椰心叶甲的天敌——椰心叶甲啮小蜂在田间扩散距离测定

采用软皮尺实地丈量,使用OP3000水准仪测量和使用地球定位仪———Gps72测定椰心叶甲的天敌———椰心叶甲啮小蜂在田间释放后的扩散距离及向上扩散的高度。结果表明:椰心叶甲啮小蜂在田间有一定的向周边扩散能力,在田间释放后20d,啮小蜂可向周边扩散10~50m。7个月后,最远可扩散6 km。扩散距离与风向有关,东北风较多情况下,向西南扩散最远距离可达6060 m,向东北方向扩散仅仅890 m。啮小蜂在田间扩散呈椭圆形状向各方向扩散,顺风面扩散距离远、范围较大。     

Study on Synergistic Effect of VA5 Immunopotentiator on Pigeon NDV Inactivated Vaccine

This study was designed to evaluate the synergistic effect of VA5 immunopotentiator on pigeon ND4416 inactivated vaccine.160 healthy pigeons were randomly divided into four groups,including ND4416 strain inactivated vaccine group (NDV group),ND4416 strain inactivated vaccine and immunopotentiator (VA5) mixed group (NDV+VA5 group),La Sota inactivated vaccine group (La Sota group) and normal saline as the control group (C group),to assess their vaccine efficacy against virulent pigeon NDV by serological analysis and animal testing.Pigeons sera were collected at different time points after immunization and measured the HI antibody titer of each group.The results showed that VA5 immunopotentiator significantly improved the serum antibody level of pigeons immunized with pigeon Newcastle disease inactivated vaccine (P<0.05).In addition,comparative test of spleen lymphocyte transformation was conducted at various time points after immunization.The results indicated that VA5 effectively stimulated the lymphocyte transformation of immune pigeon.Pigeons in each groups were challenged with ND4416 strain at the 30th,90th and 180th d after immunization.The results presented that the NDV+VA5 group had 100% protection rate and higher than La Sota group.The duration of immunization test showed that the antibody titer of NDV+VA5 group reached peak 11.20log₂ at the 21st d,remained 7.50log₂ at 180th d,and the protection rate remained 100% at 180th d.It indicated that VA5 immunopotentiator sustained the immune duration of pigeon NDV vaccine up to 180 d.Moreover,the in vitro detoxification test results suggested that VA5 immunopotentiator reduced the in vitro detoxification cycle of pigeons after challenge.Overall,this study suggested VA5 immunopotentiator could significantly improve the immune efficacy of pigeon Newcastle disease inactivated vaccine,which provided a basis for further enhancing the efficacy of Newcastle disease vaccine,as well increased experimental data for the application of immunopotentiators.     

Research Progress of Canine Coronavirus and Feline Coronavirus

Canine coronavirus (CCoV) and feline coronavirus (FCoV) belong to α-genus coronavirus of coronavirus family,porcine transmissible gastroenteritis virus (TGEV),porcine epidemic diarrhea virus (PEDV) also belong to the same genus.Genetic evolution analysis showed that different genotype of the virus could produce new variant strains through gene recombination,which caused great obstacles to the diagnosis and control of the disease.β-genus coronaviruses include bovine coronavirus (BCoV),canine respiratory coronavirus (CRCoV) and severe acute respiratory syndrome coronavirus (SARS-CoV).Among them,CRCoV has the highest homology with BCoV,but there are great differences in genomic structure,pathogenic mechanism and infection symptoms between this kind of coronavirus and α-coronavirus.CCoV and FCoV are widely spreading around the world,characterized by high morbidity and low mortality.Due to the characteristics of RNA virus and the influence of environmental selection pressure,the viruses continue to mutate and evolve,and new virulent strains appear one after another.The virulence of feline infectious peritonitis virus (FIPV) is greatly enhanced,some specific point mutations in the virus genome change the cellular tropism against the host.The pathogenesis of the virus mainly depends on the antibody-dependent enhancement (ADE) induced by virus infection.The epidemiological investigation and prevention and control of CCoV and FCoV should not only rely on the single factor of vaccine immunity,but also comprehensively consider the virulence of the virus,environmental conditions,pet self-immune resistance, and so on.The identification of CCoV and FCoV should be based on clinical symptoms,combined with routine hematological examination,serum biochemical examination and laboratory diagnosis techniques to prevent false positive and false negative results.     

