基于转录组学分析揭示蜡样芽胞杆菌(Bacillus cereus)AR1002阻控黄曲霉生长的生理机制研究

黄曲霉(Aspergillus flavus)极易侵染油料等农产品,其次级代谢产物黄曲霉毒素严重危害人体健康,阻控黄曲霉污染已成为亟待解决的国际难题。为探明蜡样芽胞杆菌(Bacillus cereus)AR1002抑制黄曲霉生长、阻控黄曲霉污染的生理及分子机制。该研究采用AR1002代谢产物对黄曲霉进行处理,并对黄曲霉生长及产孢表型进行鉴定,通过显微结构及转录组学分析,对AR1002的抑菌机制进行了初探。蜡样芽胞杆菌AR1002代谢物可抑制黄曲霉菌丝生长、干物质积累分别达24.73%,65.00%,减少孢子数量达98.80%;AR1002通过抑制glpA和AYR1等关键基因表达调控黄曲霉甘油磷酸代谢,通过抑制SEC61A、RAD23、ATP13A1、UBE2G2等关键基因的表达调控内质网中蛋白质加工;此外,AR1002代谢产物通过下调RAD51和RAD54B等DNA修复机制相关基因,抑制黄曲霉同源重组及非同源性末端连接过程,最终达到抑制黄曲霉生长及产孢的目的。上述结果表明,蜡样芽胞杆菌(Bacillus cereus)AR1002有望应用于实际生产中以应对黄曲霉污染问题。

蚯蚓联合生防细菌Bacillus velezensis改善连作百合土壤细菌群落结构及防治枯萎病的效果

【目的】从土壤理化性状和微生物群落结构及多样性角度,研究蚯蚓联合生防细菌改善百合连作障碍的机理和效果,为湖南百合产业化种植提供技术措施。【方法】采用盆栽试验方法,在湖南长沙连续种植了两季百合(Lilium lancifolium Thunb.)。供试土壤为红壤,生防菌为Bacillales velezensis YFB3-1菌液(菌体浓度为10^(9) CFU/mL),蚯蚓为赤子爱胜蚓(Eisenia fetida)。试验设单施菌液(T1)、只接种蚯蚓(T2)、同时使用菌液和蚯蚓(T3) 3个处理,以不使用菌剂和蚯蚓为对照(CK)。调查了两茬百合产量和枯萎病病情指数,分析了第二茬百合根际土壤理化性质、细菌群落结构和多样性,以及连作百合产量与根际土壤理化性质、细菌群落和枯萎病的相关性。【结果】与第一茬相比,同一处理的第二茬百合产量都显著下降。两年连作百合产量都以同时使用菌液和蚯蚓(T3)处理为最高,分别为16464、15674 kg/hm^(2),都显著高于同年份其他处理。两年连作百合枯萎病发病率和病情指数都以T3为最低,都显著低于同年份其他处理,T3处理百合枯萎病病情指数分别为18.15(2019年)、25.00 (2020年),并且两年的防治效果(T3)都明显优于单施菌液(T1)和接种蚯蚓(T2)。相比于对照(CK),2019、2020年T3处理百合枯萎病防治效果分别为44.55%、37.66%,分别增产15.05%、14.78%。蚯蚓联合生防菌YFB3-1增加了根际土壤Rhizobiales、Flavobacteriales、Pseudomonadales、Bacillales等有益微生物的种群丰度,蚯蚓与生防菌YFB3-1在抑制连作百合枯萎病方面具有显著的互作效应,因而缓解连作百合产量下降的效果最佳。【结论】在施用芽孢杆菌生防菌剂防治连作百合枯萎病的同时,可以增施有机肥(牛粪)来提高土壤中蚯蚓数量,有效预防百合枯萎病发生,减少连作百合产量损失。

