一例鸡马立克氏病与沙门氏菌病混合感染的实验室诊断

为确诊思南县某养殖专业合作社饲养的肉鸡发病死亡原因,采集病鸡羽髓、血样及病死鸡肝脏、脾脏进行细菌分离培养、相关病毒琼脂扩散试验和PCR检测。结果:肝脏组织中分离出沙门氏菌;羽髓琼脂扩散试验检测出马立克氏病抗原,血清琼脂扩散试验检测出马立克氏病抗体;肝脏、脾脏PCR检测出马立克氏病病毒核酸。结论:综合诊断为马立克氏病与沙门氏菌病混合感染。     

应用荧光PCR诊断鸡腺胃型传染性支气管炎

应用荧光PCR试剂盒对临床表现为消瘦和腺胃肿大为特征的患鸡病料进行鸡传染性支气管炎病毒(IBV)、禽网状内皮组织增殖病病毒(REV)和鸡马立克氏病病毒(MDV)核酸检测。结果显示:鸡传染性支气管炎病毒荧光RT-PCR检测为阳性,禽网状内皮组织增殖病病毒荧光RT-PCR检测为阴性,鸡马立克氏病病毒荧光PCR检测为阴性。结合患鸡精神沉郁,极度消瘦,腺胃肿大如球状等的临床表现,综合分析表明该患鸡发病主要病因为感染鸡腺胃型传染性支气管炎病毒。     

超强马立克氏病病毒的鉴定及病毒Meq基因序列的比较

对1例感染超强马立克氏病病毒的病例进行确诊,对感染病毒的Meq基因进行比较分析。采用病理解剖、PCR检测、病毒分离、动物试验和Meq基因序列分析,对感染病毒进行研究。病理解剖结果为病死鸡的肝脏、脾脏、腺胃、肌胃、十二指肠表现为肿瘤病变。PCR检测结果为病死鸡的组织病料感染马立克氏病病毒。病毒分离和动物试验结果证明该感染病毒是1株马立克氏病超强毒株,该病毒可以引起免疫过CVI988疫苗的鸡发病。Meq基因序列分析表明该病毒与7株马立克氏病病毒参考毒株的同源性为98.8%~99.6%,该病毒在Meq的第115、119和176位氨基酸突变同国内流行株,该检测病毒在Meq的第217位氨基酸突变同超超强马立克氏病毒株。结果表明,通过病理解剖、PCR检测、基因序列分析、病毒分离和动物试验,确诊病鸡感染超强马立克氏病病毒。     

一例鸡马立克氏病病毒和禽白血病病毒混合感染病例的实验室诊断与分析

2020年11月份,江西省宜春市某散养鸡场50日龄鸡出现消化不良、精神萎靡、死前倒地抽搐并呈"劈叉姿势"症状,部分病鸡体表存在明显肿瘤。为了对发病鸡所患疾病进行诊断,采用发病情况调查、剖检病变、病理组织学观察及PCR检测方法进行鉴定,同时对阳性条带测序比对,构建系统进化树。结果表明:剖检病死鸡可见心脏、肝脏、肾脏、肺脏有大量灰白色肿瘤灶,脾脏明显肿大。送检病料病理组织切片符合马立克氏病的病理特征;病原核酸检测显示马立克氏病病毒、禽白血病病毒和禽网状内皮组织增殖病病毒为阳性;阳性条带测序比对结果显示禽网状内皮组织增殖病病毒序列与马立克氏病流行毒株有高度相似性,为假阳性结果。说明该鸡场鸡群感染了马立克氏病病毒和禽白血病病毒,可采取改善养殖环境并结合疫苗免疫等方式科学防控,养鸡场应重视马立克氏病的实验室鉴别诊断和防控。     

鸡马立克氏病的诊断与防治

贵州省某鸡场送检发病鸡6只,根据流行病学、剖解病变疑似马立克氏病,经肝脏进行细菌分离培养24 h后观察无细菌生长,琼脂扩散试验为阳性,马立克氏病毒核酸的PCR检测能扩增出目的条带,表明该病为马立克氏病病毒感染所致。     

