应用PCR从AA肉种鸡鸡胚和1日龄雏鸡体内检测出CIAV和ALV-J

为了了解鸡传染性贫血病病毒（CIAV）和J亚群禽白血病病毒（ALV-J）对AA肉种鸡鸡胚和1日龄雏鸡的感染情况,试验直接从山东省3个不同AA肉种鸡场的鸡胚和1日龄雏鸡的肝、脾、肾、胸腺、骨髓、法氏囊等组织（器官）提取DNA,进行了PCR扩增及PCR产物的克隆和序列测定。结果显示,被检的3个肉种鸡场的鸡胚和1日龄雏鸡体内均有这2种病毒核酸的检出,其中CIAV的阳性率是20.42%,ALV-J的阳性率是15.83%,二者共感染的阳性率是6.25%;在阳性检出率中弱雏〉死胚〉健康雏〉正常胚。病毒核酸在各感染组织（器官）的含量也有所差异,CIAV以脾的含量最多,ALV-J以肾的含量最多。对肝进行细菌分离鉴定时发现,3个鸡场还存在大肠杆菌、沙门氏菌和霉形体的混合感染。结果提示,在AA肉种鸡鸡胚和雏鸡体内存在CIAV、ALV-J的感染和二者的共感染以及继发性大肠杆菌、沙门氏菌和霉形体的混合感染。     

鸡感染J亚群禽白血病病毒的免疫抑制机理

通过人工接种建立了雏鸡感染J亚群禽白血病病毒（ALV-J）后发生免疫抑制的病理模型。对免疫抑制的机理进行了初步研究。结果表明：ALV-J感染早期可诱导胸腺、法氏囊的淋巴细胞凋亡，这种早期的淋巴细胞凋亡是胸腺和法氏囊萎缩的重要原因之一。病理组织学动态观察表明。ALV-J主要引起骨髓的髓系细胞灶状或弥漫性增生，病变导致骨髓机能受损，使机体免疫机能下降。是引起免疫抑制的根本原因。病毒感染组免疫器官均发生了严重的实质萎缩性病变．这种病变的发生除与骨髓的病变及淋巴细胞凋亡有关外。还与中后期淋巴细胞的坏死有关．从而导致严重的免疫抑制。ALV-J感染可引起免疫器官及部分内脏器官中嗜酸性粒细胞样的瘤细胞浸润增生，这可以作为病毒感染早期的病理诊断指标。     

Evaluation of chicken infectious anemia virus and associated risk factors with disease and production losses in broilers

A case-control study was performed to determine the significance of chicken infectious anemia virus (CIAV) as a risk factor associated with secondary disease in commercial broilers and to identify the significance of production losses associated with CIAV. The study also examined the relationship between bursal and thymic atrophy and the presence of CIAV. Cases were defined as submissions to the Alabama Veterinary Diagnostic Laboratories with a history of clinical disease and with a diagnosis of coccidiosis, gangrenous dermatitis, or respiratory disease. Controls were selected from submissions with neither a history of disease nor evidence of disease on necropsy. CIAV was detected in fresh tissues by polymerase chain reaction. Both thymic atrophy and the detection of CIAV were significantly associated with a disease case (P < 0.05). Bursal atrophy was a significant risk factor associated with the detection of CIAV in a submission (P < 0.05). Whereas CIAV was associated with disease cases that showed production losses in both percentage of livability and percentage of condemnations (P < 0.05), detection of CIAV alone was not associated with detectable losses in production or flock performance.     

鸡传染性法氏囊病超强毒感染后SPF鸡免疫器官病理学观察

IBDV超强毒株LX株接种2周龄SPF雏鸡后，其致病性不同于经典强毒株CJ801株，它主要引起接种鸡全身性炎症反应，法氏囊、脾脏、盲肠扁桃体等免疫器官中大量异嗜性白细胞、巨噬细胞浸润，淋巴细胞严重坏死崩解，胸腺皮质严重萎缩、坏死，骨髓中造血细胞减少、巨噬细胞和脂肪细胞增生。在接种后14d法氏囊淋巴滤泡严重萎缩、淋巴细胞排空形成囊腺样结构，未见恢复正常，其它免疫器官形态基本恢复正常。电镜观察，接种后2和4d可见胸腺淋巴细胞胞浆浓集、染色质周边化形成新月形，表现细胞凋亡特征；在法氏囊坏死淋巴细胞胞浆中可见60nm大小呈晶格排列或散在的病毒粒子。研究初步探明了鸡传染性法氏囊病病毒超强毒的致病机理。     

