甜菜BvGSTU9基因的生物信息学及在镉胁迫下的表达分析

为了进一步研究甜菜谷胱甘肽转移酶BvGSTU9 (LOC104894060)在重金属胁迫过程中的功能。本研究以‘780016B/12优’为实验材料,对该基因序列特征、结构、功能进行预测分析,并利用qPCR检测该基因在不同浓度镉胁迫下的表达量变化。结果显示甜菜BvGSTU9基因全长925 bp,开放阅读框675 bp,编码了由224个氨基酸组成的不稳定膜外蛋白。BvGSTU9与菠菜、藜麦的氨基酸序列相似性较高,与系统发育进化树分析结果基本相符。二级和三级结构预测表明该基因主要由α-螺旋、β-折叠、延伸链及无规则卷曲组成。qPCR显示BvGSTU9基因在不同浓度的镉胁迫下均受到不同程度的诱导,因此可以推断甜菜BvGSTU9基因无论从结构还是功能上,与镉逆境胁迫存在着一定的应答关系。研究结果也为甜菜耐重金属镉机制研究提供参考依据。     

瓜类褪绿黄化病毒及其系统进化分析

【目的】检测新疆南疆7个县/市瓜类褪绿黄化病毒(Cucurbit chlorotic yellows virus, CCYV),进行系统发育分析,为新疆南疆CCYV预警和防控提供参考。【方法】于2019年,从中国新疆南疆7个县/市采集带有黄化特征的51份甜瓜叶片样品,RT-PCR进行CCYV的检测,进行特异性片段的克隆、测序和系统发育分析。【结果】中国新疆巴楚县、阿克苏市、莎车县、伽师县、疏勒县、疏附县均未检测到CCYV,而洛浦县的13份样本中,8份检出为CCYV阳性,检出率为61.53%,洛浦县CCYV特异性片段克隆测序获得了685 bp的核酸序列,与GenBank中的CCYV分离物外壳蛋白(Coat protein, CP)的一致性达到100%。所得序列与中国新疆(吐鲁番)、中国其他省、苏丹、日本、塞浦路斯、黎巴嫩的CP基因聚在I组(Group),沙特阿拉伯的分离物聚在Ⅱ组,伊朗的分离物聚在Ⅲ组。【结论】CCYV在中国新疆南疆洛浦县甜瓜产区发生,与中国新疆吐鲁番、中国其他省及周围国家的CCYV分离物亲缘关系很近,且群体遗传变异很小。     

Characterization of bovine ileal epithelial cell line for lectin binding,susceptibility to enteric pathogens,and TLR mediated immune responses

In this study, primary and immortalized bovine intestinal epithelial cells (BIECs) were characterized for the expression of surface carbohydrate moieties. Primary BIEC-c4 cells showed staining greater than 90 % for 16 lectins but less than 50 % staining for four lectins. Immortalized BIECs showed significantly different lectin binding profile for few lectins compared to BIEC-c4 cells. BIEC-c4 cells were studied for infectivity to E. coli, Salmonella enterica, bovine rotavirus, bovine coronavirus, and bovine viral diarrhea virus. Bovine strain E. coli B41 adhered to BIEC-c4 cells and Salmonella strains S. Dublin and S. Mbandaka showed strong cell invasion. BIEC-c4 cells were susceptible to bovine rotavirus. LPS stimulation upregulated IL-10, IL-8, and IL-6 expression and Poly I:C upregulated TLR 8 and TLR 9 expression. This study provides important knowledge on the glycoconjugate expression profile of primary and immortalized BIECs and infectivity and immune responses of primary BIECs to bacterial and viral pathogens or ligands.     

寨卡病毒SYBR GreenⅠreal-time PCR方法的建立及应用

为建立一种针对寨卡病毒的快速诊断方法,本研究根据寨卡病毒的3’端保守基因序列,设计合成1对引物,建立了检测寨卡病毒的荧光定量PCR方法。结果显示:所建立的检测方法的Ct值与标准品在1.41×10^1~1.41×10^10^ copies/μL具有良好的线性关系,相关性为1,斜率为-3.502;灵敏性结果显示,该方法的检测限度为1.41×10^1 copies/μL,是普通PCR的10000倍;特异性结果显示,对CHIKV、DENV和JEV无特异性扩增,特异性强;重复性试验结果显示,组内和组间变异系数均小于1%,重复性好。本研究建立的SYBR Green I real-time PCR检测方法,可用于寨卡病毒感染的快速诊断。     

Protective efficacy of an H5/H7 trivalent inactivated vaccine produced from Re-11, Re-12, and H7-Re2 strains against challenge with different H5 and H7 viruses in chickens

We developed an H5/H7 trivalent inactivated vaccine by using Re-11, Re-12, and H7-Re2 vaccine seed viruses, which were generated by reverse genetics and derived their HA genes from A/duck/Guizhou/S4184/2017(H5 N6)(DK/GZ/S4184/17)(a clade 2.3.4.4 d virus), A/chicken/Liaoning/SD007/2017(H5 N1)(CK/LN/SD007/17)(a clade 2.3.2.1 d virus), and A/chicken/Guangxi/SD098/2017(H7 N9)(CK/GX/SD098/17), respectively. The protective efficacy of this novel vaccine and that of the recently used H5/H7 bivalent inactivated vaccine against different H5 and H7 N9 viruses was evaluated in chickens. We found that the H5/H7 bivalent vaccine provided solid protection against the H7 N9 virus CK/GX/SD098/17, but only 50–60% protection against different H5 viruses. In contrast, the novel H5/H7 trivalent vaccine provided complete protection against the H5 and H7 viruses tested. Our study underscores the importance of timely updating of vaccines for avian influenza control.     

