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Artigo em Inglês | MEDLINE | ID: mdl-34499596


A Gram-stain-negative, coccus-shaped, obligately anaerobic, non-motile and non-spore-forming bacterium, designated strain JN500902T, was isolated from the mud in a fermentation cellar used continuously over 30 years for Chinese strong-flavour baijiu production. Colonies were white, circular, convex and smooth-edged. Growth was observed at 20-40 °C (optimum, 37 °C), at pH 5.0-10 (optimum, pH 7.5), with 0-2 % (w/v) NaCl and with 0-4 % (v/v) ethanol. The Biolog assay demonstrated positive reactions of strain JN500902T in the metabolism of l-fucose and pyruvate. The predominant cellular fatty acids (>10 %) consisted of C16 : 0 and C14 : 0. The major end metabolites of strain JN500902T were acetic acid and ethanol when incubated anaerobically in liquid reinforced clostridial medium. Acetate was the major organic acid end product. The complete genome size of strain JN500902T was 3 420 321 bp with 3327 identified genes. The G+C content was 43.5 mol%. Phylogenetic analysis based on 16S rRNA gene sequences affiliated strain JN500902T with the family Lachnospiraceae, having low sequence similarity (92.8 %) to the nearest type strain, Syntrophococcus sucromutans DSM 3224T and forming a clearly distinct branch. Core genome phylogenetic analysis of the isolate and 134 strains belonging to the family Lachnospiraceae also revealed that strain JN500902T was well-separated from other genera of this family as a monophyletic clade. The average nucleotide identity and amino acid identity values between strain JN500902T and 134 Lachnospiraceae strains were less than 74 and 65 %, respectively. Considering its polyphasic characteristics, strain JN500902T represents a novel genus and species within the family Lachnospiraceae, for which the name Novisyntrophococcus fermenticellae gen. nov., sp. nov. is proposed. The type strain is JN500902T (=CICC 24502T=JCM 33939T).

Clostridiales/classificação , Fermentação , Filogenia , Microbiologia do Solo , Anaerobiose , Técnicas de Tipagem Bacteriana , Composição de Bases , China , Clostridiales/isolamento & purificação , DNA Bacteriano/genética , Ácidos Graxos/química , RNA Ribossômico 16S/genética , Análise de Sequência de DNA
Appl Environ Microbiol ; 87(17): e0088521, 2021 08 11.
Artigo em Inglês | MEDLINE | ID: mdl-34160281


The mud cellar creates a unique microenvironment for the fermentation of strong-flavor baijiu (SFB). Recent research and long-term practice have highlighted the key roles of microbes inhabiting pit mud in the formation of SFB's characteristic flavor. A positive correlation between the quality of SFB and cellar age was extracted from practice; however, the evolutionary patterns of pit mud microbiome and driving factors remain unclear. Here, based on the variation regularity analysis of microbial community structure and metabolites of samples from cellars of different ages (∼30/100/300 years), we further investigated the effects of lactate and acetate (main microbial metabolites in fermented grains) on modulating the pit mud microbiome. Esters (50.3% to 64.5%) dominated the volatile compounds identified in pit mud, and contents of the four typical acids (lactate, hexanoate, acetate, and butyrate) increased with cellar age. Bacteria (9.5 to 10.4 log10 [lg] copies/g) and archaea (8.3 to 9.1 lg copies/g) mainly constituted pit mud microbiota, respectively dominated by Clostridia (39.7% to 81.2%) and Methanomicrobia (32.8% to 92.9%). An upward trend with cellar age characterized the relative and absolute abundance of the most predominant bacterial and archaeal genera, Caproiciproducens and Methanosarcina. Correlation analysis revealed significantly (P < 0.05) positive relationships between the two genera and major metabolites. Anaerobic fermentation with acetate and lactate as carbon sources enhanced the enrichment of Clostridia, and furthermore, the relative abundance of Caproiciproducens (40.9%) significantly increased after 15-day fed-batch fermentation with lactate compared with the initial pit mud (0.22%). This work presents a directional evolutionary pattern of pit mud microbial consortia and provides an alternative way to accelerate the enrichment of functional microbes. IMPORTANCE The solid-state anaerobic fermentation in a mud cellar is the most typical feature of strong-flavor baijiu (SFB). Metabolites produced by microbes inhabiting pit mud are crucial to create the unique flavor of SFB. Accordingly, craftspeople have always highlighted the importance of the pit mud microbiome and concluded by centuries of practice that the production rate of high-quality baijiu increases with cellar age. To deepen the understanding of the pit mud microbiome, we determined the microbial community and metabolites of different-aged pit mud, inferred the main functional groups, and explored the forces driving the microbial community evolution through metagenomic, metabolomic, and multivariate statistical analyses. The results showed that the microbial consortia of pit mud presented a regular and directional evolutionary pattern under the impact of continuous batch-to-batch brewing activities. This work provides insight into the key roles of the pit mud microbiome in SFB production and supports the production optimization of high-quality pit mud.

Archaea/isolamento & purificação , Bactérias/isolamento & purificação , Argila/microbiologia , Aromatizantes/metabolismo , Microbiota , Vinho/análise , Archaea/classificação , Archaea/genética , Archaea/metabolismo , Bactérias/classificação , Bactérias/genética , Bactérias/metabolismo , China , Fermentação , Aromatizantes/análise , Armazenamento de Alimentos/instrumentação , Vinho/microbiologia
Genome Announc ; 2(5)2014 Oct 23.
Artigo em Inglês | MEDLINE | ID: mdl-25342677


The complex microbiota of pit mud of solid-state fermentation reactors used for the production of Chinese liquor is responsible for producing one of the oldest distillates in the world. We apply a deep-sequencing approach to characterize the microbiota from pits that have been in use for up to 440 years.

