淡色库蚊肌动蛋白全长基因的克隆及生物信息学分析

为阐明肌动蛋白抗药性相关机制及研制新型卫生杀虫剂奠定基础,根据库蚊抗性与敏感品系差异表达的EST片段,设计特异扩增引物,运用RACE技术从淡色库蚊抗性品系中扩增出该抗性相关基因的全长cDNA序列,分析其生物信息学特性。结果表明,获得淡色库蚊肌动蛋白基因cDNA全长1 708bp序列,其编码377个氨基酸;该基因编码的蛋白为膜蛋白,具有27个跨膜螺旋、1个信号肽切割位点、27个磷酸化位点。     

河北邢台和江苏南京地区棉蚜对丁硫克百威和吡虫啉的抗性

【目的】对河北邢台和江苏南京地区棉蚜（Aphis gossypii Glover）进行抗性监测,了解棉蚜对丁硫克百威和吡虫啉的抗性发展情况,为抗性治理提供理论依据。【方法】以用药水平最低的新疆南疆种群作为相对敏感种群,通过室内毒力测定、离体酶活力测定、相对表达量分析,研究新疆南疆、河北邢台及江苏南京3个地区棉蚜对丁硫克百威和吡虫啉潜在的代谢抗性机制。【结果】河北邢台和江苏南京地区的棉蚜对丁硫克百威仍处于敏感阶段;但对吡虫啉的抗性已有所发展,抗性水平分别为4.87和8.72倍,为低水平抗性阶段。解毒代谢酶活力对比研究发现,3个地区田间种群之间多功能氧化酶、酯酶及谷胱甘肽转移酶的活力均存在显著差异,江苏南京种群的酯酶活力显著高于新疆南疆种群。定量PCR分析表明,河北邢台地区棉蚜的羧酸酯酶表达量较高,是新疆种群棉蚜的3.28倍。【结论】田间棉蚜已对吡虫啉产生了低水平的抗性,酯酶活力升高在棉蚜对吡虫啉低水平抗性形成中发挥作用。     

Sequencing of Culex quinquefasciatus establishes a platform for mosquito comparative genomics

Culex quinquefasciatus (the southern house mosquito) is an important mosquito vector of viruses such as West Nile virus and St. Louis encephalitis virus, as well as of nematodes that cause lymphatic filariasis. C. quinquefasciatus is one species within the Culex pipiens species complex and can be found throughout tropical and temperate climates of the world. The ability of C. quinquefasciatus to take blood meals from birds, livestock, and humans contributes to its ability to vector pathogens between species. Here, we describe the genomic sequence of C. quinquefasciatus: Its repertoire of 18,883 protein-coding genes is 22% larger than that of Aedes aegypti and 52% larger than that of Anopheles gambiae with multiple gene-family expansions, including olfactory and gustatory receptors, salivary gland genes, and genes associated with xenobiotic detoxification.     

Juvenile hormone induction of biting behavior in culex mosquitoes

Juvenile hormone deprivation caused by surgical removal of corpora allata shortly after adult emergence blocked the initiation of biting behavior in Culex. pipiens and Culex quinquefasciatus. Reimplantation of corpora allata or injection of a synthetic juvenile hormone (JH-I) corrected the juvenile hormone deficiency and restored biting behavior. Ovariectomy experiments demonstrated that this behavioral effect of juvenile hormone was independent of ovarian involvement.     

Environmental Control of Ovarian Development in Mosquitoes of the Culex pipiens Complex

Gonotrophic dissociation, a condition in which the ovaries remain undeveloped in female mosquitoes that have taken a full blood meal, occurs in Culex pipiens L., when incubated at low temperature (10 degrees to 15 degrees C) with short photoperiod and held at low temperature after feeding. Gonotrophic dissociation occurred sporadically in Culex quinquefasciatus Say after conditioning by low temperature, irrespective of photoperiod. Two major considerations are posed: first, the importance of gonotrophic dissociation to hibernation potential; and, second, the potential of a hibernating female mosquito to serve as a virus reservoir.     

