Infection and distribution of Candidatus Liberibacter asiaticus in citrus plants and psyllid vectors at the cellular level

Huanglongbing (HLB) is currently considered the most destructive disease of citrus worldwide. In the major citrus-growing areas in Asia and the US, the major causal agent of HLB is the bacterial pathogen Candidatus Liberibacter asiaticus (CLas). CLas is vectored by the Asian citrus psyllid, Diaphorina citri, in a persistent propagative manner. CLas cannot be cultured in vitro because of its unclear growth factors, leading to uncertainty in the infection mechanism of CLas at the cellular level in citrus and in D. citri. To characterize the detailed infection of CLas in the host and vector, the incidence of HLB was first investigated in citrus-growing fields in Fujian Province, China. It was found that the positive association of the level of CLas infection in the leaves correlated with the symptoms. Then antibodies against peptides of the outer membrane protein (OMP) of CLas were prepared and tested. The antibodies OMP-225, OMP-333 and OMP724 showed specificity to citrus plants in western blot analyses, whereas the antibodies OMP-47 and OMP-225 displayed specificity to the D. citri vector. The application of OMP-225 in the immunofluorescence assay indicated that CLas was located in and distributed throughout the phloem sieve cells of the leaf midribs and axile placenta of the fruit. CLas also infected the epithelial cells and visceral muscles of the alimentary canal of D. citri. The application of OMP-333 in immunoelectron microscopy indicated the round or oval CLas in the sieve cells of leaf midribs and axile placenta of fruit as well as in the epithelial cells and reticular tissue of D. citri alimentary canal. These results provide a reliable means for HLB detection, and enlighten a strategy via neutralizing OMP to control HLB. These findings also provide insight for the further investigation on CLas infection and pathogenesis, as well as CLas–vector interaction.     

The effect of colchicine on cholesterol processing by the progesterone-producing cells of the luteinized ovary

Previously, we indicated that luteal cells from colchicine-treated superovulatory (luteinized) rats show decreased capacity for progesterone production. The current study investigates the possibility that colchicine exerts this effect by interfering with the mechanism by which cholesterol is processed and/or synthesized by luteal cells. To this end, animals were treated with saline or colchicine after which the luteinized ovary or isolated luteal cells were assayed for their cholesterol content, their ability to synthesize cholesterol endogenously, or their ability to utilize lipoprotein-delivered cholesterol for the production of progesterone. The results show that animals treated with colchicine show a number of changes in luteal cell cholesterol metabolism: namely a 60% decline in stored cholesterol, a 3-fold rise in the activity of the cholesterol synthesizing enzyme HMG CoA reductase (although no change occurs in other cholesterol metabolizing enzymes), and a 3-fold rise in the capacity of the cells to incorporate precursor [14C]acetate into cholesterol. On the other hand, cells of animals treated with colchicine or cells treated with colchicine under in vitro circumstances are unable to fully utilize cholesterol provided by high density lipoproteins (HDL): this occurs despite the fact that the binding of HDL particles to luteal cells is quite normal after colchicine treatment. These findings are consistent with the view that a primary effect of colchicine on luteal cell progesterone production is in preventing the normal uptake of HDL-cholesterol.     

三聚氰胺胶体金免疫层析试纸条的研制

通过胶体金免疫层析技术建立一种特异、便捷、快速的三聚氰胺抗原检测方法,对奶制品及饲料中的三聚氰胺残留水平监测提供参考。用柠檬酸三钠还原法制备的胶体金,标记纯化的三聚氰胺单克隆抗体,喷于试纸的金标垫。将MEL-OVA(三聚氰胺和卵清白蛋白的偶练物)和纯化的羊抗鼠IgG分别喷于试纸的T(检测线)处和C(质控线)处,通过挑选试纸条材料和调试工艺参数,并最终组装成试纸条。结果显示,制备的试纸监测体系方法检出限为50 g/L。试纸条对牛奶、奶粉和饲料中的三聚氰胺残留的检出限分别为100 g/L、100 ng/g和200 ng/g。将该法与LC-MS/MS法对比检测牛奶、奶粉和饲料样品,在试纸条检测范围内,与LC-MS/MS法检测结果一致性好,从而验证了该方法的有效性。制备的三聚氰胺胶体金检测试纸在常温干燥环境下至少可保质6个月,能够检测出三聚氰胺含量大于50 g/L的样品,适用于现场实际样品三聚氰胺残留水平监测,具有良好的应用前景。     