紫花苜蓿根腐病活体整株接种试验

对19个苜蓿材料进行根腐病活体整株接种试验,结果表明:19个供试品种中有4个为免疫,7个为高抗,5个为中抗,3个为中感材料;从苜蓿秋眠数来看,秋眠数高的品种相应其抗病性强,而秋眠数低的品种易感染根腐病。     

3种不同精粗比日粮条件下空怀小尾寒羊母羊的消化与代谢

本试验选取18只体重约42 kg的空怀小尾寒羊母羊,按体重分为3组(n=6),分别喂给以玉米秸秆为粗饲料的精粗比为20∶80(日粮1)、30∶70(日粮2)和40∶60(日粮3)的日粮,以研究不同精粗比日粮对空怀小尾寒羊母羊消化代谢的影响。结果表明,3组羊只的有机物采食量分别为970.1±95.1、1137.3±144.8和1279.2±138.7 g/d.只;玉米秸秆的采食量分别为834.3±75.6、856.1±110.2和822.0±121.7 g/d.只;有机物表观消化率分别为52.53%±5.16%、55.73%±2.64%和59.68%±1.95%;粗蛋白质表观消化率分别为58.73%±7.60%、61.05%±2.61%和62.84%±2.33%;纤维素表观消化率分别为57.42%±6.04%、56.12%±1.06%和53.42%±3.83%;氮保留量分别为0.62±1.17、3.15±2.22和5.87±1.56 g/d.只;钙保留量分别为0.39±0.05、0.59±0.10和0.79±0.06 g/d.只;磷保留量分别为0.40±0.04、0.59±0.06和0.82±0.05 g/d.只;所获得的可消化能分别为9.40±1.37、12.31±2.05和15.09±1.83 M J/d.只;试验期体增重分别为-12.4±46.1、101.2±60.0和157.1±96.5 g/d.只。由此得出结论,在以玉米秸秆为粗饲料的条件下,当空怀小尾寒羊母羊日粮中的精料量为20%时,体重难以维持,并且氮保留率也低(3.29%);增加日粮中的精料量到30%或40%时,可提高日增重,并且使氮保留率达到13.82%或22.09%。增加日粮的精料比例可不同程度地提高有机物、能量、粗蛋白质和钙磷的表观消化率,但对纤维素类物质的消化有一定影响。增加动物的可消化能量供应可大幅度改善氮的利用率,也增加钙磷的保留量。提高日粮的精粗料比例,并不影响小尾寒羊对玉米秸秆的采食量。     

徐州地区鸭传染性浆膜炎的病原分离鉴定及药敏试验

从徐州地区临床疑似鸭传染性浆膜炎发病鸭中分离到细菌21株，通过细菌形态、培养特怔、生化试验和动物试验，鉴定为鸭疫里默氏杆菌菌株，对不同地区分离到的21株菌进行了24种常用抗菌药物的药敏试验，结果该24个菌株对头孢类药物、红霉素、链霉素、妥布霉素、新霉素、氯霉素、泰乐菌素、痢特灵等几种药物敏感性较高，对喹诺酮类、强力霉素、丁安卡那霉素、林肯霉素、庆大霉素、氟苯尼考、利福平、复方新诺明等均不同程度地产生了耐药性。不同地区分离到的鸭疫里默氏杆菌对不同抗菌药的敏感程度存在较大差异性，研究结果为该地区鸭传染性浆膜炎的药物防治提供了很好的理论依据.     

