一株嗜温高效产氢细菌Clostridium sp.08-1的分离鉴定与产氢特征

从10L连续搅拌式罐式反应器(CSTR)中分离得到1株嗜温高效产氢菌株08-1.根据菌株的形态特征和16S rDNA序列结果分析,初步鉴定菌株08-1属于Clostridium sp..同时还进一步研究了温度、pH值控制、底物浓度和种类对菌株08-1产氢的影响.结果表明.该菌株更适合利用蔗糖或成分复杂的生物质木薯粉以及废弃物厨余垃圾生长及产氢,最适产氢温度为40℃,产氢系统pH值控制在5.5时获得最大产氢量.在间歇发酵中,蔗糖浓度为20 g/L,控制温度40℃,pH值5.5,搅拌速度100 r/min时实现最大产氢速率为245 mL·(L·h)~(-1),最大产氢量达到3.06 mol.该菌株在生物制氢中具有潜在的应用价值.     

Continuous production of butanol with immobilized cells ofClostridium acetobutylicum

Spores ofClostridium acetobutylicum were immobilized in calcium alginate. An active gel preparation was obtained after outgrowth of the spores to vegetative cells within the gel matrix. A 100 mL column containing the immobilized cells was used for continuous production. At steady-state conditions the productivity of butanol was 67 g/L reactor volume/day.     

Discovery of Clostrubin,an Exceptional Polyphenolic Polyketide Antibiotic from a Strictly Anaerobic Bacterium

Genome mining of the strictly anaerobic bacterium Clostridium beijerinckii, an industrial producer of solvents, revealed the presence of several cryptic gene clusters for secondary metabolite biosynthesis. To unearth its metabolic potential, a C. beijerinckii strain was cultured under various conditions, which led to the discovery of a deep purple pigment. This novel metabolite, named clostrubin ( 1 ), was isolated and its structure was fully elucidated. The pentacyclic polyphenol features a benzo[a]tetraphene ring topology that is unprecedented for natural products. Stable‐isotope labeling experiments showed that 1 is an aromatic polyketide that folds in a noncanonical manner to form the unusual perifused ring system. In addition to being the first reported polyketide from an anaerobic bacterium, 1 is a potent antibiotic with pronounced activity against various pathogenic bacteria, such as MRSA, VRE, and mycobacteria, with minimum inhibitory concentrations (MIC) of 0.12–0.97 μM .     

Application of quantitative IR spectral analysis of bacterial cells to acetone-butanol-ethanol fermentation monitoring

Clostridium acetobutylicum ATCC 824 was grown under three different acetone-butanol-ethanol (ABE) fermentation conditions: (1) strictly anaerobic conditions with vegetative inoculum; (2) semi-anaerobic conditions with vegetative inoculum; and (3) strictly anaerobic conditions with spore inoculum. Semi-anaerobic fermentation with vegetative inoculum and strictly anaerobic fermentation with spore inoculum produced solvents at high level. Strictly anaerobic fermentation with vegetative inoculum showed an “acid crash”, i.e. produced mainly acids and did not switch to predominant solvent production. The content of carbohydrates, nucleic acids, proteins and lipids in Clostridium cells during the fermentation were evaluated from the mid-IR spectra. The content of nucleic acids decreased with process time, and the lipid content increased, corresponding to ceasing growth and formation of the toxic fermentation products. It was shown that the physiological states of either solvent production or acid crash are reflected in the microbial biomass composition, which can be assessed by IR spectroscopy.     

Continuous production of butanol from starch-based packing peanuts

Acetone, butanol, ethanol (ABE, or solvents) were produced from starch-based packing peanuts in batch and continuous reactors. In a batch reactor, 18.9 g/L of total ABE was produced from 80 g/L packing peanuts in 110 h of fermentation. The initial and final starch concentrations were 69.6 and 11.1 g/L, respectively. In this fermentation, ABE yield and productivity of 0.32 and 0.17 g/(L·h) were obtained, respectively. Compared to the batch fermentation, continuous fermentation of 40 g/L of starch-based packing peanuts in P2 medium resulted in a maximum solvent production of 8.4 g/L at a dilution rate of 0.033 h⁻¹. This resulted in a productivity of 0.27 g/(L·h). However, the reactor was not stable and fermentation deteriorated with time. Continuous fermentation of 35 g/L of starch solution resulted in a similar performance. These studies were performed in a vertical column reactor using Clostridium beijerinckii BA101 and P2 medium. It is anticipated that prolonged exposure of culture to acrylamide, which is formed during boiling/autoclaving of starch, affects the fermentation negatively.     

Properties of acetate kinase activity inClostridium thermocellum cell extracts

Acetate kinase (EC 2.7.2.1) is involved in the wasteful production of acetate during conversion of cellulose to ethanol byClostridium thermocellum. The properties of this enzyme activity inC. thermocellum cell extracts were determined. Optimum enzyme activity was at 60 degrees C and between pH 7.5 and 9.0. In the presence of air, acetate kinase was stable to temperatures up to 60 degrees C, retaining 90% activity after 2 h, and was inactivated rapidly at higher temperatures. The enzyme exhibited a wide range of stability to pH (5.0-9.0) when incubated at 50 degrees C for 2 h. As with other acetate kinases, a divalent cation, such as Mg(2+), was required for enzyme activity. Optimum activity was observed at 20mM MgCl(2) when ATP was held constant at 10 mM. Acetate kinase activity was adversely affected by KCl, a salt commonly used in ion-exchange or affinity chromatography, with 0.3M KCl inhibiting by 50%. These results will be important in optimizing the direct microbial conversion process of cellulose to ethanol usingC. thermocellum in coculture withClostridium thermosaccharolyticum.     