Ying Yong Sheng Tai Xue Bao ; 23(6): 1620-8, 2012 Jun.
Artigo em Chinês | MEDLINE | ID: mdl-22937652


Taking the nine common microbial strains in liquor-making process as test objects, this paper studied the characteristics of phospholipid fatty acid (PLFA), a characteristic component of the strains cell membrane, and the relationships between the detected amount of PLFA and the biomass of the strains. There existed significant differences in the PLFA fingerprints between test bacteria, actinomycetes, molds, and yeasts, and the PLFA fingerprint of each strain could be used as the basis to distinguish species and genus. Within a certain range of the strains biomass, the detected amount of total PLFA or 16:0 was linearly correlated with the biomass. After adding different biomass Gram positive (G+) bacteria, Gram negative (G-) bacteria, and fungi in fermented grains, a significant difference was observed in the relative amount of PLFA between experimental and control samples. It was suggested that the fingerprint of PLFA could quantitatively or semi-quantitatively characterize the microbial community structure and its dynamic variation in fermented grains. By detecting the PLFA profiles of fermented grains in various liquor industries and by analyzing the microbial community structure in the fermented grains, it was substantiated that PLFA fingerprinting was of general applicability.

Bactérias/crescimento & desenvolvimento , Ácidos Graxos/análise , Fermentação , Fungos/crescimento & desenvolvimento , Fosfolipídeos/análise , Actinomyces/crescimento & desenvolvimento , Actinomyces/isolamento & purificação , Bebidas Alcoólicas , Bactérias/isolamento & purificação , Clostridium acetobutylicum/crescimento & desenvolvimento , Clostridium acetobutylicum/isolamento & purificação , Fungos/isolamento & purificação , Lactobacillus plantarum/crescimento & desenvolvimento , Lactobacillus plantarum/isolamento & purificação , Lipídeos de Membrana/análise , Oryza , Dinâmica Populacional
Ying Yong Sheng Tai Xue Bao ; 22(4): 1020-6, 2011 Apr.
Artigo em Chinês | MEDLINE | ID: mdl-21774327


Fermentation pit is a kind of solid bioreactors with unique feature for brewing liquor, especially for Luzhou-flavor, which has significant effects on the quality of produced liquor. There exists a close and complicated correlation between pit age (using time) and microbial community. Taking the characteristic component phospholipid fatty acid (PLFA) in microbial cell membrane as an index, this paper studied the characteristics of the microbial community structure in the pit mud, fermented grains, and yellow water of different age (5-year, 100-year, and 300-year) fermentation pits. The results showed that the total PLFA content was the highest in pit mud, followed by in fermented grains, and in yellow water. The composition of PLFA differed with pit age, and the total PLFA content in yellow water decreased with increasing pit age. In pit mud, straight chain saturated fatty acid had the highest content, occupying 50.7-73.3% of total PLFA and being the highest in 300-year pit. As for the microbial community structure, the PLFA content characterizing Gram-positive (G+) anaerobic bacteria was higher in pit mud, and that characterizing Gram-negative (G-) anaerobic bacteria was higher in fermented grains and yellow water. The PLFA content characterizing G+ and G- bacteria in the pit mud of 100-year pit was higher than that in the pit mud of other ages' pits, while the PLFA content characterizing fungi was higher in the pit mud, fermented grains, and yellow water of 5-year pit, as compared with other ages' pits. Principal component analysis showed that the main varied microbial populations in 5- and 100-year pits were G- bacteria and fungi, and the main varied microbial population in 300-year pit was of bacteria. The indices frequency index, Simpson index, and Shannon index could be chosen for characterizing the diversity of microbial community in fermentation pits.

Bactérias/classificação , Reatores Biológicos/microbiologia , Fermentação , Fosfolipídeos/metabolismo , Vinho/microbiologia , Bactérias/crescimento & desenvolvimento , Bactérias/metabolismo , Biodiversidade , Contagem de Colônia Microbiana , Ácidos Graxos/metabolismo , Dinâmica Populacional
Exp Ther Med ; 1(2): 277-283, 2010 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-22993540


To investigate the in vivo and in vitro inhibitory effects of deuterium-depleted water (DDW) on human lung cancer and the possible mechanisms underlying these effects, we cultured and treated human lung carcinoma cell line A549 and human embryonic lung fibroblasts HLF-1 with various concentrations of DDW from 2 to 72 h. Cellular growth inhibition rates were determined using the 3-(4, 5-dimethyldiazol-2-yl)-2, 5-diphenyltetrazolium-bromide) (MTT) proliferation assay. A549 cells were treated with 50±5 ppm DDW, and the morphology and structure of cells were observed by scanning electron microscopy (SEM). We observed alterations in the cellular skeleton by transmission electron microscopy (TEM) and changes in cell cycle by flow cytometry. Our data showed that DDW significantly inhibited the proliferation of A549 cells at a specific time point, and cells demonstrated the characteristic morphological changes of apoptosis under SEM and TEM. The length of the S phase increased significantly in cells treated with 50 ppm DDW, whereas the G0 to G1 phase and G2 to M phase were decreased. We observed DDW-induced cellular apoptosis using terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) and DNA fragment analyses. In addition, we established a tumor transplantion model by injecting H460 tumor cells into subcutaneous tissue of BALB/c mice treated with DDW for 60 days. We determined the tumor inhibition rate of treated and control groups and found that the tumor weight was significantly decreased and the tumor inhibition rate was approximately 30% in the DDW group. We conclude that DDW is a promising new anticancer agent with potential for future clinical application.