抗马拉硫磷褐飞虱品系的交互抗性及其生化分析

用马拉硫磷筛选的褐飞虱抗性品系(R)对马拉硫磷的抗性达到184．4倍，对作用于乙酰胆碱酯酶的二嗪磷、异丙威和仲丁威有一定的交互抗性，而对氰戊菊酯不表现显的交互抗性。进一步研究显示：在抗性品系中，磷酸三苯酯对马拉硫磷和仲丁威的增效作用分别达到9．15和2．38倍，顺丁烯二酸二乙酯对这两个药剂也有一定的增效作用(2．32和1．42倍)。抗性品系的酯酶、谷胱甘肽-S-转移酶(以2，4二硝基氯苯为底物)和多功能氧化酶活力分别为敏感品系(S)的14．67、1．52和1．11倍。在抗性品系中，磷酸三苯酯对酯酶活力抑制作用很强，顺丁烯二酸二乙酯对谷胱甘肽-S-转移酶活力也存在一定的抑制作用。实验结果显示，酯酶和谷胱甘肽-S-转移酶在抗性品系对马拉硫磷抗性和对其他药剂的交互抗性中起到不同程度的重要作用。     

Isolation and characterization of larvicidal principle of garlic

The larvicidal principles of garlic, Allium sativum L.,have been isolated and identified as diallyl disulfide and diallyl trisulfide. Both natural and synthetic samples of these larvicides are fatal at 5 parts per million to Culex pipiens quinquefasciatus Say.     

家蚕核酸糖转运蛋白BmSlc35b3基因的克隆与序列分析

在电子克隆的基础上,利用RT-PCR方法克隆了家蚕的同源基因BmSlc35b3(GenBank登录号:GU244352)。生物信息学分析结果显示,该基因编码区长1128bp,包含5个外显子,由基因推定的蛋白质由375个氨基酸组成,内有9个跨膜结构域,8个N-糖基化位点,与家蚕溶质运载蛋白家族B3(ABD36159)有4个氨基酸的差异,与赤拟谷道(EFA00972)、金小蜂(XP_001601459)、致倦库蚊(EDS30138)、疟蚊(EAA11308)等的同源性均在70%以上。RT-PCR对BmSlc35b3基因在五龄3d家蚕幼虫9种组织中的表达进行检测,结果表明,其在中肠和精巢中表达,在其他组织中不表达。     

2011～2014年华东四地区口岸蚊类监测报告

[目的]掌握华东四地区口岸蚊类种群构成、分布、密度、季节消长和携带病原体情况,为蚊类的口岸防治提供基础数据.[方法]采用CO2诱蚊器、集卵器法、捞勺法、目测法对华东四地区口岸蚊类进行调查,对捕获蚊类进行分类鉴定和病原体检测.[结果]2011～2014年本底共捕获蚊类102 583只,隶属于1科6属18种,以致倦库蚊/淡色库蚊为优势种.5～10月均可捕获蚊类,以6月为高峰者最多.集卵器法阳性率为9.41％,孳生地阳性率为7.23％.入境截获蚊类数量较少,以淡色库蚊为主.登革热、疟疾等病原体检测均为阴性.[结论]华东四地区口岸以致倦库蚊/淡色库蚊为优势种,6月为高峰,口岸存在蚊类孳生环境,应在今后的防控工作中作为重点对象.     

Suppression and elimination of an island population of Culex pipiens quinquefasciatus with sterile males

The release of 8,400 to 18,000 males per day of Culex pipiens quinquefasciatus Say which had been exposed to a sterilizing agent (thiotepa), suppressed and eliminated an indigenous population of this mosquito on an island off the coast of Florida in a 10-week period. Sterile males were effective in seeking out and mating with females on the island.     

Dieldrin susceptibility: partial restoration in Anopheles selected with a carbamate

Selection of dieldrin-resistant Anopheles albimanus Wied. and normal Culex pipiens quinquefasciatus Say with m-isopropylphenyl methylcarbamate for 21 and 30 generations, respectively, resulted only in vigor tolerance to this and other aryl methylcarbamates. However, it caused substantial restoration of dieldrin- and DDT-susceptibility in Anopheles by increasing the frequency of homozygous susceptible phenotypes from 10 to 83 percent.     