Simultaneous saccharification and fermentation of cellulosic biomass to acetic acid

Astrain of Clostridium thermoaceticum (ATCC 49707) was evaluated for its homoacetate potential. This thermophilic anaerobe best produces acetate from glucose at pH 6.0 and 59°C with a yield of 83% of theoretical. Enzyme hydrolysis of two substrates, a-cellulose and a pulp mill sludge, yielded 68% and 70% digestion, respectively. The optimum conditions for the simultaneous saccharification and fermentation (SSF) were substrate dependent: 55°C, pH 6.0 for α-cellulose, and 55°C, pH 5.5 for the pulp mill sludge. In the SSF with α-cellulose, the overall yield of acetate was strongly influenced by the enzyme loading. In a fed-batch operation of SSF with α-cellulose, an overall acetic acid yield of 60 wt% was obtained. Among the factors limiting the yields were incomplete digestion by the enzyme and the end-product inhibition. In the SSF of pulp mill sludge, inhibitors present in the sludge severely limited bacterial action. A large accumulation of glucose developed over the entire process, changing the intended SSF operation into a separate hydrolysis and fermentation operation. Despite a long lag phase of microbial growth, a terminal yield of 85% was obtained with this substrate.     

Discovery of Clostrubin,an Exceptional Polyphenolic Polyketide Antibiotic from a Strictly Anaerobic Bacterium

Genome mining of the strictly anaerobic bacterium Clostridium beijerinckii, an industrial producer of solvents, revealed the presence of several cryptic gene clusters for secondary metabolite biosynthesis. To unearth its metabolic potential, a C. beijerinckii strain was cultured under various conditions, which led to the discovery of a deep purple pigment. This novel metabolite, named clostrubin ( 1 ), was isolated and its structure was fully elucidated. The pentacyclic polyphenol features a benzo[a]tetraphene ring topology that is unprecedented for natural products. Stable‐isotope labeling experiments showed that 1 is an aromatic polyketide that folds in a noncanonical manner to form the unusual perifused ring system. In addition to being the first reported polyketide from an anaerobic bacterium, 1 is a potent antibiotic with pronounced activity against various pathogenic bacteria, such as MRSA, VRE, and mycobacteria, with minimum inhibitory concentrations (MIC) of 0.12–0.97 μM .     

Total Synthesis of Five Lipoteichoic acids of Clostridium difficile

The emergence of hypervirulent resistant strains have made Clostridium difficile a notorious nosocomial pathogen and has resulted in a renewed interest in preventive strategies, such as vaccines based on (synthetic) cell wall antigens. Recently, the structure of the lipoteichoic acid (LTA) of this species has been elucidated. Additionally, this LTA was found to induce the formation of protective antibodies against C. difficile in rabbits and mice. The LTA from C. difficile is isolated as a microheterogenous mixture, differing in size and composition, impeding any structure–activity relationship studies. To ensure reliable biological results, pure and well‐defined synthetic samples are required. In this work the total synthesis of LTAs from C. difficile with defined chain length is described and the initial biological results are presented.     

基于Logistic回归建模和马尔可夫链蒙特卡罗方法计算后验描述丁酸梭菌株对于给定辐照剂量区的应答趋势

利用马尔可夫链蒙特卡罗（Markov Chain Monte Carlo,MCMC）方法估计Logistic回归模型中的参数,就是要构造一个以参数的后验分布为其平稳分布的非周期不可约的马尔可夫链,然后用该平稳分布中抽出的样本点计算蒙特卡罗积分。上述理论方法可以解决实验样本数据由于存在定和约束和多重共线性、在进行经典的logistic回归建模时的困难问题。基于此方法,研究了丁酸梭菌株对于给定辐照区间剂量的应答趋势,用模型挖掘数据所隐含的内在信息并导出了Logistic回归模型参数的贝叶斯框架下的50%,90%,95%和99%的置信区间。结果表明,运用Logistic与马尔可夫链耦合模型在有关给定辐射剂量对于微生物作用效果问题的logistic回归建模中具有较大的科学性与很好的使用性,从而可以为辐照诱变处理微生物制定辐照剂量区提供理论支持和回归技术借鉴。Using the Markov Chain Monte-Carlo method to estimate the parameters in the Logistic regression model, we constructed a non-periodic irreducible Markov Chain with the posterior distribution of the parameters as stationary distribution, and then used the sample points extracted from the stationary distribution to calculate the Monte-Carlo integral. The above theoretical method can solve the difficult problem of classical logistic regression modeling because of the existence and limitation of the experimental sample data and the multicollinearity. In the classical regression setup with a continuous response, the predicted values can range over all real numbers. Therefore, a different modelling technique is needed. In this work, the results describe in detail a previously unknown lethality trend following 12C6+ heavy-ion irradiation of Clostridium tyrobutyricum. By Markov Chain Monte-Carlo can calculate the model fit for a randomly selected subset of the chain and calculate the predictive envelope of the model. The grey areas in the plot correspond to 50%, 90%, 95%, and 99% posterior regions. More importantly, although this study focused on the use of the method in heavy-ion irradiation of microbial, its results are broadly applicable.