海兰商品蛋鸡白血病与马立克氏病混合感染案例分析

从临床流行病学、剖检症状、病理变化、马立克氏病、禽白血病、网状内皮组织增生症病毒特异基因的PCR检测等进行综合分析,以期对河南南阳地区某海兰蛋鸡场鸡群的疑似肿瘤疾病进行确诊。结果显示:鸡群大体正常,但零星出现死亡,死亡鸡极度消瘦,心、肝、脾、肺等脏器肿胀及个别病鸡脏器肿胀且有肿瘤结节。PCR检测结果显示:MDV阳性、ALV阳性、REV阴性,确诊鸡群感染马立克氏病病毒和禽白血病病毒。     

硒提高雏鸡抗MDV感染能力的试验

正马立克氏病是鸡的一种淋巴组织增生性肿瘤病,其特征为外周神经淋巴样细胞浸润和增大,引起肢(翅)麻痹,以及在性腺、虹膜、各种脏器、肌肉和皮肤上发生肿瘤病灶。病原为马立克氏病毒(MDV),属于疱疹病毒科禽疱疹病毒2型,是鸡重要的传染病病原,马立克氏病也是目前危害养鸡业健康发展的三大主要疫病(马立克氏病、新城疫、鸡传染性法氏囊炎)之一。马立克氏病在国内     

鸡马立克病毒meq基因克隆及遗传进化分析实验设计

该文设计了一个涉及兽医传染病学、兽医分子生物学等兽医学科专业的探索性实验——鸡马立克病毒meq基因克隆及遗传进化分析实验。实验内容包括通过全自动核酸提取仪、荧光定量PCR仪对鸡组织病料中马立克病毒核酸进行检测,在确定病毒核酸阳性的基础上通过分子生物学手段扩增特征毒力基因meq序列,通过与Genbank已报道的30多株参考毒株序列进行比对,分析该病例马立克病毒的分子特征及遗传进化特点。该实验可锻炼学生的实验动手能力,加深对兽医学相关内容的理解,激发对专业知识的兴趣。     

商品蛋鸡马立克氏病的诊断

某养殖场送检的23只病、死鸡,用琼脂扩散试验检出马立克氏病毒抗原阳性率为95.7%;马立克氏病毒抗体阳性率为100%。根据流行病学、临床症状及实验室诊断,确诊为鸡马立克氏病。同时对马立克氏病与禽淋巴细胞性白血病进行了鉴别诊断。     

鸡马立克氏病的病理学诊断

应用临床调查、肉眼病变和组织病理学观察的方法,对送检病鸡进行了病理学诊断,同时应用PCR快速诊断技术确诊了病原。发现病鸡临床症状表现为精神萎靡,进行性消瘦;剖检病鸡可观察到肝脏和脾脏肿大,有大小不等的白色结节;显微镜下肝脏和脾脏中有大量大小不一的淋巴细胞;PCR成功扩增出预期大小的基因片段。通过临床症状,病理变化和PCR结果判定,确诊为鸡马立克氏病。     

Marek's disease in turkeys. II. Characterization of the viral glycoprotein B gene and antigen of a turkey strain of Marek's disease virus

Marek's disease virus (MDV) causes immunosuppression and tumors in chickens. As sporadic cases of Marek's disease (MD) were recorded in turkeys, the antigenic and genomic characteristics of the MDV glycoprotein B (gB) gene and antigen of turkeys were compared to the chicken MDV gB. The whole chicken and turkey gB genes were sequenced and found identical. By immunoblotting of infected-cell culture lysates using chicken convalescent and gB monoclonal antibodies, the antigenic epitopes of the chicken and turkey viruses were found to differ. The turkey MDV had a unique epitope, compared to the chicken MDV and compared with our previous findings. While the chicken MDV had two epitope types, heat-labile but dithiothreitol (DTT)-stable and heat-stable but DTT-labile, the turkey MDV gB epitope is both heat and DTT-labile.     

