赤眼鳟线粒体NADH脱氢酶6基因克隆及序列特征分析

[目的]根据赤眼鳟线粒体NADH脱氢酶6（ND6）基因序列分析不同种鱼类的分子进化关系。[方法]根据其他鱼类的线粒体ND6基因及侧翼序列设计引物,采用PCR扩增并克隆、测序赤眼鳟ND6基因,通过MEGA4．0Neighbor—Joining法分析不同鱼类的进化关系。[结果]赤眼鳟（Squaliobarbus curriculus）线粒体基因组ND6基因序列长度为522bp,编码173个氨基酸。该基因碱基组成为：A41．5％、C32．7％、G12．4％、T13．4％,其中A＋T含量（54．9％）高于G＋C含量（45．1％）。将赤眼鳟ND6基因序列与GenBank中其他11种鱼类ND6基因序列进行比对发现,赤眼鳟ND6核酸序列与团头鲂的同源性最高,为90．8％;与红点鲑鱼的同源性最低,为73．0％。赤眼鳟ND6氨基酸序列与其他物种ND6氨基酸进行比对发现,赤眼鳟与团头鲂同源性最高,为98．3％;与大黄鱼的同源性最低,为75．7％。以ND6氨基酸序列采用NJ法构建12种鱼类的系统发生树基本反映了它们的种间、属间和科间的关系。[结论]首次得到了赤眼鳟线粒体ND6基因序列,并确定了赤眼鳟与其他11种鱼类的分子进化关系。     

内生真菌球毛壳ND35的生物安全性评价

本试验初步研究了植物内生真菌球毛壳（Chaetomium globosum）ND35抗生物质粗提物对植物生长性状和植物根际土壤微生物的影响,对小白鼠、大白兔的毒性,子囊孢子在水中的存活及对鱼类的安全性。结果表明,球毛壳ND35抗生物质粗提物对黄瓜、菜豆和小麦的种子是安全的,并且能够显著增加须根数和根鲜重,促进种苗生长,对根际菌群数量无显著影响,对动物来说属低毒级;球毛壳ND35子囊孢子在水中能自行消解,且低于10^5cfu／ml时为泥鳅生长无显著影响;表明球毛壳菌ND35无毒、无污染,用于土传病害的生物防治无明显性风险。     

中草药复方制剂对肉鸡新城疫免疫影响研究

将210只1日龄的肉仔鸡随机分成7组,每组30只,在ND免疫过程中按每公斤体重0.2 g添加中草药复方制剂,Ⅰ、Ⅱ、Ⅲ组分别在免疫前5天、3天和免疫当天开始连续饲喂3天,Ⅳ、Ⅴ、Ⅵ组分别在免疫前5天、3天和免疫当天饲喂1天,对照组（VII组）不喂药,分别于免疫前1天和免疫后14天、21天、28天和40天翅下静脉采血检测ND抗体水平。结果表明,免疫后投药组抗体效价均高于对照组,抗体达到峰值时平均高出20.2-1.6个滴度,其中Ⅲ组与对照组差异显著（P＜0.05）,其他组与对照组差异不显著（P＞0.05）。     

内生菌螺旋毛壳抗生素和水解酶的协同抗真菌作用

 生防因子螺旋毛壳(Chaetomium spirale) ND35生长于含立枯丝核菌(Rhizoctonia solani)菌丝细胞壁制备物的SM培养基中,从培养滤液提取的粗酶液强烈抑制Valsa sordida、Valsa mali、Glomerella cingulata和Curvulavia lunata病原真菌的菌丝生长和孢子萌发。经DNS法检测,粗酶液同时具有β-1,3-葡聚糖酶(包括内切和外切酶)和几丁质酶活性,分别为0.19 U.mg-1和0.09 U.mg-1。生长于3%玉米粉浸渍液的螺旋毛壳ND35的培养滤液也能抑制上述病原菌的菌丝生长和孢子萌发。离体条件下检测了ND35产生的一纯化的具有分子量为73 kD的内切β-1,3-葡聚糖酶(GLUC73)和一纯化的抗生素对苹果炭疽病菌的分生孢子萌发和芽管延长的抑制效果。当内切β-1,3-葡聚糖酶使用浓度为180 μg·mL^-1时,抑制了测试真菌的孢子萌发,造成细胞壁的改变,导致了菌丝顶端的破裂;抗生素的有效抑菌中浓度ED₅₀为1.75 μg·mL^-1。单独应用时,1.0 μg·mL^-1的抗生素或40 μg·mL^-1的内切β-1,3-葡聚糖酶没有或仅有很低的抑菌效果;两者组合导致孢子萌发约78%受抑制。甚至10 μg·mL^-1的内切β-1,3-葡聚糖酶也使1.5 μg·mL^-1抗生素的抑菌作用从低于25%增加到超过50%的抑菌效果。此外,80 μg·mL^-1的内切β-1,3-葡聚糖酶使1.0 μg·mL^-1抗生素的抑菌活性从0%提高到90%。抗生素和β-1,3-葡聚糖酶的协同抗菌活性可能在内生菌螺旋毛壳的植病生防中起重要作用。     