鸡胚胎原始生殖细胞的体外培养

旨在对影响鸡胚胎原始生殖细胞（PGCs）体外培养、传代过程的各种影响因素进行探讨。将分离到的19期鸡胚胎生殖腺的PGCs在以鸡胚成纤维细胞（CEF）为饲养层，添加细胞因子的培养体系中进行培养、传代。经冷冻保存的PGCs在添加细胞因子的培养体系中能增殖，并具有分化能力。从细胞形态、集落形态、增殖生长能力、核型检测、碱性磷酸酶测定等方面对培养过程中的鸡胚胎PGCs进行检测。结果表明，采用传至3代的CEF为饲养层，多次离散法进行传代所获得的5代鸡胚胎PGCs不但能有效维持PGCs的未分化状态和正常二倍体核型，也具有其作为多能性胚胎干细胞的一系列特征。     

牛γ-干扰素在昆虫细胞中的高效表达

应用RT-PCR技术从经植物血凝素(PHA)刺激诱导的奶牛脾脏淋巴细胞总RNA中扩增出牛-γ干扰素基因(bovine interferon-γ,BovIFN-γ)cDNA,并克隆到pGEM-T easy载体中,经过限制性酶切分析和测序证实,所克隆到的基因编码区序列与已报道的序列完全一致.将含信号肽的BoIFN-γ基因整个编码区cDNA亚克隆到杆状病毒转座载体pFastBac Ⅰ中,构建了转移载体pFastBac 1-BoIFN-γ,转座到宿主菌DH10 Bac中,在含庆大霉素、四环素、卡那霉素、IPTG和X-gal的KGTIX LB平板上筛选白色菌落,提取DNA获得重组穿梭载体Bacmid-BoIFN-γ质粒,与脂质体共转染Sf9细胞,产生有感染力的重组杆状病毒reBac-BoIFN-γ.重组病毒经过传代扩增感染Sf9细胞,通过IFA试验、Western-blot和抗病毒活性测定证实,BoIFN-γ基因在感染的昆虫细胞和上清中得到了表达,上清中活性可达2.651×105U/mL.表达条件优化结果表明,不同MOI对rBoIFN-γ的表达产量影响不大,上清中干扰素活性在感染后5 d达到高峰.     

柔嫩艾美耳球虫表面抗原SAG2基因的克隆与表达

根据生物信息学预测的基因序列设计引物,应用RT-PCR方法从柔嫩艾美耳球虫第二代裂殖子总RNA扩增获得了鸡柔嫩艾美耳球虫表面抗原2(surface antigen 2,SAG2)基因序列,将其与pGEM-T easy载体连接后转化E.coliDH5α,筛选阳性克隆,以带有限制酶切位点的特异性引物用PCR方法扩增不含SAG2 N端信号肽序列的ORF序列后克隆至表达载体pET-32 a(+),构建了重组表达质粒pET-32 a(+)-SAG2,并将其转化至E.coliBL21(DE3)。经IPTG诱导,获得了SAG2重组抗原在大肠杆菌的高效表达,重组蛋白的表达量约占菌体总蛋白的35%,融合蛋白的分子量约为47 ku。菌体经超声处理后进行SDS-PAGE分析表明,表达蛋白以包涵体的形式存在。     

基于颠茄发根的外源基因表达系统的建立

 用携带植物高效表达载体pCAMBIA1304 ⁺ “解除武装”的重组C58C1工程菌转化颠茄无菌苗真叶, 发根诱导率达100%。PCR检测证实发根型质粒pRiA4和表达型质粒pCAMBIA1304 ⁺均能整合到颠茄发根基因组内, 共转化率达30%。建立了基于颠茄发根的高效外源基因表达系统, 为将托品烷类生物碱东莨菪碱的生物合成关键酶基因导入颠茄, 实现其次生代谢工程及开展分子育种奠定了基础。