莫海威芽孢杆菌(Bacillus mojavensis)ZLX10对发酵雪茄烟叶中性致香成分的影响

为探究烟草源微生物单菌发酵对雪茄烟叶挥发性致香成分的影响,采用GC-MS检测莫海威芽孢杆菌(Bacillus mojavensis)ZLX10发酵雪茄烟叶中性致香成分,筛选差异致香成分,并与感官评价进行相关性分析,结合ROAV值变化,明确ZLX10提升雪茄烟叶感官品质的关键致香成分。结果表明,ZLX10发酵雪茄烟叶后,香气总量和大多数中性致香成分含量均显著提高,共筛选得到15种差异中性致香成分,共有10种致香成分显著上调(P<0.05),其中巨豆三烯酮A、巨豆三烯酮B、6-甲基-5-庚烯-2-酮、2,6,6-三甲基-2-环己烯-1,4-二酮、茄酮和植物醇与烟叶香气呈显著正相关(P<0.05),可认为是增加烟叶干香和花香的关键差异致香成分;共有5种致香成分显著下调(P<0.05),其中苯乙醛和2,4-二叔丁基-6-硝基苯酚与烟叶刺激性呈显著正相关(P<0.05),对烟叶香气有较大的负面影响。由此可知,ZLX10发酵能促进雪茄烟叶香气质和香气量得到提升,这为人工接菌发酵雪茄烟叶提供了基础。

1株抗真菌贝莱斯芽孢杆菌(Bacillus velezensis)lut-Y1的鉴定及抑菌活性研究

【目的】探究1株来自枸杞果实的贝莱斯芽孢杆菌lut-Y1的生长特性及抑菌活性,为微生态制剂的开发和应用提供参考。【方法】通过形态观察和16S rDNA基因测序分析对该菌株进行鉴定;测定该菌株生长曲线及最适pH、碳源、氮源及盐耐受度;采用平板对峙法观察该菌株对链格孢菌(Alternaria alternate)、小麦根腐离蠕孢(Bipolaris sorokiniana)和小孢壳二孢(Ascochyta leptospora)的颉颃作用,并分析该菌株发酵液对3种病原真菌的抑菌活性。【结果】lut-Y1菌株在LB、NA和PDA培养基上均生长良好。根据形态和16S rDNA基因序列特征将lut-Y1菌株鉴定为贝莱斯芽孢杆菌(Bacillus velezensis)。在LB液体培养基中,lut-Y1菌株在0~3 h为迟滞期,3~18 h为对数期,18~62 h为稳定期,62 h后进入衰亡期;最适pH为7.0,最适碳源和氮源分别为蔗糖和蛋白胨,对NaCl的耐受量不超过2.0 g/L。lut-Y1菌株对链格孢菌有明显的颉颃作用,对小麦根腐离蠕孢和小孢壳二孢的颉颃作用较弱。对峙平板的两菌交界处,链格孢菌基内菌丝细弱、弯曲;小麦根腐离蠕孢基内菌丝细胞壁被破坏,有泡状突起。发酵液添加量为25%时生长圈直径分别缩小72.80%和83.24%。【结论】贝莱斯芽孢杆菌lut-Y1具有较好的抑菌活性,可为开发以芽孢杆菌为主要活性成分的微生态制剂提供材料。

Mn^(2+)促进Bacillus altitudinis LZP02生物膜形成

【目的】Bacillus altitudinis LZP02是一株水稻根际促生菌(PGPR),能定殖于水稻根部,形成生物膜,且Mn^(2+)对LZP02菌株生物膜形成具有促进作用,但调控机制尚不明确,旨在探究Mn^(2+)对LZP02菌株生物膜形成的促进机制。【方法】采用结晶紫染色法进行生物膜定量分析、蒽酮-硫酸法测定胞外多糖产量、扫描电镜(SEM)观察LZP02菌株在水稻根际定殖情况、转录组学测序技术分析差异表达基因(DEGs)。【结果】添加4 mmol/L Mn^(2+)和8 mmol/L Mn^(2+)显著提高了LZP02菌株的成膜能力和胞外多糖产量,扫描电镜发现4 mmol/L Mn^(2+)和8 mmol/L Mn^(2+)提高了LZP02菌株在水稻根际的定殖能力,随着Mn^(2+)浓度的增加,DEGs数量显著增多,其主要富集在芽孢形成、毒素代谢途径和双组分系统中。1 mmol/L Mn^(2+)和4 mmol/L Mn^(2+)处理组发现skf操纵子中基因的表达均上调。扫描电镜观察发现在1 mmol/L Mn^(2+)和4 mmol/L Mn^(2+)处理组中LZP02菌体存在损伤。4 mmol/L Mn^(2+)处理组双组分系统中KinE和Spo0A基因均显著上调。【结论】Mn^(2+)通过“嗜食同类”和双组分系统中KinE基因激活Spo0A~P提高了菌株LZP02的成膜能力。

