Metagenetic analysis of the bacterial diversity of Kazakh koumiss and assessment of its anti-Candida albicans activity.

Koumiss, a five-thousand-year-old fermented mare's milk beverage, is widely recognized for its beneficial nutrient and medicinal properties. The microbiota of Chinese and Mongolian koumiss have been largely characterized in recent years, but little is known concerning Kazakh koumiss despite this drink historically originates from the modern Kazakhstan territory. In addition, while koumiss is regarded as a drink with therapeutic potential, there are also no data on koumiss anti-Candida activity. In this context, the aims of the present study were to investigate the bacterial diversity and anti-Candida albicans activity of homemade Kazakh koumiss samples as well as fermented whey and cow's milk, derived from koumiss and propagated for several months. Koumiss bacterial communities were largely dominated by lactic acid bacteria including Lactobacillus sensu lato spp. (69% of total reads), Streptococcus (8.0%) and Lactococcus (6.1%), while other subdominant genera included Acetobacter (2.6%), Enterobacter (2.4%), and Klebsiella (1.5%). Several but not all koumiss samples as well as fermented whey and cow's milk showed antagonistic activities towards C. albicans. Linear discriminant effect size (LEfSe) analysis showed that their bacterial communities were characterized by a significantly higher abundance of amplicon sequence variants (ASV) belonging to the genus Acetobacter. In conclusion, this study allowed to identify the key microorganisms of Kazakh koumiss and provided new information on the possible underestimated contribution of acetic acid bacteria to its probiotic properties.

Lactobacillus paracasei from koumiss ameliorates diarrhea in mice via tight junctions modulation.

OBJECTIVES: Probiotics are gaining interest as alternative options for antibiotic or antiinflammatory drugs. Probiotics can affect the health of the host through metabolites and competitive inhibition adhesion of pathogenic microorganisms. Koumiss is an important part of the diet of Asian nomads, and is rich in a broad array of probiotics that can benefit the body. Mongolians have developed koumiss therapy to assist in the treatment of various diseases. In the present study, we investigate the beneficial effect of Lactobacillus paracasei, a strain isolated from koumiss, on a mouse model of diarrhea induced by Escherichia coli O8 (E. coli O8). METHODS: Probiotics were isolated from Mongolian koumiss. The resistance of probiotics against acid, bile salts, gastric juice, and intestinal juice was evaluated. The mouse model of diarrhea was established by the intragastric administration of E. coli O8 after NaHCO3 treatment. L. paracasei was intragastrically administered before or after E. coli O8 exposure in mice. The plasma levels of diamine oxidase and zounlin were quantified using an enzyme-linked immunosorbent assay, and the integrity of the intestinal barrier and goblet cells of mice with diarrhea were observed using hematoxylin and eosin and Alcian blue periodic acid-Schiff staining. The expression of tight junction (TJ) proteins was detected by immunohistochemistry and Western blot. RESULTS: A total of five lactic acid bacteria and two yeast strains were isolated from koumiss, and L. paracasei was screened for animal experiments. Experimental results showed that L. paracasei could reduce the increase in diamine oxidase and zonulin caused by E. coli (P < 0.05); increase goblet cells and the expression of TJ proteins ZO-1, occludin, and claudin-1 (P < 0.05); increase the expression of mucin 2, oligomeric mucus/gel-forming (P < 0.05) protein; and reduce the level of inhibitor kappa B-alpha and myosin light-chain kinase. CONCLUSIONS: L. paracasei reduced the intestinal permeability, induced the expression of mucin 2, oligomeric mucus/gel-forming protein, and increased the number of goblet cells in mice by the upregulation of the expression of TJ proteins via the nuclear factor kappa B cells-myosin light-chain kinase signaling pathway.

Study on the effect of koumiss on the intestinal microbiota of mice infected with Toxoplasma gondii.

