鸡传染性支气管炎活疫苗(D971P株)免疫效果好

<正> 鸡传染性支气管炎(IB)是由传染性支气管炎病毒(IBV)引起的一种急性、高度接触性传染病,在世界范围内广泛流行。根据临床与组织嗜性通常可分为呼吸型、肾病变型和腺胃病变型三种类型。自1995年以来,腺胃病变型IB在我国部分省(市)流行,造成巨大的经济损失,为了控制肾型、腺胃病变型IB的发生和流行,中国农业科学院哈尔滨兽医研究所研制出鸡传染性支气管炎活疫苗(D971P株),技术特点主要有以下几个方面。1.该疫苗具有自主知识产权中国农业科学院哈尔滨兽医研究所科研究员荣骏弓等从国内发病鸡群中分离到的腺胃病变型鸡传染性支气管炎病毒(IBV-D971株)并进行了系统鉴定,该成果"腺胃病变型鸡传染性支气管炎病毒的分离鉴定"于2003年1月20日通过了农业部成果鉴定,获中国农业科学院科技进步2等奖。利用该病毒(IBV-D971株)在SPF鸡胚上连续传105代,培育出     

鸡传染性支气管炎病毒D971分离株与M41株、T株接种鸡的病理形态学比较研究

鸡传染性支气管炎病毒D971分离株感染SPF鸡后主要表现为消瘦、稀便、伴有轻微的呼吸症状；剖检可见腺胃乳头肿胀、充出血、或乳头凹陷，并附有大量黏液；组织学观察可见腺胃黏膜上皮和固有层有坏死、脱落或溃疡，个别病例见有淋巴样细胞浸润，肌胃黏膜有中度坏死，伴有少量淋巴样细胞浸润，实质器官细胞变性、坏死，脾、胸腺及法氏囊的淋巴细胞有较为明显的坏死。M41株和T株感染SPF鸡后，其腺胃变化不明显。     

腺胃病变型鸡传染性支气管炎病毒（IBV—D971株）致弱的研究

将IBV-D971株病毒在SPF鸡胚上连续传至120代,培育出了一株毒力明显减弱,并且具有良好免疫原性的腺胃病变型鸡传染性支气管炎病毒弱毒株(IBV-D971株).IBV-D971 F105代毒力明显下降,以109.25EID50接种1日龄SPF鸡5/5无不良反应.IBV-D971 F105代毒连续通过1日龄SPF鸡体5代,未见毒力返强.以103EID50免疫1日龄SPF鸡,攻毒后5/5保护,对照5/5发病.     

鸡传染性支气管炎活疫苗(D971p株)的研究

从国内发病鸡群中分离到传染性支气管炎（嗜腺胃型）病毒（IBV-D971株），在SPF鸡胚上连续传至120代，培育出IBV-D971p弱毒株。用该毒株制备活疫苗，经安全、效力等试验证明具有较高的免疫保护率和安全性，能有效的预防腺胃型、肾型鸡传染性支气管炎。     

鸡传染性腺胃炎的综述

1病原学临床表现为腺胃炎的病例，其病原学非常复杂，尚未定论。我国学者王玉东等从患腺胃肿大病鸡中分离到冠状病毒，认为是传染性支气管炎病毒（IBV）的变异株：朱国强等从江苏腺胃肿大的病鸡中分离到H95病毒，     

鸡传染性支气管炎活疫苗(D971P株)免疫效果好

鸡传染性支气管炎(IB)是由传染性支气管炎病毒(IBV)引起的一种急性、高度接触性传染病,在世界范围内广泛流行.根据临床与组织嗜性通常可分为呼吸型、肾病变型和腺胃病变型三种类型.自1995年以来,腺胃病变型IB在我国部分省(市)流行,造成巨大的经济损失,为了控制肾型、腺胃病变型IB的发生和流行,中国农业科学院哈尔滨兽医研究所研制出鸡传染性支气管炎活疫苗(D971P株),技术特点主要有以下几个方面.     