纳豆芽孢杆菌Bacillus natto NK4液态发酵产纳豆激酶的工艺优化

为提高纳豆激酶的产量,采用单因素试验及正交试验对纳豆芽孢杆菌Bacillus natto NK4进行发酵条件优化,获得该菌种液体发酵产酶的最优条件。结果表明,纳豆芽孢杆菌产纳豆激酶最佳的发酵条件为接种量2%,培养基初始pH值6.0,培养温度34℃,发酵时间72 h。此时,纳豆激酶酶活力由848.28 IU/mL提高到1569.31 IU/mL。

矿化细菌Bacillus paramycoides对重金属Pb(Ⅱ)和Cd(Ⅱ)的去除及矿化特性

微生物诱导生物矿化可有效固定重金属离子,被认为是一种重要的重金属污染环境修复工艺,对重金属污染生态修复有重要的现实意义。从贵州黔西北某铅锌矿区周边土壤中分离出一株耐Pb(Ⅱ)和Cd(Ⅱ)菌株GS-2,经形态学特征和16S rRNA鉴定为Bacillus paramycoides,菌株GS-2对Pb(Ⅱ)和Cd(Ⅱ)的耐受分别高达1500和50 mg/L,对250 mg/L Pb(Ⅱ)和5 mg/L Cd(Ⅱ)去除率分别为99.54%和69.04%,对重金属Pb(Ⅱ)和Cd(Ⅱ)(均为25 mg/L)混合去除率分别为99.73%和71.46%。SEM-EDS、FTIR和XRD结果显示,菌株通过表面的羟基、羧基、酰胺基和磷酸基团等官能团将Pb(Ⅱ)和Cd(Ⅱ)诱导矿化成Ca_(2.5)Pb_(7.5)(OH)_(2)(PO_(4))_(6)、Ca_(2.1)Pb_(7.9)Cl_(2)(PO_(4))_(6)、Pb_(3)(PO_(4))_(2)、Pb_(5)(PO_(4))_(3)OH、Pb_(9)(PO_(4))_(6)、Pb_(10)(PO_(4))_(6)(OH)_(2)和Ca_(0.67)Cd_(0.33)CO_(3)等晶体矿物。菌株GS-2具有较好的去除和矿化能力,对环境中Pb(Ⅱ)和Cd(Ⅱ)污染的生物修复具有一定应用潜力。

Antiseptic Efficacy of A Soap Made from Biosurfactants Isolated from Bacillus and Lactobacillus against Pathogenic Bacteria

The aim of our study was to use a biosurfactant produced by Bacillus and Lactobacillus isolates as an antiseptic in the formulation of local soap. A total of 60 isolates were characterized by microbiological techniques (30 Bacillus and 30 Lactobacillus) and the ability to produce biosurfactants was demonstrated by a hydrocarbon emulsification index (E24). The emulsification indexes (E24) varied from 9% to 100% for Bacillus and from 33% to 100% for Lactobacillus as well. The antagonistic assay showed that biosurfactants were able to inhibit the formation of biofilms and growth of pathogens such as Escherichia coli, Pseudomonas aeruginosa, Staphylococcus aureus, Bacillus cereus, Salmonella typhirium, Shigella boydii and Proteus mirabilis. The biosurfactant consortium (BioC) from Bacillus consortium and from Lactobacillus was able to inhibit biofilm formation and the pathogens growth. The BioC was stable to alkaline pH and the temperatures stability of Biosurfactant was ranging from 50°C to 90°C. The soap was made by the cold saponification process using one biosurfactant consortium formulated. This soap has a pH of 10 and showed good cleaning power and good foam stability. Similarly, the soap showed good antiseptic power and disinfection power against all pathogens tested. Handwashing is critical to preventing disease transmission. The persistence of pathogens in waste water was evaluated. The BioS produced showed good disinfection power against all pathogens tested. The valor of reduction on the hands and in the waste water was significantly more than compared to the control soaps used. This soap could be used in the prevention, fighting, and treatment of bacterial and viral infections.