Toxoplasma gondii is a worldwide food-borne parasite that can infect almost all warm-blooded animals, including humans. To date, there are no effective drugs to prevent or eradicate T. gondii infection. Recent studies have shown that probiotics could influence the relationship between the microbiota and parasites in the host. Koumiss has been used to treat many diseases based on its probiotic diversity. Therefore, we explored the effect of koumiss on T. gondii infection via its effect on the host intestinal microbiota. BALB/c mice were infected with T. gondii and treated with PBS, koumiss and mares' milk. Brain cysts were counted, and long-term changes in the microbiota and the effect of koumiss on gut microbiota were investigated with high-throughput sequencing technology. The results suggested that koumiss treatment significantly decreased the cyst counts in the brain (P < 0.05). Moreover, T. gondii infection changed the microbiota composition, and koumiss treatment increased the relative abundance of Lachnospiraceae and Akkermansia muciniphila, which were associated with preventing T. gondii infection. Moreover, koumiss could inhibit or ameliorate T. gondii infection by increasing the abundance of certain bacteria that control unique metabolic pathways. The study not only established a close interaction among the host, intracellular pathogens and intestinal microbiota but also provided a novel focus for drug development to prevent and eradicate T. gondii infection.

Comparative metabolomics and microbial communities associated network analysis of black and white horse- sourced koumiss.

The quality and formation of bioactive components in fermented koumiss are based on the complex metabolism of the microbial community. In the present study, changes in the bioactive metabolites and microbial communities in black and white horse-sourced koumiss were evaluated during the fermentation process. 74 and 69 differential metabolites were formed when BLM and WHM were fermentated into koumiss. Lactobacillus and Dekkera grew rapidly and became the dominant genera in the koumiss. Bioactive compounds (e.g., adenine, d-proline) were significantly enhanced after natural fermentation and were positively correlated with Lactobacillus, Dekkera and Acetobacter. The microbial metabolic network showed that Lactobacillus and Dekkera were the functional core microbiota and played significant roles in the formation of bioactive compounds, followed by Acetobacter, Streptococcus and Leuconostoc. The current study results provide new insight into the formation of bioactive components in koumiss, which is useful for directionally isolating functional microorganisms suitable for koumiss fermentation.

Determination of yeast diversity in fermented Sumbawa mare’s milk using internal transcribed spacers (ITS) fragment analysis

Aims@#Molecular identification of yeast has been conducted on various fermentation products. However, the identification of yeast in fermented Sumbawa mare’s milk based on the genotyping method has not been carried out. This study was aimed to determine the diversity profile of yeasts in fermented Sumbawa mare’s milk using phenetic characters and PCR-RFLP analysis technique based on the ITS region.@*Methodology and results@#Yeast isolates were phenotypically characterized and visualized in a dendrogram using CLAD97 software. Then, the yeast DNA was extracted using heat treatment and amplified using ITS1 and ITS4 primers. The amplicons were analyzed by RFLP using HindIII and HaeIII enzymes. The phylogenetic tree was constructed using MEGA 7.0. Based on the result of grouping by phenetic analysis and PCR-RFLP, the 12 isolates were divided into four groups with different members. The results of the phenetic analysis were divided into group I (all isolates of Dompu), group II (isolate B3, B4, S3), group III (isolate B5) and group IV (isolate S1). The types of yeast that were identified molecularly and represented each group of PCR-RFLP results included in group I were Kluyveromyces marxianus D1A and K. marxianus D1B, group II: K. marxianus D7, group III: Kazachstania humilis D4, while milk from Bima and Sumbawa has one yeast species as a member of group IV, namely Pichia kudriavzevii B3. Kluyveromyces marxianus was the yeast frequently found in Sumbawa fermented mare’s milk.@*Conclusion, significance and impact of study@#Various yeast species as a consortium of the milk samples can contribute to the increasing quality of fermented Sumbawa mare’s milk.

Novel insight into physicochemical and flavor formation in koumiss based on microbial metabolic network.

