大蒜汁对枯草芽孢杆菌抑制作用的研究

通过测定菌体浓度、抑菌圈直径和2,6-吡啶二羧酸(DPA)含量,研究大蒜汁对枯草芽孢杆菌(BS)的营养体及芽孢生长、发芽的影响,并采用响应面分析法优化确定大蒜汁抑菌适宜处理条件.结果表明:(1)大蒜汁对BS的最低抑菌浓度(MIC)和最低杀菌浓度(MBC)分别为0.4%和1%;(2)大蒜汁抑制作用主要是延长了BS的生长延缓期,0.3%的大蒜汁可使BS延缓期增加12 h;(3)大蒜汁对BS的芽孢和DPA形成有明显的抑制作用,但对芽孢的发芽无抑制作用;(4)加热温度超过35℃、时间大于5 h时处理的大蒜汁,对BS的抑制作用明显降低.在pH 3～8范围的大蒜汁都有很好的抑菌活性,但pH＞8.5时抑菌活性急剧下降;(5)响应面试验分析法优化确立了大蒜汁对BS抑制的二次回归方程和适宜处理条件,即在pH 4.5、温度45℃加热处理5 h的大蒜汁抑菌效果最好.     

枯草芽孢杆菌对几种灰霉病菌的抑制效果研究

分别研究了枯草芽孢杆菌(BacillussubtilisCohn)培养液、过滤液和灭活液对葡萄灰霉病菌(GB)、草莓灰霉病菌(SB)、辣椒灰霉病菌(PB)和番茄灰霉病菌(TB)菌丝生长的抑制作用。结果表明:枯草芽孢杆菌培养液对GB、SB、PB和TB都有很好的抑制作用。在菌液浓度达到10 5CFU/mL时,对4种灰霉病菌的抑制率均达到了10 0 % ;当浓度降低为10 4CFU/mL时,抑制率明显降抵。而菌液浓度为10 8CFU/mL时的过滤液,对GB、PB和TB的抑制率也均在5 0 %以上。灭活液对灰霉菌的抑制作用明显减弱,菌液浓度为10 8CFU/mL时,对PB、GB、TB和SB的抑制率分别为73.6 %、39.5 %、5 0 %和2 5 %。     

枯草芽孢杆菌B1-41对小麦纹枯病菌的抑制作用

从小麦根际土壤分离得到枯草芽孢杆菌(Bacillus subtilis)B1-41拮抗菌株,室内测定其带菌培养液对小麦纹枯病菌(Rhizoctonia cerealis)抑制效果为72%,盆栽试验中,其防治效果为60.3%,高于井冈霉素处理的51%的效果。试验表明,该菌株可以造成病菌菌丝发生畸变和菌丝细胞壁瓦解。     

枯草芽孢杆菌JA抗菌物特性的研究及抗菌肽的分离纯化

枯草芽孢杆菌JA分泌物中具有抑菌作用的粗提液通过DEAESepharose FF、SOURCE 15 PHE、Sephacry1 S200HR和反相HPLC多步柱层析分离纯化后,获得3种对水稻纹枯和小麦赤霉病菌均有抑菌作用的抗菌肽AFP1、AFP2和AFP3。MALDITOF质谱法测得其分子量分别为1462.645D、1476.390D和1490.530D。氨基酸组成分析结果表明,AFP1和AFP3由苏氨酸、异亮氨酸、酪氨酸、脯氨酸等多种氨基酸组成。目标产物热稳定性好、对蛋白酶有一定的耐受性,推断很可能是低分子量的环状脂肽。     

枯草芽孢杆菌感受态研究新进展

在枯草芽孢杆菌中,感受态的形成受到一种二元信号转导系统的调节,这种系统对胞外的感受态信息素浓度作出感应而激活晚期感受态基因的表达。各种晚期感受态蛋白分别负责外源DNA的吸附、吸收和内源化,它们共同构成了DNA的运输系统。初步探讨了枯草芽孢杆菌感受态调节在细胞生长和进化中的意义。     

