Bilige Menghe

A phylo-functional core of gut microbiota in healthy young Chinese cohorts across lifestyles, geography and ethnicities

Structural profiling of healthy human gut microbiota across heterogeneous populations is necessary for benchmarking and characterizing the potential ecosystem services provided by particular gut symbionts for maintaining the health of their hosts. Here we performed a large structural survey of fecal microbiota in 314 healthy young adults, covering 20 rural and urban cohorts from 7 ethnic groups living in 9 provinces throughout China. Canonical analysis of unweighted UniFrac principal coordinates clustered the subjects mainly by their ethnicities/geography and less so by lifestyles. Nine predominant genera, all of which are known to contain short-chain fatty acid producers, co-occurred in all individuals and collectively represented nearly half of the total sequences. Interestingly, species-level compositional profiles within these nine genera still discriminated the subjects according to their ethnicities/geography and lifestyles. Therefore, a phylogenetically diverse core of gut microbiota at the genus level may be commonly shared by distinctive healthy populations as functionally indispensable ecosystem service providers for the hosts.

Isolation and preliminary probiotic selection of lactobacilli from koumiss in Inner Mongolia.

From 16 samples of traditional fermented koumiss collected in Inner Mongolia Autonomous Region of China, forty‐eight lactobacilli strains were isolated and phenotypically characterized by their abilities to ferment different carbohydrates and by additional biochemical tests. The dominant lactobacilli species were identified as L. casei (17 strains), L. helveticus (10 strains) and L. plantarum (8 strains), with a lower frequency of isolation for L. coryniformis subsp. coryniformis (5 strains), L. paracasei (3 strains), L. kefiranofaciens (2 strains), L. curvatus (1 strain), L. fermentum (1 strain) and W. kandleri (1 strain). The pH values of all these samples were ranging from 3.37 to 3.94. In isolates, L. casei Zhang, L. helveticus ZL12‐1, and L. plantarum BX6‐6 were selected as potentially probiotic strains through the preliminary tests including resistance to low acid, abilities to grow in MRS with bile salts, antimicrobial activities and the viabilities during prolonged cold storage in fermented milk. Moreover 16S rDNA was conducted to confirm the identification. (© 2009 WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim)

Diversity of lactic acid bacteria associated with traditional fermented dairy products in Mongolia

Spontaneous milk fermentation has a long history in Mongolia, and beneficial microorganisms have been handed down from one generation to the next for use in fermented dairy products. The objective of this study was to investigate the diversity of lactic acid bacteria (LAB) communities in fermented yak, mare, goat, and cow milk products by analyzing 189 samples collected from 13 different regions in Mongolia. The LAB counts in these samples varied from 3.41 to 9.03 log cfu/mL. Fermented yak and mare milks had almost identical mean numbers of LAB, which were significantly higher than those in fermented goat milk but slightly lower than those in fermented cow milk. In total, 668 isolates were obtained from these samples using de Man, Rogosa, and Sharpe agar and M17 agar. Each isolate was considered to be presumptive LAB based on gram-positive and catalase-negative properties, and was identified at the species level by 16S rRNA gene sequencing, multiplex PCR assay, and restriction fragment length polymorphism analysis. All isolates from Mongolian dairy products were accurately identified as Enterococcus faecalis (1 strain), Enterococcus durans (3 strains), Lactobacillus brevis (3 strains), Lactobacillus buchneri (2 strains), Lactobacillus casei (16 strains), Lactobacillus delbrueckii ssp. bulgaricus (142 strains), Lactobacillus diolivorans (17 strains), Lactobacillus fermentum (42 strains), Lactobacillus helveticus (183 strains), Lactobacillus kefiri (6 strains), Lactobacillus plantarum ssp. plantarum (7 strains), Lactococcus lactis ssp. lactis (7 strains), Leuconostoc lactis (22 strains), Leuconostoc mesenteroides (21 strains), Streptococcus thermophilus (195 strains), and Weissella cibaria (1 strain). The predominant LAB were Strep. thermophilus and Lb. helveticus, which were isolated from all sampling sites. The results demonstrate that traditional fermented dairy products from different regions of Mongolia have complex compositions of LAB species. Such diversity of LAB provides useful information for further studies of probiotic strain selection and starter culture design, with regard to the industrial production of traditional fermented milk.

Comparative Genomic Analysis of 45 Type Strains of the Genus Bifidobacterium: A Snapshot of Its Genetic Diversity and Evolution

Bifidobacteria are well known for their human health-promoting effects and are therefore widely applied in the food industry. Members of the Bifidobacterium genus were first identified from the human gastrointestinal tract and were then found to be widely distributed across various ecological niches. Although the genetic diversity of Bifidobacterium has been determined based on several marker genes or a few genomes, the global diversity and evolution scenario for the entire genus remain unresolved. The present study comparatively analyzed the genomes of 45 type strains. We built a robust genealogy for Bifidobacterium based on 402 core genes and defined its root according to the phylogeny of the tree of bacteria. Our results support that all human isolates are of younger lineages, and although species isolated from bees dominate the more ancient lineages, the bee was not necessarily the original host for bifidobacteria. Moreover, the species isolated from different hosts are enriched with specific gene sets, suggesting host-specific adaptation. Notably, bee-specific genes are strongly associated with respiratory metabolism and are potential in helping those bacteria adapt to the oxygen-rich gut environment in bees. This study provides a snapshot of the genetic diversity and evolution of Bifidobacterium, paving the way for future studies on the taxonomy and functional genomics of the genus.

