Toxicity of Bacillus thuringiensis Crystal Toxins to Field Populations of Rice Leaf Folder, Cnaphalocrocis medinalis （Guenee） and Establishment of Baseline Susceptibility to Cry1Ab

Eight insecticidal crystal proteins of Bacillus thuringiensis, CrylAa, CrylAb, CrylAc, CrylB, Cry2Aa, CrylC, CrylDa and Cry 1Ea were assessed for toxicity against 1 st instar larvae of rice leaf folder, Cnaphalocrocis medinalis （Guenee） at 48 HAT and 72 HAT. Bioassay results depicted CrylAa was the most toxic （LCso 2.35 ppm） followed by CrylBa （LCso 8,50 ppm） and CrylAb （LCso 8.73 ppm） at 48 HAT, whereas, at 72 HAT CrylAb proved to be highly toxic （LC50 0.50 ppm） followed by CrylAa （LCso 4.07 ppm）, CrylAc （LCso 4,84 ppm） and CrylBa （LCso 6.42 ppm）. Toxins Cry2Aa, CrylCa, CrylDa and CrylEa did not resulted in any mortality at 48 HAT and 72 HAT, respectively. Baseline estimates for CrylAb against 1st instar larvae of C. medinalis sampled from seven geographical locations revealed variation in LC50＇s from 0.37 ppm to LC50 16.25 ppm at 48 HAT and LC50 0.50 ppm to LC50 6.49 ppm 72 HAT, respectively with relative resistance ratios of 44-fold and 13-fold at 48 HAT and 72 HAT over the susceptible population.     

Research Lactobacillus rhamnosusFermented Herbal and Bacillus subtilis Complex on Immune Performance,E.coli Infection of Broiler

This experiment was designed to study the complex of Lactobacillus rhamnosus fermented herbal and Bacillus subtilis on White Feather broiler immunity performance and impact of Escherichia coli infection.360 one-day-old broiler chickens were randomly divided into 3 groups with 4 replicates in each group and 30 chickens per replicate.The pretrial period lasted for 7 d,and the experiment lasted for 35 d.The chickens in the group Ⅰ(positive control group) and group Ⅱ(negative control group)were all only fed a basal diet,group Ⅲ was test group,by additive 1% fermented herbal preparations,groups Ⅱ and Ⅲ were intraperitoneal injection of 1 mL E.coli at 35 d,broiler mortality,immune organ index,cecalmicroflora content,immunoglobulin levels,IL-2 and IL-6 content were tested.The results showed that injection of E.coli caused massive death of chickens,group Ⅱtook up to 75.00%,it was significantly higher than groups Ⅰ and Ⅲ (P < 0.05),the mortality in group Ⅲ was significantly lower than group Ⅱ(P < 0.05),was only 23.33%.Injection of E.coli maked spleen index and thymus index of group Ⅱincreased significantly (P < 0.05),the spleen index and thymus index of groups Ⅰ and Ⅲ were no significant difference (P > 0.05).E.coli counts was significantly decreased after injectionin group Ⅲ (P < 0.05),but the number of intestinal Lactobacilli of group Ⅲ was significantly increased (P < 0.05),and inhibited the propagation of E.coli,the counts of E.coli in groups Ⅰ and Ⅲ were no significant difference (P > 0.05).At 42 d,the sIgA of the intestinal fluid in group Ⅲ were higher than that of groups Ⅰ and Ⅱ with 11.99% and 36.56%,respectively(P < 0.05).The serum IgG concentrations of group Ⅲ was higher than that of groupsⅠand Ⅱwith 14.68% and 28.15%,respectively(P < 0.05).At 42 d,the IL-2 content of group Ⅱ was the lowest,it was significantly lower than group Ⅲ(P < 0.05),the IL-6 of group Ⅲ was significantly lower than group Ⅱ(P < 0.05).     