1株H1N1亚型猪流感病毒的全基因组特征及其对小鼠的致病性

旨在了解河南省猪流感病毒的流行情况及其遗传进化和基因组特征。2018年4月，从河南省某一出现疑似流感症状猪群中采集鼻拭子样品150份用于分离病毒，对分离病毒的全基因组进行序列测定和分析。同时感染6周龄BALB/c小鼠，研究其对小鼠的致病性。结果显示，获得1株H1N1亚型病毒[命名为A/swine/Henan/NY20/2018（H1N1）]。遗传进化表明，其HA和NA基因属于欧亚类禽H1N1分支，PB2、PB1、PA、NP和M基因属于2009甲型H1N1分支，NS基因属于经典H1N1分支。HA蛋白的裂解位点序列为PSIQSR↓GL，具有低致病性流感病毒的分子特征，在小鼠肺和鼻甲有效复制并能引起肺组织病理学变化。本研究分离到1株3源重排H1N1亚型病毒，对小鼠呈现一定致病力，提示应进一步加强对SIV的监测。     

Identification of an H1N1 subtype of swine influenza virus and serological analysis

To investigate the epizootic of swine influenza virus(SIV), 60 nasal swabs were collected from a clinical cases of pig farm in Tai'an City, Shandong Province of China in April 2017. SIV was isolated by inoculating into 10-day-old Special Pathogen Free embryonated eggs and the whole genome was sequenced. An H1N1 subtype SIV was isolated and designated as A/swine/Shandong/TA04/2017(H1N1). Phylogenetic analysis showed that apart from the polymerase A(PA) fragment belonging to the 2009 pandemic H1N1 branch, seven genome segments belonged to avian-like H1N1 influenza virus lineage. The cleavage site sequence of the hemagglutinin(HA) protein was PSIQSR↓G, which is a typical molecular biological characteristic. Five potential N-glycosylation sites(N14, N26, N277, N484 and N543) were found in the HA gene. To further investigate the epidemiology of SIV in this farm, the 995 serum samples were assessed with EAH1N1 2009 pandemic H1N1 and H3 N2 antigens. The results showed that the total positive rate was 65.43%. The positive rates of single virus infection detected by EAH1N1, 2009 pdmH1N1 and H3 N2 for serum HI(Hemagglutination inhibition) were 48.35, 30.85 and 7.47%, respectively. The results showed that SIV in Shandong Province has been reassorted in some segments and the SIV-positive rate was high on the SIV outbreak farm. These data provide evidence of an epizootic of SIV.     

CRISPR/cas系统及其在家禽上的应用研究进展

CRISPR/cas是一种获得性免疫防御系统,广泛存在于细菌和古细菌中。Strep tococcus pyogenes cas9(Spcas9)作为目前研究最多、最清楚的效应蛋白,因其酶活性高和靶点广而被应用于生命科学各个领域,但Spcas9分子量大、脱靶率高等缺点在一定程度上限制了其应用。随着CRISPR系统技术被深入开发,一些新效应蛋白被发现,本文就CRISPR/cas系统分类、原理、新效应蛋白发现及cas9(S.Pyogenes)多种变构体技术开发在及家禽上的应用进行综述,为相关领域的研究提供参考。     

番茄褪绿病毒在湖南省首次发生

2016年7月在湖南省蔬菜病害调查中发现,4种茄科蔬菜表现出叶脉间褪绿、叶片黄化等症状,疑似被番茄褪绿病毒(Tomato chlorosis virus,To CV)感染,同时叶片背面聚集了大量烟粉虱。采用To CV热激蛋白(heat shock protein 70,HSP70)序列的特异性引物对采集的番茄、茄子、辣椒、马铃薯样品和烟粉虱样品进行RT-PCR检测,均扩增出目标条带,且扩增序列与北京番茄To CV分离物(KC887999.1)部分序列相似度为99.0%,确认采集样品被To CV感染。4种茄科蔬菜To CV的感染率达70%~100%;发病叶片上烟粉虱的带毒率为66.7%~87.5%;鉴定出烟粉虱的生物类型为MED烟粉虱。这是湖南省首次确认该病毒,需要引起关注和加强防范。     

宁南霉素等药剂对番茄黄化曲叶病毒病的田间防效

[目的]评价宁南霉素、艾德拉和碧护3种药剂混合使用防治番茄黄花曲叶病毒病的效果。[方法]通过田间试验研究8%宁南霉素、艾德拉绿(花、果)与碧护组成的药剂对番茄黄化曲叶病毒病的防治效果。[结果]8%宁南霉素、艾德拉绿(花、果)与碧护组成的试验药剂对番茄黄化曲叶病毒病具有很好的田间防治效果,第3次施药后10、20、30 d防效均在82%以上,与对照药剂番茄黄化曲叶病毒灵差异达极显著水平,且其产量比清水对照增加25 747.5 kg/hm~2,保产率为103.90%。[结论]宁南霉素、艾德拉和碧护混合使用对番茄黄花曲叶病毒病具有较好的防治效果,为番茄黄花曲叶病毒病的有效防治提供了理论依据。