枯草芽孢杆菌活菌体外拮抗6种肠道致病菌的研究

对枯草芽孢杆菌BS 3株在体外对 6种常见肠道致病菌 (肠产毒性大肠杆菌、肠侵袭性大肠杆菌、肠致病性大肠杆菌、鼠伤寒沙门菌、福氏志贺菌和宋内志贺菌 )的拮抗作用进行了研究。结果表明 ,枯草芽孢杆菌BS 3株对宋内志贺菌、肠致病性大肠杆菌及肠产毒性大肠杆菌拮抗作用较为明显。     

海参溶菌酶枯草芽孢杆菌基因工程菌构建

通过基因重组等技术构建重组海参溶菌酶的枯草芽孢杆菌基因工程菌,并对此基因工程菌进行生长曲线的测定和稳定性分析。结果表明,海参溶菌酶基因特异引物在400 bp处扩增出特异性条带,与预期的海参溶菌酶基因大小一致;重组表达质粒pHT43-SjLys经双酶切验证得8 000 bp和400 bp左右的片段;重组海参溶菌酶的枯草芽孢杆菌基因工程菌与原始菌株WB600相比生长趋势基本一致,外源基因的插入对菌体的生理代谢未造成太大影响;在无选择压力的条件下,重组质粒的稳定性良好,连续传5次后的遗传稳定性为94%,并且提取质粒和双酶切验证后发现工程菌没有发现重排或丢失现象。表明重组海参溶菌酶基因工程菌pHT43-SjLys/WB600构建成功。     

枯草芽孢杆菌ES224菌制剂防治烧伤的临床研究

本制品是根据微生态学原理,利用微生物间的拮抗作用,由枯草芽孢杆菌ＢＳ２２４活菌体制成的生物制品,喷涂于创面,达到防治各种致病菌感染的目的。按照国家卫生部申办新药的批件,由北京积水医院等四家单位对本制品进行临床研究。通过对２１６例病人的烧伤创面的细菌学校查,组织活检及主要脏器功能监测,结果表明,本制品对烧伤有良好的抗感染作用,用药后创面细菌培养阳性率显著下降,对铜绿假单胞菌更为有效,未发现本菌导致的侵袭性及脏器转移性感染,无毒副作用。     

固定化枯草杆菌生物吸附去除水中Cd的研究

采用明胶、琼脂、海藻酸钠作为载体对枯草杆菌进行固定,通过对三种载体的包埋效果、传质性能及操作难易的比较来选择适宜的固定化载体,比较固定化微生物与游离微生物及固定化载体海藻酸钠处理含镉废水的效果,并研究温度、pH值等环境因子对固定化枯草杆菌处理含镉废水效果的影响。结果表明：海藻酸钠作为固定化载体其传质性能强、方法简便,机械强度好;固定化枯草杆菌对含镉废水去除效果明显高于游离枯草杆菌。且随着废水中Cd浓度的变化,固定化枯草杆菌处理效果存在差异,在Cd浓度为1．0mg．L^-1 ～20mg．L^-1时,Cd的去除率在24h呈现3次曲线回归,而48h以4次多项式拟合;pH值对固定化枯草杆菌处理效果产生一定影响,在pH5．0-pH7．0,随pH值升高去除效率下降,温度在20℃-30℃,固定化枯草杆菌均有较好的处理效果。     

Comparative genome analysis of Bacillus cereus group genomes with Bacillus subtilis

Genome features of the Bacillus cereus group genomes (representative strains of Bacillus cereus, Bacillus anthracis and Bacillus thuringiensis sub spp. israelensis) were analyzed and compared with the Bacillus subtilis genome. A core set of 1381 protein families among the four Bacillus genomes, with an additional set of 933 families common to the B. cereus group, was identified. Differences in signal transduction pathways, membrane transporters, cell surface structures, cell wall, and S-layer proteins suggesting differences in their phenotype were identified. The B. cereus group has signal transduction systems including a tyrosine kinase related to two-component system histidine kinases from B. subtilis. A model for regulation of the stress responsive sigma factor sigmaB in the B. cereus group different from the well studied regulation in B. subtilis has been proposed. Despite a high degree of chromosomal synteny among these genomes, significant differences in cell wall and spore coat proteins that contribute to the survival and adaptation in specific hosts has been identified.     