Bacillus tropicus来源β-D-呋喃果糖苷酶基因的克隆与表达及其酶学特性研究

为获得新β-D-呋喃果糖苷酶资源并探究其酶学性质及在转化糖中的应用潜力,挖掘出Bacillus tropicus来源的β-D-呋喃果糖苷酶基因BtFFase8,成功合成后表达在大肠杆菌宿主中。通过数据库筛选β-D-呋喃果糖苷酶的基因序列,将基因克隆至载体pET-28a(+)中。将获得的重组质粒转化至大肠杆菌BL21,构建重组菌株大肠杆菌BL21/pET28a-BtFFase8。采用亲和层析纯化克隆酶,用SDS-PAGE法分析蛋白分子量,通过葡萄糖试剂盒检测酶水解产物的能力得到酶活力,用福林酚试剂法测定蛋白质含量。克隆酶BtFFase8的分子质量为57.0 kDa,对蔗糖底物的比酶活力为13.4 U/mg;BtFFase8的最适pH和最适温度分别为pH 6.5和45℃,且在pH 5.0~8.0和40℃及以下温度保持酶活力稳定,残余酶活力大于80%;BtFFase8能耐受碱性蛋白酶、酸性蛋白酶和胰蛋白酶的水解,残余酶活力大于70%。BtFFase8的发现及克隆酶优良的酶学功能为其未来应用于食品加工,特别是在转化糖的生产中奠定了良好的理论基础。

Research advance of Bacillus velezensis:bioinformatics,characteristics,and applications

Bacillus velezensis is a Gram-positive and spore-forming bacterium.It has potent antimicrobial properties that can be used to promote plant growth and as a pesticide by inhibiting pathogens.B.velezensis has the capability to generate a diverse range of enzymes that have potential applications in various fields,such as enzyme production,fermented food,degradation of pollutants,and bioenergy.In addition,B.velezensis is a promising probiotic.It possesses high bile-salt tolerance characteristics and has a high success rate of colonization in the intestinal mucosa.Besides,the strain can also regulate gut microbiota constitute by increasing the number of beneficial microorganisms and decreasing the number of pathogens.Furthermore,based on its special properties,including high-yield protease production and high salt-tolerance,B.velezensis shows potential for use in marine protein fermentation,opening up new avenues for the development of novel food products and bioactive peptides.In addition,B.velezensis can shorten the fermentation time as well as improve the nutritional value and flavor of fermented food.The safety of B.velezensis for food production was evaluated.This review provides valuable insights into the potential uses and benefits of B.velezensis,particularly in the context of fermented foods.

Enhanced extracellular production of alpha-lactalbumin from Bacillus subtilis through signal peptide and promoter screening

Alpha-lactalbumin(α-LA)is a major whey protein found in breast milk and plays a crucial role in the growth and development of infants.In this study,Bacillus subtilis RIK1285 harboring AprE signal peptide(SP)was selected as the original strain for the production ofα-LA.It was found thatα-LA was identified in the pellet after ultrasonic disruption and centrifugation instead of in the fermentation supernatant.The original strain most likely only producedα-LA intracellular,but not extracellular.To improve the expression and secretion ofα-LA in RIK1285,a library of 173 homologous SPs from the B.subtilis 168 genome was fused with target LALBA gene in the pBE-S vector and expressed extracellularly in RIK1285.SP YjcN was determined to be the best signal peptide.Bands in supernatant were observed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and purified by nickel column to calculate the highest yield signal peptide.In addition,different promoters(P_(aprE),P_(43),and P_(glv))were compared and applied.The results indicated that the strain RIK1285-pBE-P_(glv)-YjcN-LALBA had the highestα-LA yield,reaching 122.04μg/mL.This study demonstrates successful expression and secretion of humanα-LA in B.subtilis and establishes a foundation for simulating breast milk for infant formulas and developing bioengineered milk.

