First detection of koi herpesvirus from koi,Cyprinus carpio L. experiencing mass mortalities in Iran: clinical,histopathological and molecular study

Koi herpesvirus (KHV) is the aetiological agent of an emerging disease (KHVD) associated with mass mortalities in koi and common carp and reported from at least 30 countries. We report the first detection of KHV from koi in Iran using clinical, histopathological and molecular studies. KHV‐infected fish showed reduced swimming activity, sunken eyes and increased mucus production on skin and fins. On post‐mortem examination, gill necrosis was observed in the majority of fish. Histopathologically, the gill showed diffuse necrosis of the branchial epithelial cells. Margination of chromatin was detected in gills, kidney, heart, spleen, intestine and brain. In addition, sequence analyses of the TK gene, ORF 136 and marker I and II, demonstrates that Iranian KHV isolates were identical and classified as variant A1 of TUSMT1 (J strain) and displayed the I⁺⁺II⁺ allele of this Asian genotype.     

猪伪狂犬病病毒山西株胸苷激酶基因序列分析

采用PCR方法对2011-2013年山西省分离的2株猪伪狂犬病病毒（pseudorabies virus, PRV）的胸苷激酶（TK）基因进行了克隆和测序,并将其基因序列和推导的氨基酸序列与国内外主要流行毒株Bartha株、Becker株、Ea株、Kaplan株、LA株、Min-A株、NIA-3株、SH株、SL株和Yangsan株进行了同源性分析。序列分析结果表明,核苷酸同源性分别为99.7%、99.6%、99.8%、99.7%、99.7%、94.4%、99.1%、57.2%、99.5%、99.7%;氨基酸同源性分别为99.1%、99.1%、99.7%、99.4%、99.4%、90.3%、98.1%、49.7%、98.8%、99.1%。另外在核苷酸序列中起始密码子上游有3段GC box的序列,终止密码子下游有多聚腺苷加尾信号AATAAA;TK基因均由320个氨基酸组成,氨基酸序列中有疱疹病毒TK基因共同的保守序列-R*Y*DG**G*GK*T-和-FDRHP*A***C*P*AR-。     

共表达AsiaⅠ型口蹄疫病毒P1-2A和3C基因重组山羊痘病毒的构建及鉴定

将口蹄疫病毒（FMDV）衣壳蛋白前体P1-2A基因和蛋白酶3C基因与启动绿色荧光蛋白双向串连痘苗病毒启动子相连,以KpnⅠ为插入位点,将整体基因片段插入山羊痘病毒转移载体骨架pUC119-TK之中,构建共表达重组山羊痘病毒转移载体pTK-P7.5-GFP-P12A3C。与疫苗弱毒株山羊痘病毒共转染BHK21细胞,经绿色荧光筛选,RT-PCR检测,成功获得能正确表达目的蛋白的重组山羊痘病毒株vTK-3CP1-GFP。     

Effect of halomethyl-1,3,5-triazines on nitrification of ammonia

Nitrification inhibitory activity of halomethyl-1,3,5-triazines was determined by measuring the inhibitory activities on ammonia-oxidation to nitrate (NO₃⁻-N) in an upland soil and on ammonia-oxidation to nitrite (NO₂⁻-N) by Nitrosomonas europaea ATCC 25978 (ATCC) and Nitrosomonas sp TK 794 (TK). Within the chlorinated trimethyl-1,3,5-triazines, those bearing at least one trichloromethyl group inhibited nitrification more strongly, both in soil and in cell suspension of ATCC, than other mono- or dichlorinated methyl-1,3,5-triazines. Introduction of an amino group to 2,4,6-tris(trichloromethyl)-1,3,5-triazine gave 10- and 100-fold increases of nitrification inhibitory activity in soil and ATCC cell culture, respectively. Within the trihalomethyl-1,3,5-triazines, those having tribromomethyl group(s) exhibited rather weaker nitrification inhibition in soil than the corresponding trichloromethyl-1,3,5-triazines, although they showed a strong inhibition in cell suspension. Ammonium oxidation in ATCC was inhibited more strongly than that in TK. In QSAR studies, the optimum log P values were calculated as c 4.30. By using this value it will become possible to design highly active trichloromethyl-1,3,5-triazine nitrification inhibitors.     