Cloning in Bacillus subtilis of temperature-dependent promoter fragments from Bacillus stearothermophilus and Bacillus subtilis

Abstract Using promoter-probe plasmids, more than 200 promoter-containing fragments from Bacillus stearothermophilus and Bacillus subtilis were cloned in B. subtilis . Among these, 15 promoter fragments were highly temperature-dependent in activity compared to the promoter sequence (TTGAAA for the −35 region, TATAAT for the −10 region) of the amylase gene, amyT , from B. stearothermophilus . Some fragments exhibited higher promoter activities at elevated temperature (48°C), others showed higher activities at lower temperature (30°C). Active promoter fragments at higher and lower temperatures were obtained mainly from the thermophile ( B. stearothermophilus ) and the mesophile ( B. subtilis ), respectively. A promoter fragment active at high temperature was sequenced, and the feature of the putative promoter region was discussed.     

PspA外源表达对枯草芽孢杆菌168蛋白质分泌的影响

PspA同源物广泛存在于细菌和高等生物的组织中.在本研究中克隆了来源于地衣芽孢杆菌的PspA基因,并将其克隆于用于大肠-芽孢穿梭诱导表达载体pDG-StuI中构建重组质粒pDG-PspA.将构建的诱导表达型的重组质粒转化到Bacillus subtilis 168中,研究PspA的外源表达对该菌的生长,总蛋白分泌,以及Sec分泌途径中α-淀粉酶分泌的影响,结果表明,PspA基因的外源表达,在发酵过程后期能在一定程度上提高总蛋白的分泌量,在发酵过程后期能在一定程度上提高分泌的α-淀粉酶浓度.     

枯草芽孢杆菌HS-A38产抗菌脂肽Bacillomycin D的组分分析及鉴定

对一株枯草芽孢杆菌HS-A38产生的脂肽类物质进行分离鉴定及抑菌活性研究。通过酸沉淀分离和有机溶剂抽提的方法,从枯草芽孢杆菌HS-A38发酵液中得到脂肽粗提物LP,产率为1.956 g/L。利用薄层色谱和茚三酮染色法确定该脂肽粗提物中存在四个组分,分别为LP1、LP2、LP3和LP4;抑菌活性检测显示,组分LP3对两株海洋致病菌副溶血性弧菌和铜绿假单胞杆菌的活性较高。组分LP3经硅胶柱层析纯化分离后,应用反相高效液相色谱(RP-HPLC)和基质辅助激光解析电离飞行时间质谱(MALDI-TOF-MS)对该组分做进一步纯化和鉴定。分析表明,LP3样品在保留时间20 min~28 min产生单峰团LP3-1,其纯度为85.24%;经MALDI-TOF-MS分析和数据比对,组分LP3-1中的主要成分为杆菌霉素Bacillomycin D。     

枯草芽孢杆菌TF26抗菌蛋白抑菌活性的初步研究

目的:研究枯草芽孢杆菌TF26抗菌蛋白的抑菌活性和生物稳定性,为菌株及抗菌蛋白的应用提供理论依据.方法:采用硫酸铵盐析方法提取抗菌蛋白,采用菌丝生长速率法检测其对13种植物病原真菌的抑菌活性,采用抑菌圈方法对其生物稳定性进行分析.结果:抗菌蛋白粗提物能够抑制13种植物病原真菌的生长,平皿抑制率为74.3％ ～91.3％,对葵花菌核病菌、番茄和黄瓜枯萎病菌、黄瓜菌核病菌和立枯病菌、水稻恶苗病菌和大豆根腐病菌抑制作用较强.抗菌蛋白在100℃以下,pH＜ 10范围内抑菌活性稳定,对紫外线照射不敏感,室温(20℃)和4℃储存150d抑菌活性稳定.结论:抗菌蛋白具有较强的热、酸碱、紫外和储存稳定性以及广谱的抑菌活性.     