氟化物对家蚕血液羧酸酯酶及全酯酶活性的影响

为了探讨氟化物对家蚕代谢机制的影响,以家蚕耐氟品种T6和氟化物敏感品种734为研究对象,从5龄起蚕开始分别添食50、100、200、400mg/kg NaF溶液浸泡后的新鲜桑叶,检测家蚕血液中羧酸酯酶(CarE),全酯酶活性的变化.结果表明,734、T6添氟组的CarE活性分别是对照组的73％-88％和72％-81％,734两个低浓度添氟组的CarE活性与对照组和两个高浓度添氟组的差异极显著(P＜0.01),T6各处理组之间的差异不显著.734、T6添氟组的全酯酶活性分别是对照组的89％-97％和73％-92％,734各处理组之间的差异不显著,T6对照组的全酯酶活性仅与最高浓度添氟组差异极显著(P＜0.01).说明氟化物对家蚕血液CarE和全酯酶活性具有一定的抑制作用.     

棉铃虫对氰戊菊酯的抗性机理研究

以羧酸酯酶抑制剂磷酸三苯酯（ＴＰＰ）和多功能氧化酶抑制剂增效醚（ＰＢＯ）以及￣（１４）Ｃ－氰戊菊酯为材料，选择对氰戊菊酯具有不同抗性程度的棉铃虫自然种群为试虫，研究其抗性机理。结果表明：棉铃虫对氰戊菊酯的抗性与其体内酯酶活性无关，而与多功能氧化酶活性直接相关；同时，表皮对氰戊菊酯的渗透性降低也是棉铃虫对氰戊菊酯产生抗性的强化因子。     

不同酯酶型球孢白僵菌株系及相同酯酶型株系经寄主转移后毒力变化的研究

对12个酯酶型的16株白僵菌进行了毒力生物测定.结果表明,不同酯酶型株系对松毛虫的毒力差异显著,LT50可相差数倍;同一酯酶型株系经不同寄主转移后,其毒力也发生了较明显的变化.如从松灰象甲上分离出的不同酯酶型的2株白僵菌[MGS980404-21(酯酶型05),MGS980510-06(酯酶型06)]和人工释放的酯酶型02菌株Bb13毒力都很高,而从松灰象甲分离出的与释放菌株属同一酯酶型的另一个菌株MGS980507-34,毒力虽然很低,但却是林间最常见的流行菌株.上述结果理解为白僵菌在林间的宿存,尤其是它在松毛虫低虫口状态下能在林间长期宿存不绝的原因,以及对于从林间采集筛选高毒菌株提供了可能.     

Erythromycin Resistance of Streptococcus suis Isolates In Vivo

In order to study erythromycin resistance of Streptococcus suis under high or low concentration of selective drug pressure, Streptococcus suis strain LN was isolated from a diseased pig in 2005 and showed to be susceptible to erythromycin as determined by disc diffusion and tube dilution tests. In this study, clean level rabbits were divided into three groups of six rabbits each, including a prevention dosage group, a treatment dosage group, and a control group. After injection with S. suis strain LN, erythromycin （20 μg mL^-1） was taken orally in the prevention dosage group, erythromycin （5 mg kg^-1） was injected intramuscularly in the treatment dosage group, and no treatment was given in the control group. S. suis with intermediate resistance to erythromycin was isolated on the 5th day after infection from the prevention dosage group （5th PDG） and on the 7th day after infection from the treatment dosage group （7th TDG）. Both isolates were determined to be the constitutive macrolide-lincosamide-streptogramin B （cMLSB） resistance phenotype. The resistance gene ermB was detected in all of the isolates. The results suggested that both the 5th PDG and 7th TDG isolates had a mutation （A2372T） in the 23S rRNA gene. In addition, the 5th PDG isolates had a mutation in ribosomal protein L4 （detected as G268A） and a mutation in ribosomal protein L22 （A345C）; and the 7th TDG isolates had a C insertion at site 564. Each of these mutations is considered as a possible mechanism of erythromycin resistance in S. suis strain LN. This study demonstrated that erythromycin resistance was readily induced in S. suis at a low erythromycin dose creating selective pressure in vivo. Resistance appeared to be mediated by ribosome methylation, encoded by the ermB gene.     