Koumiss, naturally fermented mare's milk, has a unique flavor that is affected by its resident microflora. In this study, we analyzed time-dependent changes in the microbiota and volatile metabolite profiles of koumiss using high-throughput sequencing and liquid-liquid extraction-gas chromatography-mass spectrometry. The dominant microrganisms in mare milk were Enterobacter and Rhodotorula, which were gradually suppressed, then Lactobacillus and Dekkera became the core microorganisms of koumiss. 17 FAA and 52 volatile organic compounds (VOCs) were identified during koumiss fermentation, including 19 odor active compounds and 15 differential metabolites of VOCs, mainly acids and esters, and their anabolism showed significant positive correlations with Lactobacillus and Dekkera. The microbial metabolic network showed that Lactobacillus and Dekkera were identified as the most important flavor-producing microbiota due to their significantly positive correlation with the formation of 33 and 28 flavor metabolites, respectively. These microorganisms produce enzymes that catalyze a series of metabolic pathways that generate VOCs, including carbohydrate metabolism, amino acid metabolism, and fatty acid biosynthesis. Our findings provide new guidance for the screening and application of flavor-producing microorganisms inkoumiss.

Koumiss promotes Mycobacterium bovis infection by disturbing intestinal flora and inhibiting endoplasmic reticulum stress.

Mycobacterium bovis is the causative agent of bovine tuberculosis and also responsible for serious threat to public health. Koumiss is a fermented mare's milk product, used as traditional drink. Here, we explored the effect of koumiss on gut microbiota and the host immune response against M bovis infection. Therefore, mice were treated with koumiss and fresh mare milk for 14 days before M bovis infection and continue for 5 weeks after infection. The results showed a clear change in the intestinal flora of mice treated with koumiss, and the lungs of mice treated with koumiss showed severe edema, inflammatory infiltration, and pulmonary nodules in M bovis-infected mice. Notably, we found that the content of short-chain fatty acids was significantly lower in the koumiss-treated group compared with the control group. However, the expression of endoplasmic reticulum stress and apoptosis-related proteins in the lungs of koumiss-treated mice were significantly decreased. Collectively, these findings suggest that koumiss treatment disturb the intestinal flora of, which is associated with disease severity and the possible mechanism that induces lungs pathology. Our current findings can be exploited further to establish the "gut-lung" axis which might be a novel strategy for the control of tuberculosis.

Antibacterial spectrum of four compounds from yeasts in koumiss.

Koumiss has beneficial therapeutic effects on bacterial diseases. Four antibacterial com- pounds from yeasts (Kluyveromyces marxianus and Saccharomyces cerevisiae) in koumiss were evaluated for their antibacterial effects against three Gram-negative bacteria, three Gram-positive bacteria and five pathogenic Escherichia coli strains. The antibacterial compounds from yeasts in koumiss were extracted, and their main components were determined. The inhibition zones were analyzed, and their minimum inhibition concentrations (MICs) and minimum bactericidal concentrations (MBCs) were determined. Aqueous phases of Kluyveromyces marxianus and Saccharomyces cerevisiae at pH 2.0 and 8.0 produced larger inhibition zones than those in other phases, and then antibacterial compounds from K. marxianus (K2, pH=2.0; K8, pH=8.0) and S. cerevisiae (S2, pH=2.0; S8, pH=8.0) were obtained. Their main components were organic acids and killer toxins. K2 had more propanoic acid and S2 had more oxalic acid than others. The inhibition zones of K2, K8, S2 and S8 against three Gram-negative bacteria and three Gram-positive bacteria were 12.03-23.30 mm, their MICs were 0.01-0.13 g/mL, and MBCs were 0.03-0.50 g/mL. Meantime, the inhibition zones of K2, K8, S2 and S8 against five pathogenic E. coli were 16.10-25.26 mm, their MICs were 0.03-0.13 g/mL, and MBCs were 0.13-1.00 g/mL. These four antibacterial compounds from yeasts in koumiss had broad antibacterial spectrum. In addition, K2 and S2 were better than K8 and S8.

LuxS-mediated quorum sensing system in Lactobacillus plantarum NMD-17 from koumiss: induction of plantaricin MX in co-cultivation with certain lactic acid bacteria.