Pathogenicity of embryo-adapted serotype 2 OH strain of infectious bursal disease virus in chickens and turkeys

The pathogenicity of serotype 2 OH strain of infectious bursal disease virus (IBDV) to specific-pathogen-free (SPF) chicken embryos and 2-wk-old SPF chickens and turkey poults was investigated. The virus was pathogenic for chicken embryos after five passages as evidenced by pathologic changes in inoculated embryos. The embryo-adapted virus was not pathogenic for 2-wk-old SPF chickens and turkey poults as indicated by lack of clinical signs, gross or microscopic lesions in the bursa of Fabricius of inoculated birds. Bursa-to-body-weight ratios of the inoculated chickens and turkey poults were not significantly different from those of uninoculated controls. Virus-neutralizing antibodies to serotype 2 IBDV were detected in inoculated chickens and turkeys. Results of this study indicated that the embryo-adapted serotype 2 OH IBDV isolate that is pathogenic for chicken embryos is infectious but not pathogenic in chickens and turkeys.     

鸡传染性贫血病毒与马立克氏病病毒共感染SPF鸡群免疫抑制协同作用的研究

将鸡传染性贫血病毒（Chicken infectious anemia virus,CIAV or CAV）和马立克氏病病毒（Marek s dis-ease virus,MDV）人工单一和共同感染1日龄的SPF鸡,感染后分别于14、21、28、35日龄检测鸡体红细胞压积的变化,并检测鸡群疫苗免疫3周后的抗体反应,以探讨CAV与MDV共感染对鸡体的免疫抑制是否有协同作用。结果表明,在血液分析方面,CAV与MDV共感染组较病毒单一感染组与对照组差异极显著,共感染不仅加重了鸡群贫血现象,而且延长了贫血的病理症状;而在禽流感病毒（Avian influenza virus,AIV）H5/H9疫苗、新城疫病毒（Newcastle disease virus,NDV）疫苗和传染性法氏囊病毒（Infectious bursal disease virus,IBDV）疫苗免疫后3周的抗体检测中,CAV与MDV共感染组较其它各实验组差异极显著,抗体滴度大大低于其它实验组;此外,CAV与MDV共感染组,鸡体生长状况明显差于实验各组,有6只鸡只死亡（6/25）,比病毒单一感染时的死亡率大大增加。综上研究证明,CAV与MDV共感染在免疫抑制作用上有协同作用。     

Herpes virus of turkey vaccine: viraemias in field flocks and in experimental chickens

In a field survey of viraemias due to vaccination of chickens with herpes virus of turkey, variation was encountered in titres and percentages of birds viraemic. The incidence of viraemias was much lower in sick than in healthy birds in flocks undergoing mortality from Marek's disease. In a concurrent experiment the same strain of chicken and the same commercial vaccine were used as in the field flocks affected with Marek's disease. A high incidence of viraemia and 84.6 per cent protection against Marek's disease were obtained with single vaccination at one day of age. Revaccination at 21 days of age produced no measurable benefits in the same experiment.     

Immunopathology of Marek's disease in quails: presence of antinuclear antibody and immune complex.

Antinuclear antibody (ANA), a marker for autoimmune reactions, was detected in the sera of quails with Marek's disease (MD). The autoantibody was detected 3 weeks after infection in quails infected with chicken Marek's disease virus and 4 weeks after infection in quails infected with quail Marek's disease virus. The ANA titers were low and ranged from 10 to 40. A speckled type of nuclear fluorescence was the characteristic staining feature. In addition to the presence of ANA, immune complexes (IC) were also detected in the kidney glomeruli of quail infected with Marek's disease virus. Initially about 25-30% of the glomeruli in the kidneys of infected quails had IC deposits. In subsequent periods, the amount of IC deposit and the number of glomeruli showing IC also increased considerably. The findings of the present study suggested autoimmunity may play a pathogenic role in MD.     

