三黄鸡ST3Gal6基因的半定量RT-PCR及生物信息学分析

本研究旨在对三黄鸡ST3Gal6基因进行组织表达谱和生物信息学分析。参考三黄鸡ST3Gal6基因序列设计引物,采用PCR技术克隆三黄鸡ST3Gal6基因序列,并利用半定量RT-PCR进行组织表达谱分析;同时对其进行生物信息学分析。结果表明,克隆的三黄鸡ST3Gal6基因全长1169 bp,含有1059 bp的完整CDS编码区,编码352个氨基酸。其CDS编码区的核苷酸序列与人、黑猩猩、牛、大鼠、蟾ST3Gal6基因对应序列的同源性分别为62%、62%、61.9%、59%、54.4%。组织表达谱分析表明,ST3Gal6基因在各组织均不同程度地表达,其中在大脑表达量很高,肺脏中最低。生物信息学预测ST3Gal6蛋白结构发现,三黄鸡的ST3Gal6蛋白存在2个跨膜螺旋结构域,同时预测ST3Gal6存在22个磷酸化位点和1个特异性蛋白激酶磷酸化位点。     

流感病毒受体在三种动物气管和肺脏分布的组织化学检测

利用凝集素组织化学染色的方法,对岭南黄鸡、番鸭和BALB/C小鼠的气管和肺脏进行了流感病毒受体分布的检测。结果表明:在岭南黄鸡、番鸭和小鼠的气管粘膜层、粘膜下层、肺脏的细支气管和肺泡上皮细胞均有禽流感病毒受体的分布。番鸭和小鼠气管和肺脏的人流感病毒受体的分布范围和细胞类型与禽流感病毒受体的分布稍有差异,岭南黄鸡气管和肺脏未检测到人流感病毒受体的分布。研究结果为探讨流感病毒的感染机制和宿主特异性的差异提供了基础数据。     

Serological surveillance reveals widespread influenza A H7 and H9 subtypes among chicken flocks in Egypt

Multiple avian influenza viruses’ subtypes are circulating worldwide possessing serious threat to human populations and considered key contributors to the emergence of human influenza pandemics. This study aimed to identify the potential existence of H7 and H9 avian influenza infections circulating among chicken flocks in Egypt. Serum samples were collected from chicken flocks that experienced respiratory distresses and/or variable mortality rates. H7 and H9 virus infections were screened by haemagglutination inhibition assay using chicken erythrocytes. Serum samples were collected from 9 broiler, 12 breeder and 18 layer flocks. Out of 1,225 examined sera, 417 (34 %) from 14 flocks and 605 (49.4 %) from 21 flocks were found positive for H7 and H9, respectively. Prevalence of both H7 and H9 antibodies were higher in layer followed by breeder then broiler flocks. Special consideration should be paid to control influenza viruses in Egypt, as pandemic influenza strains may develop unnoticed given the presence of subclinical infections, and the possibility of re-assortment with the prevailing endemic H5N1 virus strains in Egypt do exist.     

禽流感病毒CH02株(H9)的鉴定及其NS 基因分子特征分析

从疑似感染H9亚型禽流感病毒的病鸡内脏组织中分离到1株能凝集鸡红细胞的病毒,通过血凝抑制试验、鸡胚中和试验、RT-PCR鉴定,确认其为H9亚型禽流感病毒,并命名为CH02株。该病毒HA效价为2^7.67±0.58;其血凝性可被抗H9亚型禽流感病毒阳性血清完全抑制,HI效价为7log2;鸡胚半数感染量(EID₅₀)为10^-8.68 EID₅₀/0.1 mL;最小致死剂量致鸡胚死亡的平均时间(MDT)为85.6 h;1日龄鸡脑内致病指数(ICPI)为0.625;其NS基因与香港株、南京株、北京株、汕头株、韩国株、巴基斯坦株的核苷酸同源性为88.6%~100%,经进化分析CH02株与香港株(A/duck/Hong Kong/Y280/97)同属一个分支。     