A bacteriocin termed plantaricin MX with a broad antimicrobial spectrum was produced by Lactobacillus plantarum NMD-17, which was isolated from Inner Mongolia traditional koumiss of china. Among 300 strains of lactic acid bacteria (LAB) belonging to the genera Lactococcus, Lactobacillus, Streptococcus, Leuconostoc, and Enterococcus, five strains including Lactobacillus reuteri NMD-86, Lactobacillus helveticus NMD-137, Lactococcus lactis NMD-152, Enterococcus faecalis NMD-178, and Enterococcus faecium NMD-219 were revealed to significantly induce the bacteriocin synthesis and greatly increase the cell numbers of Lactobacillus plantarum NMD-17 and activity of AI-2 signaling molecule. Bacteriocin synthesis was not increased by cell-free supernatants and autoclaved cultures of inducing strains, demonstrating that intact cells of inducing strains were essential to the induction of bacteriocin synthesis. The existence of bacteriocin structural plnEF genes and the plnD and luxS genes involved in quorum sensing was confirmed by PCR, and the presence of plnB gene encoding histidine protein kinase was determined by single oligonucleotide nested PCR (Son-PCR). Quantitative real-time PCR demonstrated that plnB, plnD, luxS, plnE, and plnF genes of L. plantarum NMD-17 were upregulated significantly (P < 0.01) in co-cultivation with L. reuteri NMD-86. The results showed that the bacteriocin synthesis of L. plantarum NMD-17 in co-cultivation might have a close relationship with LuxS-mediated quorum sensing system.

Isolation and identification of microorganisms in Kazakhstan koumiss and their application in preparing cow-milk koumiss.

Koumiss is a type of famous fermented mare milk and considered an important nutritious beverage in central Asian countries. However, the production of koumiss cannot meet public demand in the market due to availability of mare milk. In the present study, 52 lactic acid bacteria and 20 yeast strains from traditional homemade Kazakhstan koumiss were isolated and identified. The isolates were used in a trial that included fermented cow milk, and the flavor profiles, color, and taste to determine their contribution in the co-fermentation of cow milk. Based on the sensory evaluation, KZLAB13 and KZY10 strains were selected as the best cofermentation combinations. The optimal fermentation conditions were confirmed as the ratio of the starter culture 2.4:1.6 % (vol/vol) KZLAB13 strain to KZY10 strain and a temperature of 36°C for 16 h using response surface methodology. After evaluating the quality of the optimized cow-milk koumiss compared with the Kazakhstan koumiss, results suggested that cow milk fermented by these 2 strains possessed a promising taste, flavor, and physicochemical and rheological properties. Altogether, our results showed that cow milk fermented with a combination of KZLAB13 and KZY10 strains can simulate the taste, flavor, and quality of traditional koumiss. Our study provided a novel alternative to mare-milk koumiss and could be used in dairy programs to fulfill the needs of people.

Profiling of koumiss microbiota and organic acids and their effects on koumiss taste.

BACKGROUND: Koumiss is a naturally fermented mare's milk. Over recent decades, numerous studies have revealed the diversity of lactic acid bacteria in koumiss. However, there is limited information available regarding its secondary major component yeast profile. RESULTS: A total of 119 bacterial and 36 yeast species were identified among the 14 koumiss samples. The dominant bacterial species in koumiss were Lactobacillus helveticus, Lactobacillus kefiranofaciens, Lactococcus lactis, Lactococcus raffinolactis, and Citrobacter freundii. The main yeast species were Dekkera anomala, Kazachstania unispora, Meyerozyma caribbica, Pichia sp.BZ159, Kluyveromyces marxianus, and uncultured Guehomyces. The bacterial and yeast Shannon diversity of the Xilinhaote-urban group were higher than those of the Xilingol-rural group. The most dominant organic acids were lactic, acetic, tartaric, and malic acids. Lactic acid bacteria species were mostly responsible for the accumulation of those organic acids, although Kazachstania unispora, Dekkera anomala, and Meyerozyma caribbica may also have contributed. Redundancy analysis suggested that both bacteria and yeast respond to koumiss flavor, such as Lactobacillus helveticus and Dekkera anomala are associated with sourness, astringency, bitterness, and aftertaste, whereas Lactococcus lactis and Kazachstania unispora are associated with umami. CONCLUSIONS: Our results suggest that differences were observed in koumiss microbiota of Xilinhaote-urban and Xilingol-rural samples. The biodiversity of the former was higher than the latter group. Positive or negative correlations between bacteria and yeast species and taste also were found.