紫外辐照对产α-ALDC枯草芽孢杆菌的诱变效应

采用紫外诱变(波长260nm，功率15w)对一株产α-乙酰乳酸脱羧酶的枯草芽孢杆菌（Bacillus subtilis）BS059菌株进行诱变处理。结果表明，在紫外辐照时间为50s时，诱变效果较好。从正突变菌株中反复筛选，得到两株产酶量较高的菌株BS059-15和BS059-22，比原出发菌株的酶活分别提高了107.62%和162.25%。经过连续传代试验，证明其遗传性状稳定，为可遗传变异。     

利用抗性基因替换的方法将两个拷贝外源基因表达单元整合到枯草芽孢杆菌染色体基因组同一位点

本研究提供了一种新的整合多拷贝外源基因表达单元到枯草芽孢杆菌染色体同一位点的方法。以β-淀粉酶表达单元作为应用实例,本方法包括以下步骤：首先,构建含有一个拷贝β-淀粉酶表达单元的整合质粒pMLK83-CTBA,通过同源双交换获得单拷贝β-淀粉酶表达单元整合到枯草芽孢杆菌1A751染色体α-淀粉酶基因位点的菌株1A751[CTBA]Neo＋;然后,利用抗性基因替换质粒pVK71,将新霉素抗性基因替换为状观霉素抗性基因,得到1A751[CTBA]Neo-Spe＋;最后,整合质粒pMLK83-CTBA再以同源单交换方式整合到1A751[CTBA]Neo-Spe＋染色体,通过新霉素抗性和状观霉素抗性筛选出两个拷贝β-淀粉酶基因整合的重组菌1A751[CTBA2]Neo＋Spe＋。结果显示,利用此方法增加β-淀粉酶基因的拷贝数,能够显著和稳定地提高β-淀粉酶的表达量。     

苏云金芽孢杆菌营养期杀虫蛋白VIP3研究进展

营养期杀虫蛋白（vegetative insectcidal protein,VIP）是由苏云金芽孢杆菌（Bacillus thuringiensis,Bt）在营养生长指数中期至稳定期期间分泌产生的一类新型杀虫因子,分为VIP1、VIP2和VIP3三种,以VIP3的研究最为深入。VIP3A对鳞翅目害虫具广谱杀虫活性,可作用于敏感昆虫中肠上皮细胞刷状缘膜囊致离子通道的形成,杀虫活性达纳克级水平。利用Bt ICPs不同启动子与vip3基因重组可提高VIP3蛋白的表达量和稳定性。转vip3基因植物也已有构建成功的报道。本文从VIP3的类型和杀虫活性、作用机理、vip3基因的定位和分离、vip3基因重组和转vip3基因植物等方面详细介绍了VIP3近十年来的研究进展。     

河道淤泥和堆肥蛭石混合发酵制备基质及其育苗效果

农村河道清淤产生的淤泥,体量大、有机物浓度高,处置不当会造成二次污染。现代农业的工厂化育苗需求大量的营养土,就地取土导致耕地退化。该研究利用功能微生物发酵淤泥制备育苗基质,研究不同菌株发酵基质的物理和生物学性状,基质培育西瓜苗的生长、生理参数和抗逆性能。结果表明:微生物处理均能够提升淤泥基质物理和生物学性能,同时能够提升育苗质量。其中Trichoderma harzianum T83(T83)、Bacillus amyloliquefaciens IAE(BIAE)菌株发酵基质性能最好。相较于对照处理基质的最大持水量、总孔隙度、毛管孔隙度、通气孔隙度,T83处理分别增加了64.25%、52.65%、45.05%、56.11%;BIAE处理分别增加了101.17%、45.43%、61.43%、38.14%。相较于对照处理西瓜苗的株高、鲜质量、干质量、叶绿素含量、根系活力、根际真菌、细菌数量,T83处理分别增加了66.85%、52.07%、72.16%、43.13%、54.93%、110.62倍、1.63倍;BIAE处理分别增加了80.40%、57.34%、82.37%、54.88%、46.40%、67.26%、2.60倍、2.94倍。T83和BIAE处理西瓜苗叶片过氧化氢酶和超氧化物歧化酶酶活显著增加,根系丙二醛含量显著降低。真菌菌株T. harzianum T83和细菌菌株B. amyloliquefaciens IAE发酵淤泥,能够显著提升其农用品质,为淤泥高附加值化农用提供一条可行的途径。     