Identification and characterization of the dominant lactic acid bacteria isolated from traditional fermented milk in Mongolia

Five samples of Airag and 20 of Tarag (both in Mongolia) were collected from scattered households. One hundred strains of lactic acid bacteria (LAB) were isolated and identified from these samples according to phenotypic characterization and 16S rRNA gene sequence analysis. Eighty-five isolates belonged to the genus Lactobacillus, 15 being classified as coccoid LAB. All isolates belonged to 5 genera and 11 to different species and subspecies. Lactobacillus (Lb.) helveticus was predominant population in Airag samples, Lb. fermentum and Lb. helveticus were the major LAB microflora in Tarag.

Isolation and identification of dominant microorganisms involved in naturally fermented goat milk in Haixi region of Qinghai, China

Based upon morphological, physiological and biochemical analysis, 33 strains of lactic acid bacteria were isolated and identified from 11 samples of traditional natural fermented goat milk collected from different individual households in Haixi region of Qinghai. The counts of lactic acid bacteria were 2.5×108}3.0×109 CFU/mL, while yeasts were 2.6×106}1.6×108 CFU/mL. Strains were identified as follows: 7 strains ofLactobacillus delbrueckii susp.lactis, 4 strains ofLactobacillus delbrueckii subsp.bulgaricus, 3 strains ofLactobacillus acidophilus, 1 strain ofLactobacillus coryniformis subsp.coryniformis, 12 strains ofStreptococcus thermophilus, 1 strain ofLeuconostoc mesenteroides subsp.mesenteroides, 3 strains ofPediococcus acidilactici, and 2 strains ofAerococcus urinaeequi comb. nov.

Multilocus sequence typing of Lactococcus lactis from naturally fermented milk foods in ethnic minority areas of China

To determine the genetic diversity and phylogenetic relationships among Lactococcus lactis isolates, 197 strains isolated from naturally homemade yogurt in 9 ethnic minority areas of 6 provinces of China were subjected to multilocus sequence typing (MLST). The MLST analysis was performed using internal fragment sequences of 12 housekeeping genes (carB, clpX, dnaA, groEL, murC, murE, pepN, pepX, pyrG, recA, rpoB, and pheS). Six (dnaA) to 8 (murC) different alleles were detected for these genes, which ranged from 33.62 (clpX) to 41.95% (recA) GC (guanine-cytosine) content. The nucleotide diversity (π) ranged from 0.00362 (murE) to 0.08439 (carB). Despite this limited allelic diversity, the allele combinations of each strain revealed 72 different sequence types, which denoted significant genotypic diversity. The dN/dS ratios (where dS is the number of synonymous substitutions per synonymous site, and dN is the number of nonsynonymous substitutions per nonsynonymous site) were lower than 1, suggesting potential negative selection for these genes. The standardized index of association of the alleles IA(S)=0.3038 supported the clonality of Lc. lactis, but the presence of network structure revealed by the split decomposition analysis of the concatenated sequence was strong evidence for intraspecies recombination. Therefore, this suggests that recombination contributed to the evolution of Lc. lactis. A minimum spanning tree analysis of the 197 isolates identified 14 clonal complexes and 23 singletons. Phylogenetic trees were constructed based on the sequence types, using the minimum evolution algorithm, and on the concatenated sequence (6,192 bp), using the unweighted pair-group method with arithmetic mean, and these trees indicated that the evolution of our Lc. lactis population was correlated with geographic origin. Taken together, our results demonstrated that MLST could provide a better understanding of Lc. lactis genome evolution, as well as useful information for future studies on global Lc. lactis structure and genetic evolution, which will lay the foundation for screening Lc. lactis as starter cultures in fermented dairy products.

Molecular identification and quantification of lactic acid bacteria in traditional fermented dairy foods of Russia

