酿酒用米曲霉若干菌株的培养特性研究

对国内目前使用较广的米曲霉4#,5#,6#及日本生产用米曲霉菌株1#,2#,3#的产酶作对比试验,研究了培养温度、培养基、初始酸度和培养时间对产酶的影响,分析米曲霉制作过程中的理化性质变化,探索其最佳制曲条件。本实验三角瓶米曲霉糖化力可达1400mg葡萄糖/g曲·h,产酶最适品温为37～38℃,最适pH为5～6;用5#制的曲糖化力和酸性蛋白酶活力较高。     

A standard set of testing methods reliably enumerates spores across commercial milk powders

Contamination of dairy powders with sporeforming bacteria is a concern for dairy processors who wish to penetrate markets with stringent spore count specifications (e.g., infant powders). Despite instituted specifications, no standard methodology is used for spore testing across the dairy industry. Instead, a variety of spore enumeration methods are in use, varying primarily by heat-shock treatments, plating method, recovery medium, and incubation temperature. Importantly, testing the same product using different methodologies leads to differences in spore count outcomes, which is a major issue for those required to meet specifications. As such, we set out to identify method(s) to recommend for standardized milk powder spore testing. To this end, 10 commercial milk powders were evaluated using methods varying by (1) heat treatment (e.g., 80°C/12 min), (2) plating method (e.g., spread plating), (3) medium type (e.g., plate count milk agar), and (4) incubation time and temperature combinations (e.g., 32°C for 48 h). The resulting data set included a total of 48 methods. With this data set, we used a stepwise process to identify optimal method(s) that would explain a high proportion of variance in spore count outcomes and would be practical to implement across the dairy industry. Ultimately, spore pasteurized mesophilic spore count (80°C/12 min, incubated at 32°C for 48 h), highly heat resistant thermophilic spore count (100°C/30 min, incubated at 55°C for 48 h), and specially thermoresistant spore enumeration (106°C/30 min, incubated at 55°C for 48 h) spread plating on plate count milk agar were identified as the optimal method set for reliable enumeration of spores in milk powders. Subsequently, we assessed different powder sampling strategies as a way to reduce variation in powder spore testing outcomes using our recommended method set. Results indicated that 33-g composite sampling may reduce variation in spore testing outcomes for highly heat resistant thermophilic spore count over 11-g and 33-g discrete sampling, whereas there was no significant difference across sampling strategies for specially thermoresistant spore enumeration or spore pasteurized mesophilic spore count. Finally, an interlaboratory study using our recommended method set and a modified method set (using tryptic soy agar with 1% starch) among both university and industry laboratories showed increased variation in spore count outcomes within milk powders, which not only was due to natural variation in powders but also was hypothesized to be due to technical errors, highlighting the need for specialized training for technicians who perform spore testing on milk powders. Overall, this study addresses challenges to milk powder spore testing and recommends a method set for standardized spore testing for implementation across the dairy industry.     

一株凝结芽孢杆菌（Bacillus coagulans）发酵培养基的优化

以凝结芽孢杆菌（Bacillus coagulans）为研究对象,芽孢数为响应值,对其发酵培养基进行响应面优化分析。 在单因素试验的 基础上,采用Plackett-Burman试验筛选出对芽孢数有显著影响的因素为麸皮和牛肉膏。 由中心组合试验设计及响应面分析优化确定 最优发酵培养基配方为麸皮32.12 g/L,牛肉膏15.24 g/L,硫酸镁0.49 g/L,硫酸锰0.34 g/L,磷酸二氢钠0.31 g/L。 在此最佳条件下,发酵 液中凝结芽孢杆菌芽孢数达到3.38×108 CFU/mL,是优化前的5.28倍。     

超声协同诱导剂对枯草芽孢杆菌芽孢致死的作用

为延长食品货架期,实现中温条件下的芽孢杀灭,以枯草芽孢杆菌为对象,研究了超声处理、热处理和不 同种类诱导剂协同作用对枯草芽孢杆菌芽孢致死的最适条件。结果表明：采用温度60 ℃、功率600 W、频率25 kHz 的超声方式,辅以50 mmol/L L-丙氨酸和6 mmol/L肌苷共同作用,可以使芽孢萌发率达到98.23%。进一步改变超 声处理模式,同等条件下,采用900、600、300、100 W的功率递减超声处理,能够有效提高芽孢萌发时的处理效 率,实现体系内芽孢的大量萌发和杀灭,保证食品安全和品质。     