LTR对J亚群禽白血病病毒感染的影响

为探讨LTR基因在骨髓瘤病变型J亚群禽白血病病毒(ALV-J) NX0101致病中的作用,利用反向遗传将血管瘤病变型ALV-J HN06株中两端LTR元件替换至NX0101株的相应位置,拯救出重组病毒NX-HNLTR株.人工接种7日龄SPF雏鸡,分别检测NX0101株和NX-HNLTR株对鸡体的影响.感染鸡生长都较慢.感染NX0101株的鸡,胸腺指数和腔上囊指数明显比对照组低,脾脏指数与对照组相比波动较大,骨髓和脾脏在攻毒后3周可检测到病毒整合到基因组中,胸腺和腔上囊在攻毒后6周才检测到.感染NX-HNLTR株的鸡脾脏指数明显比对照组低,攻毒后2周可检测到病毒整合到脾脏基因组中,骨髓和胸腺分别在攻毒后3周和6周检测到.结果提示,LTR对NX0101株感染鸡的免疫器官有一定的影响.     

Lesions in the thymus and bone marrow in chicks with experimentally induced chicken infectious anemia disease

One-day-old SPF chicks were inoculated with the Cux-l strain of chicken infectious anemia virus (CIAV), and the clinical development of disease and its macroscopic and microscopic alterations in the thymus and bone marrow, were observed. Tissue sections of thymus and bone marrow were stained using the streptavidin-biotin peroxidase method and examined under light microscope for evaluation of antigenic intensities in tissues. Those findings were then compared with blood parameters and ELISA results obtained through collected sera during sacrifice procedures. We sought to determine: the localization of viral antigens in thymus and bone marrow tissues after inoculation, the correlation between antigen intensities and hematologic, serologic and histopathologic findings, definitive diagnostic criteria using histopathologic and immunoperoxidase methods, and the reliability of these methods in the diagnosis of CIAV infection. For this purpose, 83, one-day-old SPF chicks were used. The birds were divided into experimental (n = 52) and control (n = 26) groups. A virus dose of TCID50 of 100,000/ml was administered intramuscularly to every bird in the experimental group. Based on the results of this study, we have suggested that clinical examination, along with macroscopic and microscopic evaluation of the thymus and bone marrow, maybe undertaken starting from day 7 post-inoculation (PI). ELISA, might be of value, as it might give consistent results starting from day 14 PI. However, the most reliable results were obtained through examination of thymus and bone marrow sections from infected birds stained by immunoperoxidase technique, as early as day 4 PI.     

鸡传染性贫血病毒诱导细胞凋亡的研究进展

鸡传染性贫血病毒是鸡传染性贫血病的病原体,能引起雏鸡贫血、淋巴组织萎缩、出血及免疫抑制,是导致许多疫苗免疫失败及雏鸡死亡的主要原因之一。该病毒在世界范围内广泛存在,是养鸡业潜在的巨大威胁。近年来的研究表明,鸡传染性贫血病毒主要是通过其编码的凋亡素诱导雏鸡胸腺和骨髓中的造血祖细胞凋亡而致病的,而凋亡素诱导细胞凋亡的能力与其在细胞中的核定位及半胱氨酸蛋白酶(caspase)等因素密切有关。文章就鸡传染性贫血病毒生物学特征、诱导细胞凋亡的现象及机制的研究进行了综述。     

PCR和斑点杂交检测鸡传染性贫血病毒

通过聚合酶链反应（ｐｃＲ）扩增鸡传染性贫血病毒（ｃＡＶ）ＤＮＡ特定片段，将此ｐｃＲ产物用Ｄｉｇｏｘｉｇｅｎｉｎ标记后作为探针进行斑点杂交，以此检测ＣＡＶ并作流行病学调查。对从江苏省不同地市随机采集的２月龄左右不同疾病的病鸡ＤＮＡ样品进行检测，在被检的５２个不同鸡群来源的病料中，有３６个鸡群来源的病料ＤＮＡ显示ＣＡＶ阳性（群阳性率为６９．２％）；备组织中ＣＡＶＤＮＡ含量有所差异，依次为胸腺＞脾、骨髓、肝＞肾、法氏囊、脑。用ＰＣＲ检测得到的阳性鸡的组织病料感染ＳＰＦ鸡，在感染后第２４天检查，出现贫血症状。初步调查表明，在江苏一些地区ＣＡＶ感染已相当普遍，但它与发病的关系还有待于进一步研究。     