Replacement of berseem hay by Salix tetrasperma on physiological performance of New Zealand White rabbits under subtropical conditions of Egypt

Forty-eight growing New Zealand White male rabbits aged 6 weeks (874?±?1.3 g initial body weight (BW)) were used to study effects of partial replacement of berseem hay (BH) with Salix tetrasperma hay (ST) on growth and physiological responses. Rabbits were allotted to one of four diets of 12 rabbits each for 75 days in a completely randomized design. The treatments were as follows: control (30 % BH), ST25 (7.5 % ST?+?22.5 % BH), ST50 (15 % ST?+?15 % BH), ST75 (22.5 % ST?+?7.5 % BH). Nutrient digestibility coefficients, nutritive value and N utilization of rabbits fed with the ST50 rations were higher (P?P?P??ST25 and ST50?>?control. Glucose level was higher (P?P?P?相似文献   

Modeling the dynamics of backyard chicken flows in traditional trade networks in Thailand: implications for surveillance and control of avian influenza

In Southeast Asia, traditional poultry marketing chains have been threatened by epidemics caused by the highly pathogenic avian influenza H5N1 (HPAI H5N1) virus. In Thailand, the trade of live backyard chickens is based on the activities of traders buying chickens from villages and supplying urban markets with chicken meat. This study aims to quantify the flows of chickens traded during a 1-year period in a province of Thailand. A compartmental stochastic dynamic model was constructed to illustrate trade flows of live chickens from villages to slaughterhouses. Live poultry movements present important temporal variations with increased activities during the 15 days preceding the Chinese New Year and, to a lesser extent, other festivals (Qingming Festival, Thai New Year, Hungry Ghost Festival, and International New Year). The average distance of poultry movements ranges from 4 to 25 km, defining a spatial scale for the risk of avian influenza that spread through traditional poultry marketing chains. Some characteristics of traditional poultry networks in Thailand, such as overlapping chicken supply zones, may facilitate disease diffusion over longer distances through combined expansion and relocation processes. This information may be of use in tailoring avian influenza and other emerging infectious poultry disease surveillance and control programs provided that the cost-effectiveness of such scenarios is also evaluated in further studies.     

融合表达猪源C3d与猪流感病毒HA DNA疫苗的构建

为了构建融合表达分子佐剂猪补体C3d与猪流感病毒血凝素(HA)的真核表达载体,论文克隆了猪C3d全长基因与H3N2亚型猪流感病毒的HA基因,并构建了包含3拷贝的C3d和经过改造的HA(替换信号肽并去除跨膜区)的质粒pHA-C3d3.序列分析表明,获得的猪C3d与参考序列相比核苷酸同源性达到99.7%,氨基酸同源性达99...     

Consequence of Cryptosporidiosis on the immune response of vaccinated broiler chickens against Newcastle disease and/or avian influenza