Potency of Bacillus thuringiensis and Bacillus subtilis against the cotton leafworm,Spodoptera littoralis (Bosid.) Larvae

The biological activities of two species of bacteria isolated from soil of cotton fields identified as Bacillus subtilis strain NRC313 (BS NRC313) and Bacillus thuringiensis strain NRC335 (BT NRC335) were evaluated against the third larval instar of the cotton leafworm, Spodoptera littoralis (Boisd.). The different entomopathogenic bacteria of BS NRC313and BT NRC335 contained 10 × 10⁸ cell/ml, and caused mortality of 100 and 97.3% for the above mentioned strains, respectively. Concentrations of 2.5 × 10⁸ to 10 × 10⁸ cell/ml of strains BS NRC313 and BT NRC335 were applied to the larvae: LC₅₀ were 3.3 × 10⁸ and 3.9 × 10⁸ cell/ml respectively. The influence of exposure to toxin concentrations manifested in terms of decreasing the adult emergence and prolongation of the generation period. The percentage of larvae that survived and succeeded to pupate increased by decreasing the concentration. The longevity of adult emergence that resulted from larvae treated with Bacillus subtilis were 6.0 ± 0.51 and 9.0 ± 0.63 days at 5 × 10⁸ and 2.5 × 10⁸ cell/ml, respectively compared with 9.8 ± 0.47 in control. The results indicated that Bacillus subtilis was more potent than Bacillus thuringiensis. Field applications of B. thuringiensis, B. subtilis and Reldan achieved 55.6, 67.4 and 89.4% reduction of the cotton leafworm larvae Spodoptera littoralis in clover plants under field conditions.     

The oxidative stress response in Bacillus subtilis

Abstract Bacillus subtilis undergoes a typical bacterial stress response when exposed to low concentrations (0.1 mM) of hydrogen peroxide. Protection is thereby induced against otherwise lethal, challenge concentrations (10 mM) of this oxidant and a number of proteins are induced including the scavenging enzymes, catalase and alkyl hydroperoxide reductase, and a putative DNA binding and protecting protein. Induced protection against higher concentrations (10–30 mM) of hydrogen peroxide is eliminated in a catalase-deficient mutant. Both RecA and Spo0A influence the basal but not the induced resistance to hydrogen peroxide. A regulatory mutation has been characterized that affects the inducible phenotype and is constitutively resistant to high concentrations of hydrogen peroxide. This mutant constitutively overexpresses the proteins induced by hydrogen peroxide in the wild-type. The resistance of spores to hydrogen peroxide is partly attributable to binding of small acid soluble proteins by the spore DNA and partly to a second step which coincides with the depletion of the NADH pool, which may inhibit the generation of hydroxyl radicals from hydrogen peroxide.     

The safety of Bacillus subtilis and Bacillus indicus as food probiotics

Aims:  To conduct in vitro and in vivo assessments of the safety of two species of Bacillus , one of which, Bacillus subtilis , is in current use as a food supplement.
Methods and Results:  Cultured cell lines, Caco-2, HEp-2 and the mucus-producing HT29-16E cell line, were used to evaluate adhesion, invasion and cytotoxicity. The Natto strain of B. subtilis was shown to be able to invade and lyse cells. Neither species was able to adhere significantly to any cell line. The Natto strain was also shown to form biofilms. No strain produced any of the known Bacillus enterotoxins. Disc-diffusion assays using a panel of antibiotics listed by the European Food Safety Authority (EFSA) showed that only Bacillus indicus carried resistance to clindamycin at a level above the minimum inhibitory concentration breakpoints set by the EFSA. In vivo assessments of acute and chronic dosing in guinea pigs and rabbits were made. No toxicity was observed in animals under these conditions.
Conclusions:  Bacillus indicus and B. subtilis should be considered safe for oral use although the resistance of B. indicus to clindamycin requires further study.
Significance and Impact of the Study:  The results support the use of B. subtilis and B. indicus strains as food supplements.