大熊猫精液和包皮菌群组成及潜在致病菌调查

[目的] 大熊猫是我国国家一级保护动物,其种群面临着传染病和栖息地破碎化等持续威胁,其中生殖系统的细菌感染和菌群失衡会影响大熊猫生殖健康,严重者可导致流产,是引起大熊猫繁殖障碍的原因之一。本研究对精液与包皮分泌物样本的菌群组成情况及分离培养潜在致病菌开展研究。[方法] 通过采集13份大熊猫包皮分泌物和12份精液样本,采用16S rRNA扩增子测序技术、细菌培养及PCR鉴定的方法,确定样本中的细菌种类。[结果] 菌群组成分析结果显示,在门水平上,厚壁菌门（Firmicutes）的细菌丰度在大熊猫包皮与精液中均为最高;在属水平上,不同时期的雄性大熊猫包皮的菌群可能会发生改变,棒状杆菌属（Corynebacterium）和Dolosicoccus是Ⅰ期包皮样本中最丰富的微生物菌群,相对丰度分别为15.45%和12.40%;链球菌属（Streptococcus）和埃希氏菌属（Escherichia）是Ⅱ期包皮样本中最丰富的微生物菌群,相对分度分别为37.94%和9.68%;拟杆菌属（Bacteroides）和普雷沃氏菌属（Prevotella）是精液样本中最丰富的微生物菌群,相对丰度分别为14.40%和12.88%。菌群多样性分析结果显示,精液样品高于Ⅰ期包皮样品和Ⅱ期包皮样品,Ⅰ期包皮样品和Ⅱ期包皮样品之间无显著差异。通过细菌分离培养得到肺炎克雷伯菌（Klebsiella pneumonia）在内的多种潜在性致病菌。[结论] 本研究分析了大熊猫精液和不同时期包皮分泌物的菌群组成,其优势菌属存在差异,大熊猫包皮与精液中存在潜在性致病菌,这可能对大熊猫的生殖系统健康带来威胁,其致病性有待进一步研究。     

AFLP analysis of genetic diversity and relationship among some Chinese domestic ducks and wild ducks

The amplified fragment length polymorphic(AFLP)technique was used to analyze the genome DNA polymorphism among 8 breeds of domestic ducks and 2 species of wild ducks.Nine of the 17 selected primers pairs gave reproducible polymorphic DNA amplification bands.The amplified bands ranged from 44 to 83 per primer pair.Of the 513 AFLP markers obtained.498 were polymorphic.The proportion of polymorphic loci was 97.1%.The genetic distance(D)and similarity coefficients(GS)were calculated based on the polymorphic data.Between domestic ducks D ranged from 0.331 to 0.589,while between domestic ducks and the wild ducks,it ranged from 0.298 to 0.520(vs.Anas Platyrhynchos)and from 0.316 to 0.522(vs.A.Poecilorhyncha),respectively.The variance analysis showed no significant difference between the two groups of data,which indicated that both mallard and spot-billed ducks made contributions to domestic duck evolution.A dendrogram was constructed according to the D value.     

蜘蛛杀虫肽基因的合成及其在植物中表达质粒的构建

近年来用生物制剂防治害虫,虽然可以避免环境污染,但效果往往不稳定,而用基因工程方法,将抗虫基因导入植物的基因组中,让植物自身产生抗虫物质,将是一种理想的途径［１,２］。抗虫的植物基因工程主要是利用苏云金杆菌的内毒素蛋白,转化此基因的烟草和番茄显示了对虫的抗性［３—６］。澳大利亚Ｄｅａｋｉｎ大学从一种蜘蛛毒液中分离纯化到一种只有３７个氨基酸的小肽,体外实验发现其能杀死多种对农业生产有害的昆虫,但对哺乳动物没有毒害作用［７］。我们根据此肽的氨基酸序列,采用植物偏爱的密码子,人工合成并克隆了此肽的基因…     