The consequence of cryptosporidiosis on the immune response of vaccinated chickens against Newcastle disease and/or avian influenza was studied by using 240, 1 day old, male, white Hy-Line chicks and divided into 8 groups and subgroups. Each group or subgroup was consisting of 30 chicks (15?×?2 replicates). The first and second groups were kept as unvaccinated control, G1uninfected and G2 infected. G3, G4 and G5 contained 2 subgroups A&B (G3A, G3B, G4A, G4B, G5A and G5B). Chicks of subgroup A were vaccinated only while chicks of subgroup B were infected and vaccinated. These chicks were orally inoculated with 5?×?10⁵ oocysts of Cryptosporidium baileyi (C. baileyi) at 2 days of age. Chickens were vaccinated intraocular with live Newcastle disease (ND) vaccine (Hitchner on day 7th and LaSota on day 17th of chicken life) (G3) or vaccinated by subcutaneous route with Volvac®- H5N2- AI vaccine on day 10 of chicken life (G4). Last group (G5) was infected similarly and vaccinated with ND and AI vaccines with the same day, dose and route of vaccination for each one. Random blood samples were collected for 3 weeks post-vaccination for investigation of humoral immune response against Newcastle and/or avian influenza vaccines by the haemagglutination inhibition (HI) test. The results showed that H5N2 vaccine at day 10 of chicken life is effective in chickens indicated by the geometric mean of HI titer against AI virus. The findings of this study showed that the infection with Cryptosporidia in the broiler chicken has a depressive effect on the immune status of the birds vaccinated against ND and/or AI vaccination. Moreover, the obtained protection rates against challenge with virulent ND virus observed to be parallel to the results of HI- test. Also, by using 2 different antigens (one commercial and field prepared antigen) to avian influenza virus, lower Geometric mean (GM) HI titer were appeared in infected and vaccinated group than vaccinated group only. A study of the relative lymphoid organs weight such as bursa of Fabricius from the experimental chicks indicated that those organs were comparable between the groups infected-vaccinated and vaccinated only. Non significant variations in final live weight between uninfected control and infected groups were indicated. Also, H5N2-AI vaccination at 10 days old did not affect the final live weight. ND and/or AI Vaccination could not be a substitute to application of good hygienic measures and fecal examination of the birds especially for protozoal diseases such as cryptosporidiosis. It could be concluded that cryptosporidiosis could be one cause of ND and/or AI vaccination failure in poultry farms.     

鸭源H5N1亚型高致病性禽流感病毒的分离鉴定及其HA和NA基因的生物信息分析

分离到1株 H5N1亚型高致病性禽流感病毒, 经序列测定发现HA蛋白裂解位点上插入多个连续的碱性氨基酸（PQREIRRKKR*G）,从分子上证实是一株高致病性禽流感病毒。核酸序列比较分析结果表明,分离的流感病毒HA基因与A/duck/VietNam/Ncvd1/2002(H5N1)同源率最高,达到98.8%;NA基因与A/duck/VietNam/Ncvd1/2002(H5N1) 和A/chicken/Jiangsu/cz1/2002(H5N1)同源率最高,达到98.7%。氨基酸水平上,HA与A/duck/Viet Nam/Ncvd1/2002(H5N1)同源率最高,可达99.3%;NA与A/chicken/Jiangsu/cz1/2002(H5N1)同源率最高,达98.7%。HA与NA基因的潜在糖基化位点与作者所选参比毒株一致。通过遗传进化树分析结果表明,A/duck/VietNam/Ncvd1/2002(H5N1)可能是该毒株的来源株。     

H5N1亚型禽流感病毒和新城疫病毒感染鸡胚成纤维细胞IFN-β基因的动态表达

根据鸡β-干扰素（ChIFN-β）基因保守序列设计引物建立了检测鸡IFN-β mRNA表达水平的荧光定量RT-PCR (RRT-PCR)方法,并对H5N1亚型禽流感病毒（AIV）和新城疫病毒强毒株（vNDV）感染后3、6、12、24、30 h的鸡胚成纤维细胞（CEF）中IFN-β mRNA表达水平进行了检测。结果显示,建立的RRT-PCR特异性好,对鸡IFN-β mRNA的扩增效率为94.18%,线性范围为10^-8～10^-3,相关系数为0.992,最低能检出48拷贝/反应。AIV在感染CEF后6、12 h显著抑制IFN-β mRNA的表达,24 h开始诱导IFN-β mRNA表达,30 h时IFN-β mRNA水平显著升高;vNDV在感染CEF后的24 h内显著抑制IFN-β mRNA的表达,30 h时则显著诱导 IFN-β mRNA表达。本试验结果为进一步研究H5N1和NDV与机体的相互作用提供了有价值的信息。      