A UPLC-Q-TOF-MS-based metabolomics approach for the evaluation of fermented mare's milk to koumiss.

The fermentation of mare's milk into koumiss produces many beneficial functional compounds depending on the metabolism of the initial microbial flora. In this study, metabolites found in mare's milk and resulting koumiss were identified. Major metabolic pathways in the fermentation were also identified using an UPLC-Q-TOF-MS-based metabolomics method. In total, 354 metabolites were identified: 61 were up-regulated and 105 were down-regulated. Metabolic pathway analyses revealed that c-5-branched dibasic acid metabolism, valine, leucine and isoleucine degradation, arginine and proline metabolism, valine, leucine and isoleucine biosynthesis, vascular smooth muscle contraction, aminoacyl-tRNA biosynthesis and ß-alanine metabolism showed significant increases. A hierarchical cluster analysis of metabolites indicated a clear grouping pattern in which the relative concentrations of p-pyruvate, 20-HETE, 4-aminobutanoate, uracil, acetoacetate, and γ-linolenic acid differed significantly between milk and koumiss. This study provides reference values for metabolic isolates and bioactive compounds purification in mare's milk and koumiss.

Comparison of backslopping and two-stage fermentation methods for koumiss powder production based on chemical composition and nutritional properties.

BACKGROUND: Koumiss is a traditional fermented beverage made from mare's milk. The traditional backslopping method for koumiss production has shortcomings in terms of microbiological diversity and nutritional characteristics. In this study, a two-stage fermentation method was established to scale up the production of koumiss powder. The chemical composition and nutritional properties of a novel two-stage fermentation koumiss powder (TKP) were compared with backslopping koumiss powder (BKP). RESULTS: The TKP exhibited important nutritional and functional properties, including a high percentage of essential amino acids, and high polyunsaturated fatty acid, vitamin, and mineral content. The essential amino acid content of TKP was not significantly different from that of BKP. The oleic acid, linoleic acid, linolenic acid, and water-soluble vitamin content of TKP was higher than that of BKP. The Ca:P ratio of TKP was also close to the optimal Ca:P ratio in humans. CONCLUSION: The novel method could be applied for the scaled-up production of koumiss powder with similar nutritional properties to traditional backslopping koumiss powder. The successful production of koumiss powder could also promote the development of the koumiss industry. © 2019 Society of Chemical Industry.

Antibacterial and antioxidant activity of exopolysaccharide mediated silver nanoparticle synthesized by Lactobacillus brevis isolated from Chinese koumiss.

Recently, silver nanoparticles gain significant attention due to their applications in various fields. The aim of present study was to develop the eco-friendly, cost effective, and simple method to biosynthesized the silver nanoparticle using sliver nitrate as precursor. In this study, we investigated the physical characterization and biotechnological applications of biosynthesized silver nanoparticle using exopolysaccharide of probiotic Lactobacillus brevis MSR104 isolated from Chinese koumiss. Biosynthesized silver nanoparticles were characterized using the fourier-transform infrared spectroscopy, scanning electron microscopy, transmission electron microscopy, X-ray diffraction analysis, and elemental analyzer. The achieved results indicate that silver nanoparticles varied in sized with an average size of 45 nm. The X-ray diffraction analysis results showed that the silver nanoparticles have a crystalline nature. The results of antimicrobial assay indicated that the silver nanoparticles exhibited outstanding antimicrobial activity in dose dependent manner against both Gram's negative as well as Gram's positive. The antioxidant results indicate that the silver nanoparticles showed excellent scavenging rate against DPPH free radicals (81.4 ±â€¯1.2%) and nitric oxide free radicals (75.06 ±â€¯0.4%). Furthermore, the results of MTT assay revealed that the AgNPs significantly reduced the percentage of live HT-29 cells at higher concentration. This study concluded that the newly synthesized silver nanoparticles have antibacterial, antioxidant, and anticancer applications in agricultural and food industries.