鸡腺胃型传染性支气管炎的诊治与基因序列分析

从广东省某鸡场疑似腺胃型传染性支气管炎病死鸡的腺胃中分离到一株病毒,初步鉴定为QX型传染性支气管炎病毒(IBV)。通过疫苗免疫与其他辅助措施,疫病得到有效控制。但动物回归试验表明,IBV毒株单独感染并不能导致鸡产生腺胃炎症状。     

鸡传染性腺胃炎的诊断

<正>1病原学临床表现为腺胃炎的病例,其病原学非常复杂,因分离到的病原难以复制出临床典型病例,故真正病原尚未定论。我国学者朱国强等从江苏腺胃肿大的病鸡中分离到H95病毒,认为与传染性支气管炎病毒有密切的血清学关系;王玉东等从患腺胃肿大病鸡中分离到冠状病毒,认为是传染性支气管炎病毒(IBV)的变异株。     

肉仔鸡肾型传染性支气管炎的诊治研究

鸡传染性支气管炎是由传染性支气管炎病毒（IBV）引起的危害养鸡业的重要传染病之一。该病的特点是病情复杂，血清型众多，又具有高度传染性，鸡群同时感染不同毒株或使用弱毒疫苗发生基因重组而不断变异，使IBV侵害鸡体的靶器官也发生了变化。IgnjatovicJ等用澳大利亚分离的25株IBV人工感染SPF鸡，比较了其致病性，发现12株肾致病型IBV，10株呼吸型IBV，3株混合致病型IBV，侵害肺、肾。为有效防制本病，我们对其病原进行了分离与鉴定。     

1株肾型鸡传染性支气管炎病毒的鉴定

鸡传染性支气管炎病毒(IBV)具有不同致病特性,将IBV XDC-2株接种9日龄SPF鸡胚培养,可引起鸡胚死亡和出现侏儒胚,病毒EID50达5×10-5.33/mL。将IBV XDC-2株接种18日龄SPF鸡,饲养观察14 d,病鸡临床症状表现为:精神沉郁,羽毛凌乱,双翅下垂,轻微腹泻,多数拉白色水样稀粪。病死鸡出现肾肿大、呈花斑状、大量尿酸盐沉积。鸡发病率为100%,死亡率为25%。死亡鸡肺脏、肾组织制作组织切片,发现病理变化明显,主要为:肾小管扩张,上皮细胞呈玻璃样变性,部分管腔内可见坏死脱落之上皮细胞,于肾间质可见大量单核细胞浸润,肾间质有充血、出血现象;肺内动脉、毛细血管充血,淋巴细胞浸润。死亡鸡肺脏、肾组织接种鸡胚分离病毒,RT-PCR检测结果为阳性,表明该分离株为鸡传染性支气管炎病毒,具有很强的嗜肾性。     

鸡传染性支气管炎H120、W93株活疫苗生产工艺的改进

分别以鸡传染性支气管炎病毒(In-fectiousbronchitisvirus,IBV)H120株、W93株不同病毒量接种10、12日龄SPF鸡胚,并改变后孵化温度生产抗原液,接种后不同时间收获胚液,根据鸡胚的早死率以及胚液的收获量、病毒滴度,选择最佳的生产工艺。结果表明,IBVH120株、W93株以102.7EID50/0.1mL的病毒量接种12日龄鸡胚尿囊腔,在34.5℃孵育35h,可较为显著地降低早死胚率,提高产毒量,且抗原液病毒滴度符合疫苗制备规程的要求。     

鸡新城疫－传染性支气管炎－传染性法氏囊病三联灭活疫苗对不同日龄雏鸡的免疫效力研究

为评估鸡新城疫（ND）-传染性支气管炎（IB）-传染性法氏囊病（IBD）三联灭活疫苗对不同日龄和不同水平母源抗体雏鸡的免疫效力和持续期,本试验用该疫苗免疫7、14、21日龄SPF雏鸡和有母源抗体的普通雏鸡,免疫后采血测定ND血凝抑制抗体（HI Ab）、IB血凝抑制抗体（HI Ab）及IBD中和抗体（NA）,并用传染性法氏囊病病毒（IBDV）强毒攻击。结果显示,7日龄SPF雏鸡免疫后21 d ND HI抗体、IB HI抗体及IBD中和抗体效价分别为7.9log2、6.9log2和14.1log2,SPF鸡日龄越大,抗体水平越高;28日龄SPF鸡免疫后3个月,0.3 mL免疫剂量组试验鸡ND HI、IB HI及IBD中和抗体效价分别达6.5log2、6.1log2和13.6log2,IBDV攻毒保护率均为100%（10/10）;不同日龄普通雏鸡免疫效果与SPF鸡试验一致,抗体水平随鸡日龄增大而升高,IBD攻毒保护率也都达到100%（10/10）。试验结果证实,鸡新城疫-传染性支气管炎-传染性法氏囊病三联灭活疫苗可使7、14及21日龄SPF雏鸡和普通雏鸡产生良好的免疫力,对雏鸡的免疫期至少为3个月。     