拮抗细菌A2菌株的鉴定及其对小麦白粉病的防治效果

为适应现代无公害农业的发展,针对小麦白粉病的防治,从保护地土壤中分离得到一株生防细菌A2。通过形态学特征、生理生化特性的观察和测定,结合16S rDNA序列分析并构建系统发育树,对菌株A2进行了初步鉴定。同时,以清水为对照,测定了菌株A2对3432品种小麦白粉病的防治效果。结果显示该菌株主要特征是菌体杆状,内生芽孢、芽孢卵圆形,革兰氏染色阳性,好氧,接触酶反应、乙酰甲基甲醇实验为阳性,能利用柠檬酸盐;该菌株与亲缘关系较近菌株B.subtilis isolate G8的同源性达99%。结合以上两点,将菌株A2初步鉴定为枯草芽孢杆菌（B. subtilis）。防效实验显示菌株A2发酵液处理最好防治效果为72.22%;无菌滤液处理最好防治效果为29.24%,与对照相比均具有显著性差异（α=0.05）。表明,拮抗菌株A2菌体与代谢产物对小麦白粉病都具有较强的防治作用,为研制型、无毒、无公害生物农药提供了依据。     

Phosphate-solubilizing microorganisms associated with the rhizosphere of mangroves in a semiarid coastal lagoon

 The phosphate-solubilizing potential of the rhizosphere microbial community in mangroves was demonstrated when culture media supplemented with insoluble, tribasic calcium phosphate, and incubated with roots of black (Avicennia germinans L.) and white [Laguncularia racemosa (L.) Gaertn.] mangrove became transparent after a few days of incubation. Thirteen phosphate-solubilizing bacterial strains were isolated from the rhizosphere of both species of mangroves: Bacillus amyloliquefaciens, Bacillus licheniformis, Bacillus atrophaeus, Paenibacillus macerans, Vibrio proteolyticus, Xanthobacter agilis, Enterobacter aerogenes, Enterobacter taylorae, Enterobacter asburiae, Kluyvera cryocrescens, Pseudomonas stutzeri, and Chryseomonas luteola. One bacterial isolate could not be identified. The rhizosphere of black mangroves also yielded the fungus Aspergillus niger. The phosphate-solubilizing activity of the isolates was first qualitatively evaluated by the formation of halos (clear zones) around the colonies growing on solid medium containing tribasic calcium phosphate as a sole phosphorus source. Spectrophotometric quantification of phosphate solubilization showed that all bacterial species and A. niger solubilized insoluble phosphate well in a liquid medium, and that V. proteolyticus was the most active solubilizing species among the bacteria. Gas chromatographic analyses of cell-free spent culture medium from the various bacteria demonstrated the presence of 11 identified, and several unidentified, volatile and nonvolatile organic acids. Those most commonly produced by different species were lactic, succinic, isovaleric, isobutyric, and acetic acids. Most of the bacterial species produced more than one organic acid whereas A. niger produced only succinic acid. We propose the production of organic acids by these mangrove rhizosphere microorganisms as a possible mechanism involved in the solubilization of insoluble calcium phosphate. Received: 21 April 1999     

胨冻样芽孢杆菌(Bacillus mucilaginosus)的分离、筛选及其发酵培养基研究

本文报道了胨冻样芽孢杆菌 (Bacillusmucilaginosus)的分离、筛选及其对小麦苗期生长的作用 ,并对胨冻样芽孢杆菌的发酵培养基进行了优化 .结果表明 ,胨冻样芽孢杆菌 (B .mucilaginosus)生长的最适培养基 (g/L)为 :蔗糖 1 4、Na2 HPO4 1 .5、FeCl3 0 .0 0 2、CaCO3 1 .5、酵母膏 1 0 ,自来水 1 0 0 0ml、pH7.0～ 7.5;有机氮和无机铵态氮抑制胨冻样芽孢杆菌的生长 ;胨冻样芽孢杆菌液体培养物对小麦苗期生长具有促进作用 .