生孢梭菌芽孢萌发条件的优化

本文研究了不同温度、时间和pH值条件下,生孢梭菌芽孢萌发所需的最佳条件.本研究将活化传代好的生孢梭菌菌液培养7 d,多次离心后,加入无菌水制成生孢梭菌芽孢悬浮液,处理温度为70℃～90℃,处理时间为5～25 min,处理pH值为4～8,用平板计数法观察计算不同条件处理后的芽孢萌发率,用响应面优化法对影响生孢梭菌芽孢萌发...     

酿造工业中黑曲霉孢子检测方法的改良

根据黑曲霉孢子菌的特征，对其检测技术进行探讨。筛选出检测无菌液，找出适于此类菌系检测方法关键技术。研究结果表明，以０．１５％琼脂无菌液代替无菌水对样品进行稀释，每个稀释液制成后，用磁力搅拌器搅拌，可使样品达到均匀分散程度，不再产生上浮、聚集的不均匀现象，再经接种、培养，即可获得准确结果，从根本上解决了原稀释平板法检测孢子菌类数值重现性差、难以确定真值的问题。     

响应面法优化卡氏芽孢杆菌ST-1产芽孢发酵培养基

以卡氏芽孢杆菌(Bacillus cabrialesii)ST-1为研究对象，芽孢数为响应值，采用单因素试验、Plackett-Burman试验、最陡爬坡试验及响应面试验对其发酵培养基进行优化。结果表明，影响卡氏芽孢杆菌ST-1芽孢数的主要因素为蔗糖、麸皮和蛋白胨、磷酸二氢钾，卡氏芽孢杆菌ST-1产芽孢的最适宜培养基配方为：蔗糖13.1 g/L，麸皮+蛋白胨(1∶2)17.0 g/L，磷酸二氢钾2.0 g/L。在此最优条件下，卡氏芽孢杆菌ST-1发酵液中芽孢数达到5.96×10⁹ CFU/mL，是优化前的32.21倍。     

Ger1 is a secreted aspartic acid protease essential for spore germination in Ustilago maydis

Ustilago maydis expresses a number of proteases during its pathogenic lifecycle. Some of the proteases including both intracellular and extracellular ones have previously been shown to influence the virulence of the pathogen. However, any role of secreted proteases in the sporulation process of U. maydis have not been explored earlier. In this study we have investigated the biological function of one such secreted protease, Ger1 belonging to aspartic protease A1 family. An assessment of the real time expression of ger1 revealed an infection specific expression of the protein especially during late phases of infection. We also evaluated any contribution of the protein in the pathogenicity of the fungus. Our data revealed an involvement of Ger1 in the sporulation and spore germination processes of U. maydis. Ger1 also showed positive influence on the pathogenicity of the fungus and accordingly the ger1 deletion mutant exhibited reduced pathogenicity. The study also demonstrated the protease activity associated with Ger1 to be essential for its biological function. Fluorescence microscopy of maize plants infected with U. maydis cells expressing Ger1-mcherry-HA also revealed that Ger1 is efficiently secreted within maize apoplast.     

Synthesis of Muramyl-δ-Lactam in Spore Peptidoglycan of Clostridioides difficile

Clostridioides difficile is a spore-forming human pathogen responsible for significant morbidity and mortality. Infections by this pathogen ensue dysbiosis of the intestinal tract, which leads to germination of the spores. The process of spore formation requires a transition for the cell-wall peptidoglycan of the vegetative C. difficile to that of spores, which entails the formation of muramyl-δ-lactam. We describe a set of reactions for three recombinant C. difficile proteins, GerS, CwlD, and PdaA1, with the use of four synthetic peptidoglycan analogs. CwlD and PdaA1 excise the peptidoglycan stem peptide and the acetyl moiety of N-acetyl muramate, respectively. The reaction of CwlD is accelerated in the presence of GerS. With the use of a suitable substrate, we document that PdaA1 catalyzes a novel zinc-dependent transamidation/transpeptidation reaction, an unusual reaction that requires excision of the stem peptide as a pre-requisite.