧化氢试验对分离的菌株进行初步鉴定,基于BOXA1R-PCR图谱遗传多样性选取代表菌株进行16S rRNA基因测序分析并进行主成分分析(principal component analysis, PCA),同时分析乳酸菌菌株安全性和益生特性。【结果】通过初步鉴定共分离获得58株乳酸菌,根据BOXA1R-PCR结果挑选20株菌进行测序,结果显示20株菌分属于明串珠菌属(Leuconostoc)、肠球菌属(Enterococcus)、魏斯氏菌属(Weissella)和链球菌属(Streptococcus)这4个属,主成分分析结果表明不同年龄段的大熊猫肠道乳酸菌群落结构存在差异。20株乳酸菌均不溶血,17株乳酸菌对11种抗生素均敏感;11株菌耐pH值2.0酸性条件,14株菌对0.3%的胆盐具有良好...     

Characterization and comparative genomic analysis of gamma-aminobutyric acid (GABA)-producing lactic acid bacteria from Thai fermented foods

Objectives

This study aimed to screen, characterize, and annotate the genome along with the comparison of GABA synthesis genes presented in lactic acid bacteria (LAB).

Results

Thirty-five LAB isolates from fermented foods were screened for GABA production using thin-layer chromatography (TLC). Fifteen isolates produced GABA ranging from 0.07 to 22.94 g/L. Based on their GTG₅ profiles, phenotypic, and genotypic characteristics, isolates LSI1-1, LSI1-5, LSI2-1, LSI2-2, LSI2-3, LSI2-5, and LSM3-1-4 were identified as Lactobacillus plantarum subsp. plantarum; isolate LSM1-4 was Lactobacillus argentoratensis; isolates CAB1-2, CAB1-5, CAB1-7, and LSI1-4 were Lactobacillus pentosus; and CAB1-1, LSM3-1-1 and LSM3-2-3 were Lactobacillus fermentum. Strains LSI2-1 and CAB1-7 from pickled vegetables were selected for genome analysis. The gadA gene (1410 bp, 470aa) was encountered in GABA production of both strains and no other glutamate decarboxylase (GAD) genes were found in the genomes when compared with other LAB strains. The presence of gadA is evidence for GABA production. Strains LSI2-1 and CAB1-7 produced 22.94 g/L and 11.59 g/L of GABA in GYP broth supplemented with 3% (w/v) MSG at 30 °C for 72 h, respectively.

Conclusions

Our report highlights the characterization of LAB and GABA production of L. plantarum LSI2-1 strain with its GABA synthesis gene.

Graphic abstract

GABA production of strains LSI2-1 and CAB1-7 in GYP broth with 3% (w/v) MSG and comparative GAD genes

     

Diversity and Probiotic Potential of Lactic Acid Bacteria Isolated from Horreh,a Traditional Iranian Fermented Food

The aim of this study was to evaluate the probiotic potential of lactic acid bacteria (LAB) strains isolated from Horreh. Some probiotic properties, e.g., resistance to acid, bile tolerance, antibacterial activity, and antibiotic susceptibility, were investigated. A total of 140 Gram-positive and catalase-negative isolates from Horreh were subjected to identification and grouping by cultural methods and the 16S rRNA sequencing. The new isolates were identified to be Lactobacillus (fermentum, plantarum, and brevis) Weissella cibaria, Enterococcus (faecium and faecalis), Leuconostoc (citreum and mesenteroides subsp. mesenteroides) and Pediococcus pentosaceus. Probiotic potential study of LAB isolates showed that Lb. plantarum and Leu. mesenteroides subsp. mesenteroides isolates were able to grow at pH 2.5 and 3.5. Lactobacillus plantarum (isolate A44) showed the highest cell hydrophobicity (84.5%). According to antibacterial activity tests, Listeria innocua and Staphylococcus aureus were the most sensitive indicators against the selected LAB strains, while Escherichia coli and Bacillus cereus were the most resistant. In addition, all the isolated LAB species were resistant to vancomycin. The results of the present study suggested that the Lactobacillus fermentum and plantarum isolated from Horreh, characterized in this study, have potential use for industrial purposes as probiotics.     

Lactobacillus rhamnosus Strain GG Modulates Intestinal Absorption, Fecal Excretion, and Toxicity of Aflatoxin B1 in Rats

In this study, the modulation of aflatoxin B₁ (AFB₁) uptake in rats by administration of the probiotic Lactobacillus rhamnosus GG was demonstrated. Fecal AFB₁ excretion in GG-treated rats was increased via bacterial AFB₁ binding. Furthermore, AFB₁-associated growth faltering and liver injury were alleviated with GG treatment.     