Investigating the bacterial microbiota of traditional fermented dairy products using propidium monoazide with single-molecule real-time sequencing.

Traditional fermented dairy foods have been the major components of the Mongolian diet for millennia. In this study, we used propidium monoazide (PMA; binds to DNA of nonviable cells so that only viable cells are enumerated) and single-molecule real-time sequencing (SMRT) technology to investigate the total and viable bacterial compositions of 19 traditional fermented dairy foods, including koumiss from Inner Mongolia (KIM), koumiss from Mongolia (KM), and fermented cow milk from Mongolia (CM); sample groups treated with PMA were designated PKIM, PKM, and PCM. Full-length 16S rRNA sequencing identified 195 bacterial species in 121 genera and 13 phyla in PMA-treated and untreated samples. The PMA-treated and untreated samples differed significantly in their bacterial community composition and α-diversity values. The predominant species in KM, KIM, and CM were Lactobacillus helveticus, Streptococcus parauberis, and Lactobacillus delbrueckii, whereas the predominant species in PKM, PKIM, and PCM were Enterobacter xiangfangensis, Lactobacillus helveticus, and E. xiangfangensis, respectively. Weighted and unweighted principal coordinate analyses showed a clear clustering pattern with good separation and only minor overlapping. In addition, a pure culture method was performed to obtain lactic acid bacteria resources in dairy samples according to the results of SMRT sequencing. A total of 102 LAB strains were identified and Lb. helveticus (68.63%) was the most abundant, in agreement with SMRT sequencing results. Our results revealed that the bacterial communities of traditional dairy foods are complex and vary by type of fermented dairy product. The PMA treatment induced significant changes in bacterial community structure.

Characterization of a broad spectrum bacteriocin produced by Lactobacillus plantarum MXG-68 from Inner Mongolia traditional fermented koumiss.

An agar well diffusion assay (AWDA) was used to isolate a high bacteriocin-producing strain with a broad spectrum of antibacterial activity, strain MXG-68, from Inner Mongolia traditional fermented koumiss. Lactobacillus plantarum MXG-68 was identified by morphological, biochemical, and physiological characteristics and 16S rDNA analysis. The production of antibacterial substance followed a growth-interrelated model, starting at the late lag phase of 4 h and arriving at a maximum value in the middle of the stationary phase at 24 h. Antibacterial activity was abolished or decreased in the presence of pepsin, chymotrypsin, trypsin, proteinase, and papain K. The results showed that antibacterial substances produced by L. plantarum MXG-68 were proteinaceous and could thus be classified as the bacteriocin, named plantaricin MXG-68. The molar mass of plantaricin MXG-68 was estimated to be 6.5 kDa, and the amino acid sequence of its N-terminal was determined to be VYGPAGIFNT. The mode of plantaricin MXG-68 action was determined to be bactericidal. Bacteriocin in cell-free supernatant (CFS) at pH 7 was stable at different temperatures (60 °C, 80 °C, 100 °C, 121 °C for 30 min; 4 °C and - 20 °C for 30 days), as well as at pH 2.0-10.0. Antibacterial activity maintained stable after treatment with organic solvents, surfactants, and detergents but increased in response to EDTA. Response surface methodology (RSM) revealed the optimum conditions of bacteriocin production in L. plantarum MXG-68, and the bacteriocin production in medium optimized by RSM was 26.10% higher than that in the basal MRS medium.

Study of bacterial and fungal community structures in traditional koumiss from Inner Mongolia.

Koumiss is notable for its nutritional functions, and microorganisms in koumiss determine its versatility. In this study, the bacterial and fungal community structures in traditional koumiss from Inner Mongolia, China, were investigated. Our results demonstrated that 6 bacterial phyla represented by 126 genera and 49 species and 3 fungal phyla represented by 59 genera and 57 species were detected in 11 samples of artisanal koumiss. Among them, Lactobacillus was the predominant genus of bacterium, and Kluyveromyces and Saccharomyces dominated at the fungal genus level. In addition, there were no differences in the bacterial and fungal richness and diversity of koumiss from 3 neighboring administrative divisions in Inner Mongolia, and the bacterial and fungal community structures (the varieties and relative abundance of bacterial and fungal genera and species) were clearly distinct in individual samples. This study provides a comprehensive understanding of the bacterial and fungal population profiles and the predominant genus and species, which would be beneficial for screening, isolation, and culture of potential probiotics to simulate traditional fermentation of koumiss for industrial and standardized production in the future.