斜纹夜蛾溴氰菊酯抗性和敏感品系酯酶的生化特性研究

测定了斜纹夜蛾酯酶的动力学参数,酯酶同工酶电泳和杀虫剂(溴氰菊酯和丙溴磷)对酯酶的抑制效应。结果表明,抗性品系与敏感品系酯酶的Km值之间均没有显著差异,但抗性品系酯酶的Vmax值比敏感品系的高且差异显著。在酯酶的非变性聚丙烯酰胺凝胶电泳图谱中,抗性品系中所含的酯酶同工酶条带数比敏感品系的多;溴氰菊酯浓度在12.5μmol/L时,对敏感品系酯酶的抑制率最大,约为30%,而对抗性品系酯酶的抑制率则随着溴氰菊酯浓度的升高而升高,溴氰菊酯浓度在50μmol/L时抑制率最大,约为75%;但是丙溴磷对抗、感品系的酯酶的抑制效应较为一致。上述结果说明在抗性品系中出现了与敏感品系不同的酯酶同工酶,但这些同工酶并没有显著改变抗性品系全酯酶的Km值,仅是增强了其酯酶活性,提高了对溴氰菊酯的代谢能力。     

宠物源大肠杆菌中质粒介导喹诺酮类耐药基因qnrD的研究

 【目的】对临床分离的宠物源大肠杆菌进行质粒介导喹诺酮类耐药基因qnrD的检测,并对该基因进行序列分析和传播机制的研究。【方法】采用微量稀释法对阳性菌株进行15种抗生素的最小抑菌浓度试验;通过PCR对基因克隆,并对克隆产物和菌株阳性质粒进行转化;同时对菌株进行接合转移试验。【结果】从164株宠物源大肠杆菌中检测出1株qnrD阳性菌株(GP2009-036),GP2009-036对14种兽医临床常用抗生素耐药严重,表现为多重耐药(14耐);PCR产物经连接PMD19-T载体后可转化入DH5α感受态细胞中;接合转移试验成功地将质粒转移到大肠杆菌J53中;可从GP2009-036与其转化子中抽提出阳性质粒。【结论】该qnrD阳性菌株对临床常用抗生素耐药严重,且阳性质粒可在病原微生物间进行水平传播,其水平传播机制可能使该基因在宠物临床上进行传播。     

田间灰飞虱抗药性相关细胞色素P450基因和酯酶基因的筛选

利用毒死蜱、吡虫啉和溴氰菊酯等3类不同药剂,通过毒力生测首先确定了田间灰飞虱对杀虫剂的低水平多重抗性特性;其次通过解毒酶活力测定,发现抗性品系的细胞色素P450氧化酶和酯酶明显升高;进而根据灰飞虱转录组数据进行引物设计,通过PCR克隆对转录组中的细胞色素P450及酯酶基因进行验证,在得到53条细胞色素P450基因序列及72条酯酶基因序列的基础上,利用半定量PCR比较这些基因在灰飞虱田间抗性品系和敏感品系间的表达差异。结果发现:细胞色素P450的CYP6家族中有10条,CYP4家族中有6条,这16条基因序列在抗性品系中的表达量明显高于敏感品系;酯酶中也有12条基因系列在抗性品系中的表达量明显高于敏感品系,它们中3条来自羧酸酯酶家族,8条来自磷酸酯酶家族,1条来自其他酯酶家族。提示:解毒酶基因的过量表达是灰飞虱田间低水平多重抗性的主要机制,也是抗药性发展初期的关键因素,利用分子生物学技术进行田间监测,有利于了解田间灰飞虱的抗药性发展。