脑源性神经营养因子促进海马神经干细胞的存活、增殖及向神经元分化

探讨脑源性神经营养因子(brain derived neurotrophic factor, BDNF)对海马神经干细胞(neural progenitor/stem cells, NPCs)的存活、增殖及分化的影响.采用无血清培养基体外分离、纯化、扩增胎鼠海马NPCs.通过细胞形态观察、nestin免疫荧光染色及血清促分化检测NPCs的干细胞特性; 采用神经球计数及神经球直径测定观察BDNF对NPCs的促增殖作用, 筛选出在适当细胞密度下, 促进NPCs增殖的有效浓度; 采用Tunel染色及全自动生化分析仪测定细胞培养上清液乳酸脱氢酶(lactic dehydrogenase, LDH)的含量探讨BDNF对海马NPCs存活的影响; 采用抗-b-微管蛋白(tubulin) III (Tuj-1)染色检测NPCs分化成神经元的百分率, 同时测定分化神经元突起的长度.分离的海马NPCs表现为nestin 免疫染色阳性, 具有自我增殖能力、且能分化为神经元和星形胶质细胞; 当细胞密度为5×105个/ ml 时, 10～200 ng/ml BDNF能显著促进NPCs的增殖, 其中40 ng/ml BDNF促增殖作用最强, 40 ng/ml BDNF能显著增大神经球直径; 40 ng/ml BDNF 显著减少NPCs的凋亡率(Tunel /DAPI ), 抑制LDH漏出; 40 ng/ml BDNF能显著促进NPCs分化为Tuj-1免疫染色阳性神经元, 且分化后神经元的突起长度显著大于对照组.上述结果提示: BDNF促进海马NPCs的存活、增殖及向神经元方向分化.     

感染羊瘙痒因子263K株或139A株仓鼠脑组织中tau蛋白和磷酸化tau蛋白变化的研究

为探讨朊病毒病的神经病理特征,应用Western blot方法检测了感染羊瘙痒因子263K株或139A株的仓鼠脑组织中总tau蛋白和Ser396和Ser404位点发生磷酸化tau蛋白表达水平的变化;并应用Real Time PCR方法检测了tau mRNAs转录活性的改变。结果表明总tau蛋白含量升高而Ser396和Ser404位点发生磷酸化的tau蛋白含量降低,该现象与羊瘙痒因子毒株类型和临床潜伏期无关;感染羊瘙痒因子的仓鼠脑组织中Tau2和Tau4这两个异构体的转录水平升高。这些结果表明tau蛋白在Ser396和Ser404位点的去磷酸化可能与朊病毒病发病相关。     

过氧化物酶体增殖剂对稻瘟病菌生长发育及致病性的影响

【目的】探索过氧化物酶体增殖剂(Peroxisome proliferators,PPs)对稻瘟病菌生长发育及致病性的影响。【方法】在6种不同的PPs诱导下,观察比较稻瘟病菌过氧化物酶体数量及相关基因表达、生长速率、孢子萌发、附着胞形成与致病性的差异。【结果】在不同的PPs诱导下,稻瘟病菌过氧化物酶体数量均呈现明显的增加,同时过氧化物酶体形成相关基因PEX8、PEX11、PEX14的表达量升高;PPs影响病菌菌丝生长、分生孢子萌发及附着孢形成,并导致致病性的减弱。其中,2,4-D与阿司匹林的抑制效果最为显著。同时,2,4-D与ASA对稻瘟病菌过氧化物酶体形成突变体Δpex5和Δpex7的生长抑制效果与野生菌株相比明显增加。【结论】首次将PPs类化合物用于模式丝状病原真菌稻瘟病菌的研究。研究发现6种PPs均能够引起过氧化物酶体的增殖,并可抑制稻瘟病菌生长发育,降低致病性。     

鼎突多刺蚁群体结构和生活史的研究

本文报告了鼎突多刺蚁(Polyrhachis vicina Roger)的群体结构和生活史。通过研究表明,鼎突多刺蚁一年发生一代,以蚁后、雄蚁、工蚁、幼虫和卵越冬。卵和幼虫在冬天发育停滞(或极慢)。到了春天,随着气温的上升,卵和幼虫又恢复正常发育。据室内人工饲养观察,卵的发育历期为23.8±2.5天(平均温度26℃)幼虫为20.4±4.4天(26℃),工蚁蛹为19.8±5.5天(27℃)。成长工蚁在5—11月出现;8—11月,雄蚁从蛹中羽化;10月,雌蚁从蛹中羽化。雌蚁分飞交尾后,进入邻近蚁巢或回到原巢,脱翅成为蚁后。蚁巢内存在着多后现象。