Characterization of a lactose-responsive promoter of ATP-binding cassette (ABC) transporter gene from Lactobacillus acidophilus 05-172.

A novel lactose-responsive promoter of the ATP-binding cassette (ABC) transporter gene Lba1680 of Lactobacillus acidophilus strain 05-172 isolated from a traditionally fermented dairy product koumiss was characterized. In L. acidophilus 05-172, expression of Lba1680 was induced by lactose, with lactose-induced transcription of Lba1680 being 6.1-fold higher than that induced by glucose. This is in contrast to L. acidophilus NCFM, a strain isolated from human feces, in which expression of Lba1680 and Lba1679 is induced by glucose. Both gene expression and enzyme activity assays in L. paracasei transformed with a vector containing the inducible Lba1680 promoter (PLba1680) of strain 05-172 and a heme-dependent catalase gene as reporter confirmed that PLba1680 is specifically induced by lactose. Its regulatory expression could not be repressed by glucose, and was independent of cAMP receptor protein. This lactose-responsive promoter might be used in the expression of functional genes in L. paracasei incorporated into a lactose-rich environment, such as dairy products.

Investigating bacterial population structure and dynamics in traditional koumiss from Inner Mongolia using single molecule real-time sequencing.

Koumiss is considered as a complete dairy product high in nutrients and with medicinal properties. The bacterial communities involved in production of koumiss play a crucial role in the fermentation cycle. To reveal bacterial biodiversity in koumiss and the dynamics of succession in bacterial populations during fermentation, 22 samples were collected from 5 sampling sites and the full length of the 16S ribosomal RNA genes sequenced using single molecule real-time sequencing technology. One hundred forty-eight species were identified from 82 bacterial genera and 8 phyla. These results suggested that the structural difference in the bacterial community could be attributed to geographical location. The most significant difference in bacterial composition occurred in samples from group D compared with other groups. The sampling location of group D was distant from the city and maintained the primitive local nomadic life. The dynamics of succession in bacterial communities showed that Lactobacillus helveticus increased in abundance from 0 to 9h and reached its peak at 9h and then decreased. In contrast, Enterococcus faecalis, Enterococcus durans, and Enterococcus casseliflavus increased gradually throughout the fermentation process, and reached a maximum after 24h.

Purification and characterization of a novel bacteriocin produced by Lactobacillus crustorum MN047 isolated from koumiss from Xinjiang, China.

The growing emergence of antibiotic-resistant bacteria in the food industry needs to be controlled with effective antimicrobials. In this study, bacteriocin MN047 A (BMA) was found to have antibacterial activity against multidrug-resistant bacteria. It was produced by Lactobacillus crustorum MN047, which was first isolated from koumiss, a traditional fermented dairy product from Xinjiang Autonomous Region, China. It was purified by ammonium sulfate precipitation, ion-exchange chromatography, and reversed-phase chromatography. It had a low molecular mass of 1,770.89 Da according to matrix-assisted laser desorption/ionization time-of-flight mass spectrometry, and the sequence was identified as QLPWQILGIVAGMFQA by liquid chromatography-tandem mass spectrometry analysis and MASCOT searching. It was proteinaceous in nature: the bacteriocin was digested by protease but not by α-amylase or lipase. It showed broad pH toleration (pH 2-11), good thermostability, and good storage stability. It had a broad inhibitory spectrum, including both gram-positive and gram-negative bacteria. Growth curve and time-kill kinetics indicated that it was bactericidal to the indicator strains, and this finding was verified by scanning electron microscope and transmission electron microscope after treatment with BMA. As well, BMA halted the growth of Staphylococcus aureus and Escherichia coli in the G1 and G2/M phases according to cell-cycle analysis by flow cytometry, indicating that BMA had comprehensive inhibitory effects against foodborne pathogens.