Pathogenicity of a low-virulence duck virus enteritis isolate with apparent immunosuppressive ability

Duck enteritis virus (DEV) was isolated from commercial 2-to-6-wk-old white Pekin ducks experiencing 25%-30% mortality and high morbidity. Secondary infections with Pasteurella multocida, Riemerella anatipestifer, and Escherichia coli were frequently seen in affected ducks. The isolated virus was identical to the prototype DEV by virus neutralization test but differed from the classic DEV by causing lymphoid organ atrophy and inconsistent hemorrhagic lesions in the intestinal annular bands. Attempts to reproduce the disease in white Pekin ducks were unsuccessful until the virulence of the virus was increased by three passages in Muscovy ducklings. Significant thymic atrophy (P < or = 0.001) was detected during the first 10 days postinfection (DPI), but thymus size returned to normal by 17-24 DPI. However, bursal atrophy increased significantly (P < or = 0.001) from 4 DPI until the end of the experiment (39 DPI). Reduction in body weight was significant (P < or = 0.05) between 4 and 6 DPI. There was massive depletion of thymic and bursal lymphocytes with lymphoid necrosis in the thymus, bursa, spleen, and Harderian gland. Eosinophilic intranuclear inclusions were observed in thymus, bursa, spleen, esophagus, cloaca, liver, conjunctiva, and Harderian gland. Occasional intracytoplasmic inclusions were also found scattered in the epithelial cells of conjunctiva, esophagus, bursa of Fabricius, and cloaca. Virus was recovered from experimentally infected ducks from thymus, bursa, spleen, liver, kidneys, trigeminal ganglion, and cloaca during the first 10 days of infection. These findings suggest that a low-virulent DEV can cause a massive lymphoid atrophy and can sustain immunosuppression as noted by the secondary bacterial infection.     

Evaluation of reconstitution of immunodeficient mice by partially purified thymic extracts.

Partially purified thymus products were used to evaluate the maturation of T lymphocytes in immunodeficient mice. Three different bovine thymic extracts, designated thymic extracts A, B, and C, were successful in increasing the longevity of conventionally raised nude mice. Daily injection of bovine thymic extracts A, B, and C and mouse thymus extracts failed to mature a population of T lymphocytes and restore the capacity to reject heart allografts. Preincubation of normal syngeneic bone marrow with thymic extract B in vitro before injection into nude mice also failed to reconstitute the host's ability to reject heart grafts. The number of antibody plaque-forming cells of sheep red blood cells in lethally irradiated, bone marrow-reconstituted mice could be increased by preincubating the bone marrow cells with bovine thymic extract fraction B before injection followed by daily injections. A similar but less marked increase in plaque-forming cells was obtained by the daily injection of bovine thymic extract fraction C. Complete functional maturation of T lymphocytes in immunodeficient animals appears to require more than thymic extract stimulation of bone marrow cells or pre-T lymphocytes.     

谷氨酰胺对肉鸡免疫器官胚后发育的影响

160只1日龄艾维茵肉仔鸡随机分成4组。分别饲喂添加0％、0．2％、0．4％和0．8％谷氨酰胺（Gln）饲粮28d。每周末取鸡24只，每组6只，颈静脉放血致死，取胸腺、法氏囊和脾脏，称重．制作切片．光镜观察，研究基础日粮中添加Gln对肉鸡免疫器官胚后发育的影响。结果显示：（1）添加0．8％Gln组1、4周龄和添加0．4％Gln组2、4周龄胸腺重量显著高于对照组（P〈0．05），添加0．8％Gln组2周龄极显著高于对照组（P〈0．01）；添加0．4％Gln组1、2、4周龄和添加0．8％Gln组4周龄法氏囊重量显著高于对照组（P〈0．05），添加0．8％Gln组1、2周龄极显著高于对照组（P〈0．01）；试验各组脾脏重量均高于对照组。添加0．8％Gln组4周龄显著高于对照组（P〈0．05）。（2）试验组胸腺小叶皮质增厚，淋巴细胞密集，髓质比例减小。胸腺小体减少、减小。试验组法氏囊皱襞内淋巴滤泡增多，皮质增厚．淋巴细胞增多。试验组脾脏脾小结增多、增大；动脉周围淋巴鞘增厚；椭球增多、增大．细胞排列疏松．鞘毛细血管管腔变大．内皮细胞增高。间隙增大。添加0．8％Gln对肉鸡免疫器官发育影响最为明显。结果表明，日粮中添加Gln对肉鸡中枢淋巴器官胚后发育的促进作用优于外周淋巴器官。并有延缓法氏囊退化作用。本试验从组织学角度证明Gln能够促进机体淋巴细胞的增殖分化及免疫器官的胚后发育，进而提高机体的免疫功能。     