锦鲤疱疹病毒双基因检测方法的研究

为建立锦鲤疱疹病毒(KHV)双基因检测方法,本研究根据KHV聚合酶基因(Sph)和胸苷激酶基因(TK)的保守序列设计2对引物,建立两基因同时检测的PCR方法.结果表明,利用双基因同时检测KHV,可以同时特异扩增出570 bp和155 bp片段,而与鲤春病毒血症病毒、传染性造血器官坏死病毒和鲤鱼疱疹病毒无扩增反应.而且所建立的双基因检测方法对临床样品的检测结果与单一PCR方法比较符合率为100％.为进一步研究KHV以及诊断方法奠定了基础.     

Effect on Proliferation of Goat Pox Virus of TK Gene Deficiency

In this study, the proliferation of goat pox virus (GPV) was researched using the recombinant GPV of TK gene insertion inactivation. Different length DNA fragments X1, X2, X3 and X4 were inserted into the transfer plasmid pTKfpgigp to constructed the recombinant transfer plasmids and these plasmids were transfected into lamb testis cells infected GPV. Recombinant GPVs of TK gene inactivation by inserting foreign DNA fragment of 2 800, 4 000, 5 200 and 7 700 bp,respectively, were generated via homologous recombination. Four recombinant GPVs (rGPV/tk^-) were obtained by selective culture and the virus titer were mensurated by 50% tissue culture infective dose. The results showed that the rGPV/tk^- virus titers were decreased obviosuly which was comparable with the wild type GPV AV41. Furthermore, the virus titer of the recombinant virus was decreased more obviously with the extension of the foreign DNA fragment. The result indicated that the proliferation of the GPV was depressed when GPV TK gene was deficient.     

表达H5N1亚型禽流感病毒HA和NA基因的重组鸭瘟病毒的构建

利用PCR技术扩增出鸭瘟病毒(DPV)生长非必需的TK基因(约1.1kb),将其克隆入pGEM-Teasy载体获得载体pGTK。根据已知的绿色荧光蛋白载体pEGFP-C1的序列设计了一对引物,PCR扩增出pEGFP-C1上含CMV启动子、EGFP及其多克隆位点的完整的基因表达盒插入pGTK的TK上,获得质粒pGTK-EGFP。根据Genbank已发表的H5N1亚型禽流感病毒的血凝素(HA)和神经氨酸酶(NA)基因序列,设计了两对引物,分别从pT-HA和pT-NA两个质粒上扩增出HA和NA基因,克隆到pGTK-EGFP的表达盒的多克隆位点Kpn2I与SmaI之间,构建含EGFP及HA和NA基因的转移质粒载体pGTK-EGFP-HA-NA。将这质粒载体与DPV34F2疫苗毒共转染鸡胚成纤维细胞(CEF),通过荧光方法筛选,获得了表达HA和NA基因的重组DPV(rDPV-HA-NA)。     

我国微生物农药常见剂型种类及管理

本文总结了微生物农药的母药和常见剂型的定义、特性和登记现状,汇总了常用剂型产品质量控制的各项目和指标,分析了目前微生物农药剂型管理中存在的问题,并提出了建议,以推进微生物农药剂型科学管理和行业发展。     

鸡传染性喉气管炎病毒的分离和TK基因鉴定

从临床表现以呼吸困难、喉头出血为主要特征的某蛋鸡场患鸡喉头和气管等组织分离到一株病毒。该分离毒无血凝特性,应用检测ILTVTK基因的PCR能从分离毒中扩增出大小为427bp的特异性目的片段。测序结果与GenBank收录的ILTV不同毒株序列的同源性为100%。结果初步表明该分离毒为鸡传染性喉气管炎病毒(命名为ILTV-FJ)。