Functional Characterization of Potential Probiotic Lactic Acid Bacteria Isolated from <Emphasis Type="Italic">Kalarei</Emphasis> and Development of Probiotic Fermented Oat Flour

Considerable variations among probiotics with respect to their health benefitting attributes fuel the research on bioprospecting of proficient probiotic strains from various ecological niches especially the poorly unexplored ones. In the current study, kalarei, an indigenous cheese-like fermented milk product, and other dairy-based sources like curd and raw milk were used for isolation of lactic acid bacteria (LAB). Among 34 LAB isolates, 7 that could withstand simulated gastrointestinal (GI) conditions were characterized for functional probiotic attributes, viz. adhesion ability, aggregation and coaggregation, extracellular enzyme producing capability, antibacterial activity against pathogens and antibiotic resistance. The isolate M-13 (from kalarei) which exhibited most of the desirable probiotic functional properties was identified as Lactobacillus plantarum based on 16S ribosomal DNA sequence analysis and designated as L. plantarum M-13. The sequence was submitted to GenBank (accession number KT592509). The study presents the first ever report of isolation of potential probiotic LAB, i.e. L. plantarum M-13 from indigenous food kalarei, and its application for development of potential probiotic fermented oat flour (PFOF). PFOF was analysed for parameters like viability of L. plantarum M-13, acidity and pH. Results show that PFOF serves as a good matrix for potential probiotic L. plantarum M-13 as it supported adequate growth of the organism (14.4 log cfu/ml after 72 h of fermentation). In addition, appreciable acid production by L. plantarum M-13 and consequential pH reduction indicates the vigorous and active metabolic status of the potential probiotic organism in the food matrix. Thus, study shows that fermented oat flour may possibly be developed as a potential probiotic carrier especially in view of the problems associated with dairy products as probiotic vehicles.     

Molecular cloning, characterization and heterologous expression of bile salt hydrolase (Bsh) from Lactobacillus fermentum NCDO394

Bile salt hydrolase (Bsh) active probiotic strains hydrolyze bile acid amino conjugates in vivo, which triggers cholesterol consumption in liver to synthesize new bile leading to consequential cholesterol lowering. Hence, bile salt hydrolyzing potential was the criterion to select L. fermentum NCDO394 for this study and its gene encoding Bsh was identified and cloned. The resulting nucleotide sequence of bsh gene contained an open reading frame (ORF) of 978 nucleotides encoding a predicted protein of 325 amino acids with a theoretical pI of 6.39. Moreover, deduced Bsh protein had high similarity with the Bshs of L. fermentum only and also exhibited significant similarity to the Pencillin V amidases of other Lactobacillus spp. Five catalytically important amino acids were highly conserved in L. fermentum Bsh while four amino acid motifs around these active sites, were not as consistent as in other Bsh proteins. Furthermore, L. fermentum bsh gene was sub-cloned into pET-28b(+) vector, and its expression was induced with 0.05 mM isopropylthiogalactopyranoside (IPTG) in Escherichia coli BL21(DE3). The recombinant Bsh (rBsh) was purified with homogeneity using Ni⁺²-NTA column and characterized for substrate specificity, pH and temperature. The rBsh hydrolyzed six major human bile salts with a slight preference towards glycine-conjugated bile salts. The optimum pH of rBsh was six, and its enzymatic activity declined below pH 5 and above pH 7. The enzyme was stable and functional even at 65 °C while showed its maximum activity at 37 °C. In conclusion, L. fermentum NCDO394 may be a promising candidate probiotic which may affect cholesterol metabolism in vivo.     

A selected probiotic strain of <Emphasis Type="Italic">Lactobacillus fermentum</Emphasis> CM33 isolated from breast-fed infants as a potential source of β-galactosidase for prebiotic oligosaccharide synthesis

Lactic acid bacteria from healthy breast-fed infants were isolated and screened for β-galactosidase production in MRS broth. Among 49 isolates that exhibited the yellow clear zone on MRS agar supplemented with bromocresol blue, the isolate CM33 was selected as being the highest β-galactosidase producer and was identified as Lactobacillus fermentum based on its morphological characteristics and 16S rDNA nucleotide sequence. L. fermentum CM33 exhibited a good survival rate under the simulated stomach passage model, comparable to known probiotic strains L. gallinarum JCM2011 and L. agilis JCM1187. L. fermentum CM33 was antagonistic to pathogenic bacteria Listeria monocytogenes, Escherichia coli 0157:H7, Salmonella typhi, and Salmonella enteriditis, using the well diffusion method. In addition, the selected lactobacilli exhibited a high growth rate when cultivated in modified MRS containing commercial galactooligosaccharide (GOS) as a sole carbon source, as well as in glucose. A preliminary study on the enzymatic synthesis of oligosaccharide using crude β-galactosidase revealed the capability for oligosaccharide synthesis by the transgalactosylation activity.     

Control of spoilage fungi by lactic acid bacteria

The evaluation of the potentiality of lactic acid bacteria (LAB) strains isolated from different origins to inhibit mould growth and to identify and characterize the antifungal metabolites were the aims of this study. From a total of ninety-one LAB strains tested, ten were selected due to their high inhibitory effect (>80%). The antifungal activity of the majority of the selected LAB strains was lost after the neutralization treatment determining the acidic nature of the antifungal metabolites. Lactic, acetic and phenyllactic (PLA) acids were identified as being responsible for antifungal effect in the 10 cell-free supernatants (CFS) evaluated. Amongst the strains evaluated, only Lactobacillus fermentum CRL 251 produced fungus inhibitory peptide/s, smaller than 10 kDa, thermostable, active in the pH range of 4–7 and sensitive to trypsin. This is the first report on antifungal peptide/s produced by a L. fermentum strain.     