DNA探针在鸡传染性贫血流行病学调查中的应用

用地高辛标记的核酸(DNA)探针对江都、连云港、苏州、无锡等地3个市县9个养鸡场(户)随机取样,采集106份样品,分别为12日龄～60日龄的肉鸡、蛋鸡的胸腺、法氏囊和脾脏组织,抽提DNA检测CIAV感染情况。鸡传染性贫血病毒阳性鸡只22份,阳性率为20.7%,各鸡场阳性率从5%到50%不等。结果表明,在江苏省的一些地区鸡群中CIAV的感染已非常普遍,有的地区甚至十分严重,控制该病的发生已刻不容缓。     

Coinfection of specific-pathogen-free chickens with Marek's disease virus (MDV) and chicken infectious anemia virus: effect of MDV pathotype

Both Marek's disease virus (MDV) and chicken infectious anemia virus (CIAV) infections are prevalent in chickens throughout the world. In the past decade, MDV strains with increased virulence (very virulent plus MDV pathotype [vv+MDV]) have been isolated. The purpose of this experiment was to determine the effects of coinfection of chickens with CIAV and a vv+MDV isolate. Specific-pathogen-free chickens were inoculated at 1 day posthatch with RB1B (very virulent MDV pathotype [vvMDV]) only, 584A (vv+MDV) only, CIAV only, RB1B + CIAV, 584A + CIAV, or nothing. Samples of spleen, thymus, and bursa of Fabricius were collected at 4, 7, 10, and 13 days postinoculation (DPI). Thymic and bursal atrophy at 13 DPI and final mortality at 30 DPI were significantly greater in chickens inoculated with 584A with or without added CIAV, or with RB1B plus CIAV, compared with birds inoculated with RB1B alone. Both amounts of virus reisolated and levels of virus detected by quantitative-competitive polymerase chain reaction were greater at 4 DPI in 584A inoculates compared with RB1B inoculates. To monitor the early cytolytic infection, northern analysis was done with a probe for the MDV immediate early gene ICP4 (infected cell protein 4). In the absence of CIAV, ICP4 expression was more apparent in chickens inoculated with 584A than in those inoculated with RB1B. CIAV coinfection increased ICP4 expression in the spleens of chickens infected with RB1B. These results indicated that inoculation of chickens with the 584A isolate caused a more robust early cytolytic infection compared with inoculation with RB1B alone and support the classification of 584A as a vv+MDV strain. Coinfection with CIAV exacerbated vvMDV strain RB1B infection. The extent of this exacerbation was less evident when birds were coinfected with 584A and CIAV.     

一株鸡传染性贫血病毒对不同日龄SPF鸡的致病性研究

为评价鸡传染性贫血病毒AV1550株的致病性,取1日龄、7日龄和14日龄SPF鸡分别经胸部肌肉注射不同病毒含量的病毒液,同时设置正常对照,隔离饲养观察21日。感染后14日采血测定红细胞压积,21日统计死亡率、体重变化以及胸腺、骨髓、法氏囊病变情况并测定1日龄SPF鸡感染后不同组织中的病毒载量。结果表明,1日龄SPF鸡感染AV1550株后,表现出精神沉郁、增重减缓、贫血等明显的临床症状,死亡率为53.9%;死亡鸡或观察期结束时存活鸡剖检,可见胸腺萎缩,骨髓变成淡黄色;不同剂量感染后14日,均能引起红细胞压积显著下降;21日时,胸腺病毒载量最高,可达106.7copies/mg 。7日龄SPF鸡感染后,出现增重减缓,高感染剂量（100000EID50）出现贫血,部分鸡出现胸腺萎缩和骨髓病变,但病变率低于30%。14日龄SPF鸡感染后,不引起明显临床症状。研究证实,CAV对SPF鸡的致病性具有明显的日龄依赖性,红细胞压积降低、骨髓病变、胸腺萎缩以及胸腺病毒载量测定可作为评价CAV致病的指标。     