Russian traditional fermented dairy foods have been consumed for thousands of years. However, little research has focused on exploiting lactic acid bacteria (LAB) resources and analyzing the LAB composition of Russian traditional fermented dairy foods. In the present study, we cultured LAB isolated from fermented mare and cow milks, sour cream, and cheese collected from Kalmykiya, Buryats, and Tuva regions of Russia. Seven lactobacillus species and the Bifidobacterium genus were quantified by quantitative PCR. The LAB counts in these samples ranged from 3.18 to 9.77 log cfu/mL (or per gram). In total, 599 LAB strains were obtained from these samples using de Man, Rogosa, and Sharpe agar and M17 agar. The identified LAB belonged to 7 genera and 30 species by 16S rRNA and murE gene sequencing and multiplex PCR assay. The predominant LAB isolates were Lactobacillus helveticus (176 strains) and Lactobacillus plantarum (63 strains), which represented 39.9% of all isolates. The quantitative PCR results revealed that counts of 7 lactobacilli species and Bifidobacterium spp. of 30 fermented cow milk samples ranged from 1.19±0.34 (Lactobacillus helveticus in Tuva) to 8.09±0.71 (Lactobacillus acidophilus in Kalmykiya) log cfu/mL of fermented cow milk (mean ± standard error). The numbers of Bifidobacterium spp., Lb. plantarum, Lb. helveticus, and Lb. acidophilus revealed no significant difference between the 3 regions; nevertheless, Lactobacillus paracasei, Lactobacillus fermentum, Lactobacillus sakei, and Lactobacillus delbrueckii ssp. bulgaricus exhibited different degrees of variation across 3 regions. The results demonstrate that traditional fermented dairy products from different regions of Russia have complex compositions of LAB species. The diversity of LAB might be related to the type of fermented dairy product, geographical origin, and manufacturing process.

Characterization of angiotensin-converting enzyme inhibitory activity of fermented milk produced by Lactobacillus helveticus

Hypertension affects up to 30% of the adult population in most countries. It is a known risk factor for cardiovascular diseases, including coronary heart disease, peripheral artery disease, and stroke. Owing to the increased health awareness of consumers, the application of angiotensin-converting enzyme (ACE)-inhibitory peptides produced by Lactobacillushelveticus to prevent or control high blood pressure has drawn wide attention. A total of 59 L. helveticus strains were isolated from traditional fermented dairy products and the ACE-inhibitory activity of the fermented milks produced with the isolated microorganisms was assayed. The ACE-inhibitory activity of 38 L. helveticus strains was more than 50%, and 3 strains (IMAU80872, IMAU80852, and IMAU80851) expressing the highest ACE-inhibitory activity were selected for further studies. Particularly, the gastrointestinal protease tolerance and thermostability of the ACE-inhibitory activity in the fermented milks were assessed. Based on these 2 criteria, IMAU80872 was found to be superior over the other 2 strains. Furthermore, IMAU80872 exhibited a high in vitro ACE-inhibitory activity at the following fermentation conditions: fermentation temperature at 40°C, inoculation concentration of 1×10(6) cfu/mL, and fermentation for 18h. Finally, by using ultra-performance liquid chromatography coupled with electrospray ionization quadrupole time-of-flight tandem mass spectrometry analysis, we observed changes of the metabolome along the milk fermentation process of IMAU80872. Furthermore, 6 peptides were identified, which might have ACE-inhibitory activity. In conclusion, we identified a novel ACE-inhibitory L. helveticus strain suitable for the production of fermented milk or other functional dairy products.

Multilocus sequence typing of Streptococcus thermophilus from naturally fermented dairy foods in China and Mongolia

BackgroundStreptococcus thermophilus is a major dairy starter used for manufacturing of dairy products. In the present study, we developed a multilocus sequence typing (MLST) scheme for this important food bacterium. Sequences of 10 housekeeping genes (carB, clpX, dnaA, murC, murE, pepN, pepX, pyrG, recA, and rpoB) were obtained for 239 S. thermophilus strains, which were isolated from home-made fermented dairy foods in 18 different regions of Mongolia and China.MethodsAll 10 genes of S. thermophilus were sequenced, aligned, and defined sequence types (STs) using the BioNumerics Software. The nucleotide diversity was calculated by START v2.0. The population structure, phylogenetic relationships and the role of recombination were inferred using ClonalFrame v1.2, SplitsTree 4.0 and Structure v2.3.ResultsThe 239 S. thermophilus isolates and 18 reference strains could be assigned into 119 different STs, which could be further separated into 16 clonal complexes (CCs) and 38 singletons. Among the 10 loci, a total of 132 polymorphic sites were detected. The standardized index of association (IAS = 0.0916), split-decomposition and ρ/θ (relative frequency of occurrence of recombination and mutation) and r/m value (relative impact of recombination and mutation in the diversification) confirms that recombination may have occurred, but it occurred at a low frequency in these 10 loci. Phylogenetic trees indicated that there were five lineages in the S. thermophilus isolates used in our study. MSTree and ClonalFrame tree analyses suggest that the evolution of S. thermophilus isolates have little relationship with geographic locality, but revealed no association with the types of fermented dairy product. Phylogenetic analysis of 36 whole genome strains (18 S. thermophilus, 2 S. vestibularis and 16 S. salivarius strains) indicated that our MLST scheme could clearly separate three closely related species within the salivarius group and is suitable for analyzing the population structure of the other two species in the salivarius group.ConclusionsOur newly developed MLST scheme improved the understanding on the genetic diversity and population structure of the S. thermophilus, as well as provided useful information for further studies on the genotyping and evolutionary research for S. thermophilus strains with global diversity.