安徽省合肥地区家禽交易市场禽流感疫情定点监测情况分析

为了了解安徽省合肥地区家禽禽流感免疫及带毒情况,2011年4—8月,在合肥地区家禽交易市场,采集不同来源、不同品种家禽的血清和同份棉拭子样品（咽喉、泄殖腔双份）603份（实测589份）,分别进行高致病性禽流感免疫抗体和病原学检测,结果表明,全部样品平均免疫抗体合格率为70.80%。对监测结果进行分析发现,不同品种、不同养殖规模、不同月份的高致病性禽流感免疫抗体存在差异,其中,蛋鸡、种鸡高致病性禽流感免疫抗体合格率达80%以上,而肉鸡及水禽的免疫抗体水平则较差;饲养规模在1000羽以上的养禽场家禽高致病性禽流感平均免疫抗体合格率均达到70%以上,而饲养规模在1000羽以下的养禽场和农村散养家禽免疫抗体合格率分别为27.45%和32.84%,尚未达到农业部规定标准;在6月采集的样品,其平均免疫合格率偏低,其他月份差异不显著。病原学检测结果表明,全部检测样品高致病性禽流感病原检测结果均为阴性。     

H9N2亚型禽流感病毒的遗传进化分析及F株灭活疫苗的免疫效果评价

为了解H9N2亚型禽流感病毒(AIV)变异情况及评价作为H9亚型F株禽流感灭活疫苗的免疫效果,本研究对2009年～2010年期间从免疫鸡群中分离的9个H9N2亚型AIV株进行血凝素(HA)基因序列测定和分析。这9个病毒株HA基因编码区长度均为1 683 nt,HA基因核苷酸序列同源性为90.7%～98.9%,其推导氨基酸序列同源性为92.2%～98.8%;分离株与F株的HA基因之间的核苷酸序列同源性为91.6%～99.6%,氨基酸序列同源性为92.9%～99.3%;9个分离株与F株均属于Beijing/1/94-like进化分支,但分别属于3个不同的基因亚型。免疫试验结果显示:3周龄SPF鸡接种F株灭活疫苗21 d后,产生的HI抗体效价在log2 9以上;而且免疫鸡对分离株及F株攻毒后的喉头和泄殖腔排毒产生明显的抑制作用。本研究数据表明,F株灭活疫苗可以提供对这些分离株的有效免疫保护。     

Expression of adiponectin and its receptors (AdipoR1 and AdipoR2) in chicken ovary: potential role in ovarian steroidogenesis

Adiponectin and its receptors (AdipoR1 and AdipoR2) mRNAs are expressed in various chicken tissues including ovary. However, the cellular expression and the role of adiponectin system have never been investigated in chicken ovary. Here, we have shown that the level of adiponectin mRNA is about 10- to 30-fold higher (p < 0.001) in theca cells than in granulosa cells from each hierarchical yellow follicle studied (F4–F1). In contrast, the level of AdipoR1 mRNA expression was about two-fold lower in theca cells than in granulosa cells (p < 0.05) whereas those of AdipoR2 was similar in both ovarian cells. Whereas expression of adiponectin mRNA increased with follicular differentiation in theca cells, it decreased in granulosa cells. In contrast, mRNA expression of AdipoR1 and AdipoR2 in both theca and granulosa cells remained stable during yellow follicle development. To determine whether adiponectin is involved in the ovarian steroidogenesis, LH (100 ng/ml)-, FSH (100 ng/ml)- and IGF-1 (100 ng/ml)-induced progesterone production was measured in absence or presence of human recombinant adiponectin (10 μg/ml) for 36 h in cultured granulosa cells from F1, F2 and mixed F3 and F4 follicles. In absence of LH, FSH and IGF-1, adiponectin treatment had no effects on progesterone production whatever vitollegenic follicle studied. However, it increased by about two-fold IGF-1-induced progesterone secretion in F2 and F3/4 follicles whereas it halved progesterone production in response to gonadotropins (LH and FSH) in F3/4 follicles. Thus, in chicken, adiponectin, mainly expressed in theca cells, could exert paracrine or autocrine effect on the ovarian steroidogenesis.     