应用双重实时荧光定量PCR方法检测鸡马立克氏病血清1型病毒

本研究建立了鸡马立克氏病血清1型病毒（MDV1）绝对定量检测方法。研究中选择MDV1特有的Meq基因的一段保守序列作为检测对象，将其克隆到质粒载体中，作为阳性标准品；同时将管家基因．鸡卵铁转蛋白（Ovo）特异性基因片段克隆到质粒载体上作为内参照的标准品。经荧光定量PCR（FQ-PCR）法扩增获得MDV1的FQ-PCR两条标准曲线，建立了MDV1双重FQ-PCR检测方法。应用该方法绝对定量检测了实验攻毒鸡及吉林省某地发病鸡只的羽髓、淋巴细胞等组织样本中单位细胞病毒拷贝数，并与琼脂扩散（AGP）、常规PCR等检测方法进行比较。结果表明，不论实验攻毒鸡还是自然发病鸡，羽髓中病毒富含量均高于其它组织，每百万宿主细胞内病毒含量为10^7～10^8拷贝；FQ-PCR检测MD发病鸡只的阳性率高于AGP，达100％；该方法的灵敏度比常规PCR检测高10～100倍，在单位细胞内可灵敏地检测到2．78个拷贝的病毒。该方法可以在不同的样品中有效的绝对定量检测MDVl。     

分子修饰前后人参皂苷体外抗马立克氏病毒鸡胚成纤维细胞模型的建立

用雏鸡将马立克氏病毒(MDV)血毒复壮,分离发病鸡淋巴细胞并接种于鸡胚成纤维细胞,观察其病变。获得适应鸡胚成纤维细胞(CEF)的MD强毒,通过电镜观察、琼脂扩散实验进一步鉴定病毒,并建立了MDV感染CEF细胞模型,为研究人参皂苷及其衍生物的体外抗病毒作用及其机制奠定基础。     

Experimental infection with avian leukosis virus isolated from Marek's disease vaccines

Recently, avian leukosis virus (ALV) was isolated from four lots of Marek's disease vaccine produced by two laboratories. The ALVs isolated were characterized by examination of their interactions with cells of two phenotypes (C/E and C/A,E), subgroup-specific polymerase chain reaction (PCR), virus neutralization, envelope gene sequencing, and phylogenetic analysis. All four ALVs are exogenous, belong to subgroup A, and appear to be virtually identical to each other based on PCR and envelope gene nucleotide sequences. We describe herein the characterization of the contaminant viruses in vivo by means of experimental infection in chickens. The contaminant viruses established transient viremia in specified pathogen-free (SPF) Leghorn chickens and elicited a robust and lasting antibody response detectable by enzyme-linked immunosorbent assay. None of the contaminant ALVs induced tumors up to 31 wk of age, and mortality was insignificant. Despite a strong antibody response against the contaminant ALVs, vertical (congenital) transmission to the progeny of experimentally infected SPF chickens took place, albeit at a very low rate (< or = 1.6%). Experimental infection in meat-type chicken embryos resulted in viremia at hatch, suggesting that some meat-type chickens are susceptible to infection and support virus replication.     

用斑点杂交法同时检测鸡群中的CAV MDV和REV

为研究鸡传染性贫血病毒(CAV)在鸡群中的感染状况以及马立克氏病病毒(MDV)和网状内皮细胞增生病病毒(REV)在鸡群中的发病率，用斑点杂交法对山东省4个肉鸡场和2个肉种鸡场进行CAV、MDV和REV的检测，结果表明除一个肉种鸡场没有检测出REV以外，其他鸡场均同时检测出CAV、MDV和REV，并且发现直接从病科中检测CAV的阳性率(20％)远远低于将病科接种SPF鸡胚后的检出率(80％)。