Bioleaching of vanadium from stone coal vanadium ore by Bacillus mucilaginosus:Influencing factors and mechanism

Vanadium and its derivatives are used in various industries,including steel,metallurgy,pharmaceuticals,and aerospace engineering.Although China has massive reserves of stone coal resources,these resources have low grades.Therefore,the effective extraction and recovery of metallic vanadium from stone coal is an important way to realize the efficient resource utilization of stone coal vanadium ore.Herein,Bacillus mucilaginosus was selected as the leaching strain.The vanadium leaching rate reached 35.5%after 20 d of bioleaching under optimal operating conditions.The cumulative vanadium leaching rate in the contact group reached 35.5%,which was higher than that in the noncontact group(9.3%).The metabolites of B.mucilaginosus,such as oxalic,tartaric,citric,and malic acids,dominated in bioleaching,accounting for 73.8%of the vanadium leaching rate.Interestingly,during leaching,the presence of stone coal stimulated the expression of carbonic anhydrase in bacterial cells,and enzyme activity increased by 1.335-1.905 U.Enzyme activity positively promoted the production of metabolite organic acids,and total organic acid content increased by 39.31 mg·L^(-1),resulting in a reduction of 2.51 in the pH of the leaching system with stone coal.This effect favored the leaching of vanadium from stone coal.Atomic force microscopy illustrated that bacterial leaching exacerbated corrosion on the surface of stone coal beyond 10 nm.Our study provides a clear and promising strategy for exploring the bioleaching mechanism from the perspective of microbial enzyme activity and metabolites.

Oral administration of Bacillus coagulans TQ-35 alleviates allergic responses in OVA-sensitive BALB/c mice

Bacillus coagulans has been extensively studied so far,but there has been a lack of research on its usage in allergy.In this study,we designed to assess the effect of different concentrations of B.coagulans on food allergy in a BALB/c mouse model of ovalbumin(OVA)-induced food allergy and its effect on gut microbes.The assessment of symptoms,specific immunoglobulin E(IgE),T-cell differentiation,and related gene expression levels in sensitized mice by assay indicated that high doses of oral B.coagulans could alleviate allergic symptoms.Treatment with B.coagulans,in the high-dose group,significantly reduced IgE and IgG1 levels and modulated the balance of T helper type 1 cell(Th1)and Th2 and the expression of relevant genes in the spleen.16S rRNA analysis showed that probiotics improved the structure of the microbiota,in particular by boosting the percentage of Clostridia,Bacteroides vulgatus and Enterococcus faecium,and by increasing the abundance of microbial species,thereby modulating the immune system.Therefore,this study can provide insights into the practical application of B.coagulans doses to alleviate OVA allergy.

Low-Level Antibiotic Resistance among Staphylococcus aureus and Gram-Negative Pathogens from Infected Skin and Soft Tissues in Rural Kenya

Introduction: Bacterial skin and soft tissue infections (SSTIs) are a cause of frequent inpatient and outpatient care visits whose causative agents are associated with a high antimicrobial resistance burden. For insights on antimicrobial susceptibilities in a rural setting, we examined specimens from suspected SSTIs from two public health facilities in Kenya. We additionally assessed antibiotic use, appropriateness of empiric therapy and risk factors for SSTI. Methodology: Between 2021 and 2023, 265 patients at Kisii and Nyamira County Referral hospitals were enrolled. Wound swabs/aspirates were collected and processed following standard microbiological procedures. Identification and antimicrobial susceptibility were performed using the VITEK 2 Compact platform. Demographic, clinical, and microbiological data were analyzed with R Statistical software. Results: S. aureus was isolated in 16.2% (43/265) of patients with a methicillin resistance (MRSA) proportion of 14% (6/43). While 13/15 drugs elicited susceptibilities ranging from 84% - 100%, penicillin (16%) and trimethoprim-sulfamethoxazole [TMP-SXT] (23%) yielded the lowest susceptibilities. Escherichia coli (n = 33), Klebsiella pneumoniae (n = 8), Pseudomonas aeruginosa (n = 8), and Citrobacter species (n = 4) were the most commonly isolated gram-negative species. Gram-negative strains showed high susceptibilities to most of the tested drugs (71% - 100%) with the exception of ampicillin (18%), TMP-SXT (33%), and first and second generation cephalosporins. Conclusions: The low MRSA prevalence and generally high antibiotic susceptibilities for S. aureus and gram-negative bacteria present opportunities for antibiotic stewardship in the study setting. Diminished susceptibilities against penicillin/ampicillin and TMP-SXT accord with prevailing local data and add a layer of evidence for their cautious empiric use.