鸡贫血病—法氏囊病联合免疫母鸡后子代雏鸡免疫器官T细胞和抗体生成细胞数量的动态变化

对鸡传染性贫血病（CIA）－传染性法氏囊病（IBD）联合免疫母鸡后的子代雏鸡免疫器官的免疫学化变化进行了研究。结果发现,混合感染CIAV、IBDV雏鸡免疫器官T细胞和IgG,IgM,IgA抗体生成细胞数量在27日龄内明显未免疫对照组、联合免疫组和联合免疫攻毒组,表明感染CIAV、IBDV的雏鸡全身免疫功能显著下降,CAI－IBD联合免疫母鸡后,子代雏鸡T细胞胞和IgG,IgM,IgA抗体生成细胞数量在27日龄内,较未免疫对照组明显增加,表明CIA－IBD联合免疫母鸡可使子代雏鸡免疫器官的免疫功能增强,能抵御强毒攻击。     

Pathogenesis of duck plague in the bursa of Fabricius, thymus, and spleen.

White Pekin ducks were inoculated orally with duck plague virus and killed at 24-hour intervals after inoculation. Spleen, thymus, and bursa of Fabricius were collected and examined by light, fluorescent, and electron microscopy. Necrosis of lymphocytes occurred in the bursa of Fabricius, thymus, splenic periarteriolar lymphoid sheath (T lymphocytes), and splenic germinal centers (B lymphocytes). Viral nucleocapsids were present in the karyoplasm of lymphocytes, but these cells necrotized before virions were formed. Periarteriolar reticular sheath cells and sinusoidal lining cells in the spleen, epithelial cells in Hassall's corpuscle of the thymus, epithelial cells between the cortex and medulla of the follicles in the bursa of Fabricius, and macrophages in all 3 tissues contained nucleocapsids in the nuclei and virions in cytoplasmic vacuoles before necrosis occurred.     

Atrophy of the primary lymphoid organs of meat pigeons in Italy associated with circoviruslike particles in the bursa of Fabricius

During a survey effected in a meat pigeon slaughterhouse of central Italy, atrophy of primary lymphoid organs (bursa of Fabricius and thymus) and hypoplasia of bone marrow were observed. Histologic, ultrastructural, and hematologic examinations were performed on a total of 80 randomly selected 30-day-old meat pigeons. By histologic studies, lymphocytic depletion and necrosis with cyst formations in the bursa of Fabricius were detected in all subjects that showed thymus and bursa atrophy at necropsy. Basophilic intranuclear inclusions were also observed in bursal cells. After ultrastructural studies, these inclusions were proved to be viral particles resembling circoviruslike particles in morphology and size. Severe lymphocytic depletion of the bursa was plausibly associated with the presence of the viral particles.     

鸡传染性贫血病毒感染对雏鸡免疫系统的影响Ⅱ．几种细胞因子体外诱生活性动态变化

１日龄雏鸡人工感染鸡传染性贫血病毒（ＣＩＡＶ）后７、１４ｄ，脾脏Ｔ淋巴细胞ＩＬ－２活性明显降低（Ｐ＜０．０１，Ｐ＜０．０５），２１ｄ明显回升，２８ｄ显著升高（Ｐ＜０．０５），３５ｄ后降至对照水平；感染后７、１４和２１ｄ，胸腺Ｔ淋巴细胞ＩＬ－２诱生活性明显降低（Ｐ＜０．０５），２８ｄ后恢复至对照水平。感染后７、１４ｄ，脾脏淋巴细胞ＩＦＮ诱生活性明显低于对照鸡（Ｐ＜０．０５），２１ｄ后恢复至对照水平。感染后１４ｄ，胸腺淋巴细胞集落刺激因子（ＣＳＦ）诱生活性未见明显变化，２１ｄ显著升高（Ｐ＜０．０５），２８ｄ降至正常水平。