VA5免疫增强剂对鸽新城疫病毒灭活疫苗的增效作用研究

试验旨在探究VA5免疫增强剂对鸽新城疫病毒（NDV）灭活疫苗（ND4416株）的免疫增效作用。选取160只健康鸽随机分为4组：ND4416株灭活疫苗组（NDV组）、ND4416株灭活疫苗与免疫增强剂（VA5）混合组（NDV+VA5组）、La Sota灭活疫苗组（La Sota组）及生理盐水空白对照组（C组）,进行免疫效力及免疫持续期试验。采用血凝抑制试验检测各组鸽免疫后不同时间点的血清抗体效价,结果显示,VA5免疫增强剂能够显著提高鸽NDV灭活疫苗血清抗体水平（P<0.05）。脾脏淋巴细胞转化试验结果显示,VA5能够有效刺激免疫鸽的淋巴细胞转化。在免疫后第30、90、180天,各组鸽子采用ND4416株进行攻毒试验,结果显示,NDV+VA5组在3个时间点的攻毒保护率均为100%,均高于La Sota组。免疫持续期试验结果显示,NDV+VA5组血清抗体水平在免疫后第21天达峰值11.20log₂,第180天仍能达到7.50log₂,攻毒保护率达100%,表明VA5免疫增强剂可延长鸽新城疫疫苗的免疫效力持续期至180 d。攻毒后体外排毒检测结果表明,VA5免疫增强剂能够缩短攻毒后鸽的体外排毒周期,减少体外排毒。综上,VA5免疫增强剂能够显著提高鸽NDV灭活疫苗的免疫效果,为研制效果更好的鸽NDV疫苗提供了基础依据,同时也为免疫增强剂的应用增加了试验数据。     

抗鸡新城疫和传染性法氏囊病二联卵黄抗体的研制及应用

用新城疫疫苗和传染性法氏囊病疫苗免疫健康高产蛋鸡制备抗鸡新城疫和传染性法氏囊病二联卵黄抗体,并在40个规模化肉鸡场做推广应用,结果表明,预防保护率达100%,治愈率为63%~85%。     

绿头鸭线粒体ND2基因全序列测定及其研究(英文)

[Objective] The study was to analyze the phylogenesis of Anas platyrhynchos. [Method] Complete sequence of mitochondrial ND2 gene of 4 Anas platyrhynchos was determined by direct DNA sequencing based on PCR products. Combined with ND2 gene sequences of the Anas Linnaeus accessed in GenBank, phylogenetic tree was constructed by Neighbor-joining and maximum parsimony methods. [Result] The ND2 gene sequences of 4 Anas platyrhynchos were identical(1 041 bp in length; the nucleotide contents of A, G, T, and C were 28.91%, 13.35%, 20.75% and 36.98% respectively; A+T content approximated to that of C+G). Sequences of ND2 gene of mallard were same as spotbill duck, and had high homology with others. The phylogenetic trees indicated mallard and spotbilled duck were close in genetic relationship, both shared a haplotype; then Philippine duck, green-winged teal and northern pintail fell into branch 'A". [Conclusion] The domestic duck may be domesticated from mallard and spotbilled duck.     

感染鸡传染性贫血病毒的雏鸡新城疫疫苗免疫后局部体液免疫球蛋白含量变化

本试验对感染鸡传染性贫血病毒（ＣＩＡＶ）雏鸡新城疫（ＮＤ）疫苗免疫及其强毒攻击后，其泪液、气管液、肠液和胆汁的免疫球蛋白ＩｇＧ、ＩｇＭ、ＩｇＡ含量的动态变化进行了检测。结果发现，感染雏鸡泪液的ＩｇＧ、ＩｇＡ和气管液的ＩｇＧ及胆汁的ＩｇＧ、ＩｇＭ、ＩｇＡ含量均于接种ＮＤ疫苗后７～２８天明显低于未感染ＣＩＡＶ的免疫对照雏鸡；泪液的ＩｇＭ和气管液的ＩｇＡ含量在ＮＤ免疫后１４～２８天明显降低；肠液的ＩｇＡ和ＩｇＭ、ＩｇＧ含量分别于免疫后７～１４和７、１４天也显著减少。表明ＣＩＡＶ感染雏鸡对ＮＤ疫苗免疫的局部体液免疫应答能力降低。ＮＤ强毒攻击后，ＣＩＡＶ感染ＮＤ免疫雏鸡泪液、气管液、肠液和胆汁的ＩｇＧ、ＩｇＡ、ＩｇＭ含量及免疫保护率，均明显低于未感染ＣＩＡＶ的免疫对照雏鸡。     

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.     

中药穿心莲对新城疫HI抗体滴度的影响

1日龄肉仔鸡随机分为A、B、C、D、E、F、G7个试验组,每组组内设2个重复小组,每个重复小组35只鸡。A、B、C、D、E、F试验组添加穿心莲,G组不添加。各组均对新城疫进行首免和二免。结果发现,穿心莲能显著提高肉鸡的新城疫HI抗体滴度水平。另外,穿心莲的抗体滴度水平有一定的剂量依赖性,随着剂量的上升效果有提高的趋势。