H9N2亚型禽流感病毒抗原性变异的研究

对1998—2002年间在河南省豫北地区分离到的5株H9N2亚型禽流感病毒的抗原性变异进行了研究。经HI试验、鸡胚中和试验、细胞中和试验及攻毒保护试验证明，5株H9N2亚型间已经发生了抗原性漂移。98A5和99S毒株间的保护力接近100％，HI试验、鸡胚中和试验、细胞中和试验的相关性均在0．74以上。表明2毒株间的抗原性相近；用00Y毒株攻击其他4株免疫的鸡，其保护率仅为60％～80％；而02Y株对除00Y株外的4株的免疫保护率分别为60％、75％、80％、100％，与分离年代呈负相关性，HI、鸡胚中和试验、细胞中和试验也取得类似结果，说明2000年后的毒株间已发生抗原性变异。     

Cloning of comparative gene identification-58 gene in avian species and investigation of its developmental and nutritional regulation in chicken adipose tissue

Adipose triglyceride lipase (ATGL) is the rate-limiting enzyme of lipolysis in chicken adipose tissue. Its regulation is not fully understood. Recent studies suggest ATGL may be regulated by physical protein-protein interactions. Comparative gene identification 58 (CGI-58) has been identified as an activator of ATGL in mice. The purpose of the current study was to clone and sequence the CGI-58 gene in avian species and to investigate its regulation during development, fasting, and refeeding. Here, we report the cloning and sequencing of the complete coding sequence of CGI-58 and the deduced AA sequences for the domestic chicken, turkey, and Coturnix quail. The CGI-58 protein is a 343-AA protein in the chicken and quail, and a 344-AA protein in the turkey. Sequence comparisons with the human and mouse show that the CGI-58 gene is highly conserved among avian and mammalian species, with complete identities at the predicted lipid-binding site. Cell fractionation of chicken fat cells and stromal-vascular cells revealed that CGI-58 is expressed primarily in mature adipocytes (P < 0.01). When compared in multiple organs and tissues, avian CGI-58 is expressed predominantly in the adipose tissue (P < 0.001), similar to ATGL. To understand CGI-58 expression during adipose tissue development, its mRNA expression was measured along with ATGL and stearoyl CoA desaturase (SCD-1) mRNA, an adipogenic marker, in embryos and adults. Messenger RNA expression of CGI-58 increased (P < 0.05) immediately after hatching, concurrent with peak ATGL expression. It is interesting that CGI-58 remained somewhat increased at posthatch d 11 and 33 as SCD-1 mRNA expression increased (P < 0.05). To evaluate the response of CGI-58 to nutritional status, chickens and quail were fasted for 24 h and subsequently refed. After the fasting period, CGI-58 mRNA was induced (P < 0.05) for both chickens and quail and was returned to control levels upon refeeding. The ATGL mRNA responded similarly, increasing dramatically after fasting and quickly decreasing with refeeding. The direct relationship between CGI-58 and ATGL mRNA expression indicates a role for CGI-58 in activating ATGL-mediated lipolysis in avian species.     

The susceptibility of culture cells to avian influenza viruses

The susceptibilities of culture cells to twelve avian influenza virus strains were determined with ten established cell lines including MDCK and ESK cells and three primary culture cells. The established cell lines derived from embryonic swine kidney (ESK) and chicken kidney (CK) primary culture cells were more sensitive to the avian influenza viruses than the other eleven cells. The ESK cell had a particularly higher infective titer than the MDCK cell with and without trypsin supplement in culture medium, and dispersion of the infective titers was narrower than that of the MDCK cell. The ESK cell is a suitable candidate for routine work on avian influenza viruses in laboratories.