Probiotic potential of lactic acid bacteria isolated from chicken gastrointestinal digestive tract

This study was conducted in order to evaluate the probiotic properties of lactic acid bacteria (LAB) isolated from intestinal tract of broilers and Thai indigenous chickens. The major properties, including the gastric juice and bile salts tolerance, starch, protein and lipid digesting capabilities, and the inhibition on certain pathogenic bacteria were investigated. Three-hundred and twenty-two and 226 LAB strains were isolated from ten broilers and eight Thai indigenous chickens, respectively. The gastrointestinal transit tolerance of these 548 isolates was determined by exposing washed cell suspension at 41°C to simulated gastric juice (pH 2.5) containing pepsin (3 mg ml⁻¹), and to simulated small intestinal juice (pH 8.0) in the presence of pancreatin (1 mg ml⁻¹) and 7% fresh chicken bile, mimicking the gastrointestinal environment. The survival of 20 isolates was found after passing through the gastrointestinal conditions. The survival rates of six strains; KT3L20, KT2CR5, KT10L22, KT5S19, KT4S13 and PM1L12 from the sequential study were 43.68, 37.56, 33.84, 32.89, 31.37 and 27.19%, respectively. Twelve isolates exhibited protein digestion on agar plate but no isolates showed the ability to digest starch and lipid. All 20 LAB showed the antimicrobial activity against Salmonella sp., Staphylococcus aureus and Escherichia coli except one strain which did not show the inhibitory activity toward E. coli. Accordingly, five isolates of selected LAB (KT2L24, KT3L20, KT4S13, KT3CE27 and KT8S16) can be classified as the best probiotics and were identified as Enterococcus faecalis, Enterococcus durans, Enterococcus faecium, Pediococcus pentosaceus, and Enterococcus faecium, respectively. The survival rate of microencapsulation of E. durans KT3L20 under simulated small intestine juice after sequential of simulated gastric juice was also investigated. An extrusion technique exhibited a higher survival rate than emulsion technique and free cell, respectively.     

Isolation and characterization of probiotic properties of Lactobacilli isolated from rat fecal microbiota

The objective of the present study was to characterize lactobacilli isolates from the feces of male Wistar rats. Various physiological features of the candidate probiotic isolates were preliminarily investigated, including tolerance to simulated gastric juice and bile salts, antimicrobial activity, antibiotic susceptibility and in vitro aggregation. Based on their morphological and biochemical characteristics, four potential probiotic isolates (CS2, CS3, CS4, and CS7) were screened. The isolates showed good tolerance to stimulated gastric juice and bile salts. CS4 and CS7 exhibited strong antibacterial activities against the pathogens tested as assessed in neutral pH culture supernatants. All lactobacilli isolates were susceptible to all the tested antibiotics, except vancomycin. Moreover, the isolate CS4 and CS7 were found to possess stronger cell surface traits such as hydrophobicity, auto‐aggregation and co‐aggregation capacity. In addition, CS4 and CS7 had greater β‐galactosidase activities than the others. Biochemical tests and 16S rRNA gene sequencing confirmed that CS2, CS3, CS4 and CS7 are Lactobacillus intestinalis PJ2, L. sakei PJ3, L. helveticus PJ4, and L. plantarum PJ7, respectively. Based on the obtained results, L. helveticus PJ4 and L. plantarum PJ7 are ideal in vitro probiotic candidates and require further in vivo evaluation.     

Bioconversion of isoflavone glycosides to aglycones,mineral bioavailability and vitamin B complex in fermented soymilk by probiotic bacteria and yeast

Aim: To study the role of β‐glucosidase producing probiotic bacteria and yeast in the biotransformation of isoflavone glycosides to aglycones, mineral bioavailability and vitamin B complex in fermented soymilk. Methods and Results: Five isolates of probiotic lactic acid bacteria (LAB), Lactobacillus acidophilus B4496, Lactobacillus bulgaricus CFR2028, Lactobacillus casei B1922, Lactobacillus plantarum B4495 and Lactobacillus fermentum B4655 with yeast Saccharomyces boulardii were used to ferment soymilk to obtain the bioactive isoflavones, genistein and daidzein. High‐performance liquid chromatography was used to analyse the concentration of isoflavones. Bioactive aglycones genistein and daidzein after 24 and 48 h of fermentation ranged from 97·49 to 98·49% and 62·71 to 92·31% respectively with different combinations of LAB with yeast. Increase in bioavailability of minerals and vitamin B complex were also observed in fermented soymilk. Conclusions: LAB in combination with yeast S. boulardii has great potential for the enrichment of bioactive isoflavones, enhancing the viability of LAB strains, decreasing the antinutrient phytic acid and increasing the mineral bioavailability in soymilk fermentation. Significance and Impact of the Study: Fermentation of soymilk with probiotic organisms improves the bioavailability of isoflavones, assists in digestion of protein, provides more soluble calcium, enhances intestinal health and supports immune system. Increased isoflavone aglycone content in fermented soymilk improves the biological functionality of soymilk.