Animal Safety Test of Bacillus thuringiensis BT Protein

[Objectives]To determine the biological safety of BT protein from Bacillus thuringiensis(Bt)fermentation broth to mammals at high doses.[Methods]Healthy mice were randomly divided into 4 groups with 10 mice in each group.The experimental groups were fed with Bt fermentation supernatant at 0.2,0.6 and 1.0 mL/kg,respectively,once a day for 7 consecutive days.The blank control group was fed normally without intragastric administration.[Results]There was no significant difference in blood routine and blood biochemical analysis between the experimental group and the control group.After intragastric administration,the mice were dissected,and no obvious pathological changes were found;the heart,liver,spleen,lung and kidney were taken to make tissue sections,and no pathological changes were found by microscopic observation.[Conclusions]Mice can tolerate high doses of BT protein from B.thuringiensis fermentation broth.

Isolation and Experimental Study on Bacteriostasis of Bacillus haynesii

This experiment aims to isolate and inhibit three bacteria strains to provide candidate strains for the development and application of probiotics.Using bacterial morphological identification,16S rDNA sequence alignment,and genetic evolution analysis,three strains were identified as Bacillus haynesii,named HP01,HD02,and HK03.Through biosurfactant activity tests,C-TAB tests,hemolysis tests,and antibacterial activity analyses,the results showed that all three strains of B.haynesii exhibited significant biosurfactant activity.Additionally,the solutions of the three strains demonstrated a pronounced antibacterial effect on Staphylococcus aureus.The resistance and safety of commonly used drugs were evaluated using the tablet diffusion method and a mouse feeding test.The results indicated that the three strains were not resistant to commonly used antibacterial drugs,and the oral bacterial solution was not pathogenic and had high safety in mice.The study concluded that all three B.haynesii strains met the basic conditions for use,with B.haynesii HP01 being the most promising candidate.

贝莱斯芽孢杆菌(Bacillus velezensis)MCCC 1A15695抑菌蛋白分离纯化及应用研究

对虾“玻璃苗”病的暴发,会导致对虾幼苗大批死亡,给水产养殖带来巨大损失。利用本实验室最近鉴定的“玻璃苗”致病菌副溶血弧菌(Vibrio parahaemolyticus)vp-HL,从海洋来源芽孢杆菌中筛选拮抗菌,获得一株能够高效抑制vp-HL菌株的贝莱斯芽孢杆菌(Bacillus velezensis)MCCC 1A15695。经分离鉴定与活性测定,其有效抑菌物质为蛋白类,该蛋白抑菌谱较广,对包括多种弧菌病原在内的17种病原微生物有抑制作用。经Q Seharose Fast Flow阴离子柱层析、聚丙烯酰胺凝胶电泳(SDS-PAGE)及液相色谱-质谱联用技术(LC-MS)分离纯化鉴定,确定活性蛋白组分中的有效抑菌物质为枯草杆菌素(subtilisin)、未知蛋白(hypothetical protein)及α-淀粉酶(Alpha-amylase)。蛋白类物质具有高生物安全性,本研究为后续益生菌的开发利用提供了理论基础。

Bacillus sp.B110胞内麦芽糖淀粉酶基因克隆与酶学特性

目的:对菌株Bacillus sp.B110的胞内麦芽糖淀粉酶BMAL进行基因克隆、异源表达、纯化及酶学性质研究,为后期开发新的淀粉加工用酶打下基础。方法:使用PCR技术对Bacillus sp.B110的胞内麦芽糖淀粉酶bmal基因序列进行全长克隆,异源表达,使用Ni2+-NTA进行纯化,再对其酶学特性进行测定,使用序列分析工具BioEdit、MEGA等对其氨基酸序列进行分析,使用AlphaFold2对其三级结构进行预测分析。结果:BMAL基因全长1770 bp,编码一个589氨基酸残基的蛋白。重组酶rBMAL经Ni2+-NTA亲和层析纯化后,SDS-PAGE电泳结果显示其分子量大小为63 kDa。氨基酸序列分析和三维建模表明BMAL与来源于B.subtilis 168和B.subtilis SUH4-2的麦芽糖淀粉酶有较高的一致性,且BMAL具有一个麦芽糖淀粉酶所独有的N端结构域以及由Asp328-Glu357-Asp424三个氨基酸残基所构成的催化中心。重组酶rBMAL最适反应温度为45℃,最适反应pH为6.0。重组酶rBMAL在30℃条件下保藏7 h残留酶活为60%,但在60℃条件下保藏2 h残留酶活力下降98%,说明BMAL对热敏感。重组酶rBMAL在4℃,pH7.0~9.5保藏12 h活性稳定。当存在1 mmol/L的金属离子Mg2+时,重组酶rBMAL活力提高36%,而Ni2+、Fe3+、Co2+、Cu2+、Zn2+、Al3+、Ca2+对重组酶rBMAL有抑制作用,酶活力减少85%~48%。有机溶剂和化学试剂甲醇、乙醇、丙酮、异腈、EDTA和SDS对重组酶rBMAL有较强的抑制作用,酶活力减少至32.3%~64.8%。底物特异性实验结果证实BMAL最适底物为环精糊。结论:Bacillus sp.B110的胞内麦芽糖淀粉酶BMAL具有良好的催化特性和pH稳定性,在面包烘焙工业上具有潜在的应用价值。

贝莱斯芽孢杆菌(Bacillus velezensis)YFB3-1高密度液体发酵条件优化

旨在获得高密度的贝莱斯芽孢杆菌(Bacillus velezensis)YFB3-1生防菌剂,用于预防作物土传病害的发生。采用单因素和正交试验对菌株YFB3-1的液体培养基和培养条件进行优化。在酵母粉8 g/L、复合氨基酸10 g/L、氯化钠1 g/L、硫酸镁(MgSO_(4)·7H_(2)O)1 g/L的液体培养基中培养菌株YFB3-1,初始pH7.0、接种量2.5%、发酵温度30℃、转速180 r/min和培养时间48 h为发酵条件时,获得的菌体浓度为2.89×10^(10)cfu/mL,是菌株YFB3-1液体发酵的最优培养基和培养条件,比牛肉膏蛋白胨液体培养基(NB)的菌体浓度4.77×10^(9)cfu/mL,提高了5.06倍,在相同时间内可以提高发酵效率。本研究为生防菌B velezensis YFB3-1的产业化扩大培养和推广应用提供了理论依据和技术参数。

U(Ⅵ)在芽孢杆菌Bacillus sp.dwc-2上的矿化动力学研究

微生物诱导的U(Ⅵ)生物矿化会影响铀在环境中的化学形态和迁移行为,但目前对于生长和繁殖阶段的微生物诱导U(Ⅵ)矿化的动力学行为研究不多,相关的机制也不明确。本文从某拟用作极低放射性废物处置场场所的土壤中分离出1株芽孢杆菌Bacillus sp.dwc-2,考察了pH值、甘油磷酸钠(SGP)、U(Ⅵ)初始浓度等因素对Bacillus sp.dwc-2在生长繁殖过程中诱导U(Ⅵ)矿化动力学行为的影响,并对其矿化机制进行了探讨。结果表明,Bacillus sp.dwc-2可诱导U(Ⅵ)形成钠铀云母(NaUO_(2)PO_(4)·3H_(2)O)的晶体矿物,该过程作为一种酶促反应,受pH值、SGP浓度、U(Ⅵ)初始浓度等因素的显著影响;当U(Ⅵ)初始浓度从50 mg/L增加至150 mg/L时,刺激了处于生长迟滞期至对数期(培养0~12 h)菌体磷酸酶活性的表达和磷酸盐的代谢,使其诱导U(Ⅵ)的矿化行为发生在生长对数期(培养12 h);SGP浓度的增加促进了磷酸酶的催化反应速率,进而促进了U(Ⅵ)的矿化,当SGP的浓度从25 mmol/L增加到125 mmol/L时,Bacillus sp.dwc-2诱导U(Ⅵ)的矿化时间从其生长稳定期(培养24 h)提前至生长对数期(培养12 h);中性和碱性条件下,磷酸酶活性在生长迟滞期至对数期(培养0~12 h)快速增加,从而促进了磷酸盐的消耗和U(Ⅵ)的矿化,特别是在碱性条件下,沉积物在生长对数期(培养8 h)时即转化为明显的晶体矿物。