Attenuation of avian infectious bronchitis virus by cold-adaptation.

Avian infectious bronchitis virus (IBV) Arkansas-type DPI strain was passaged 10 times in specific-pathogen-free (SPF) chicken embryos incubated at 28 C and 37 C. Virus grown at 28 C acquired cold-adapted (CA) and temperature-sensitive (TS) characteristics based on more-rapid growth at 28 C and a reduced ability to grown at 41 C, respectively, compared with non-cold-adapted (non-CA) virus grown at 37 C. The pathogenicity and immunogenicity were determined for CA and non-CA IBV in 1-day-old SPF chickens following intratracheal inoculation. The percentage of CA IBV-vaccinated chicks exhibiting respiratory disease exceeded 30% on only 1 day postinoculation (PI) (day 5 PI), compared with 8 days (days 2-9 PI) for birds given non-CA IBV. Mortality was 0% for CA IBV-vaccinated chickens and 6% for non-CA virus-vaccinated chickens. Microscopically, both CA and non-CA IBV caused diffuse tracheal deciliation, although mucosal hyperplasia, necrosis, and heterophil infiltration were more severe with non-CA IBV. Virus was reisolated from kidneys of chickens given CA IBV, suggesting the loss of the TS property. The instability of the TS property was confirmed by growth of the reisolated virus at 41 C. Both CA and non-CA viruses induced complete protection against homologous challenge virus infection of the upper respiratory tract.     

Pathologic study of specific-pathogen-free chicks and hens inoculated with adenovirus isolated from hydropericardium syndrome.

The mortality and pathology caused by serotype 4 adenovirus, isolated from chickens with hydropericardium syndrome (HPS) in Japan, was investigated in specific-pathogen-free (SPF) chickens. One-day-old to 15-mo-old SPF chickens were inoculated intramuscularly, orally, and intranasally with liver homogenates from HPS chickens or isolated serotype 4 adenovirus. There were no clinical signs before death. The mortality rate in all groups of 1-day-old chicks was 100%, irrespective of the inoculum or inoculation route. Four-week-old chickens inoculated with liver homogenate also had a 100% mortality rate. Five-week-old chickens inoculated with cell culture of HPS adenovirus had a 40% mortality rate. The mortality rates of 7-mo-old hens inoculated with liver homogenates intramuscularly and orally were 75% and 25%, respectively. In 15-mo-old hens inoculated with liver homogenates intramuscularly, the mortality rate was 70%. Gross lesions were hydropericardium and swelling and congestion of the liver with occasional petechial hemorrhages. Histologically, the liver had diffuse or multifocal hepatic necrosis and hemorrhage with intranuclear inclusion bodies noted within hepatocytes. In the spleen, macrophages containing erythrocytes and yellow pigment were prominent in the red pulp. In the lung, a moderate diffuse macrophage infiltration was noted throughout the lung parenchyma, and these macrophages contained yellow pigment. In the pancreas of the chicks inoculated at 1 day old, there was multifocal necrosis of glands with intranuclear inclusion bodies. Intranuclear inclusion bodies were seen also in the gizzard, proventriculus, duodenum, cecum, kidney, and lung of the chicks inoculated at 1 day old. Immunohistochemically, the intranuclear inclusion bodies of various organs showed positive reactions against group I avian adenovirus. Adenovirus was recovered from the liver of chickens with HPS. This study indicates that HPS adenovirus is able to reproduce HPS lesions and mortality in SPF chicks and even adult chickens and that it is a highly pathogenic strain.     

Differences in the induction of urate deposition of specific-pathogen-free chicks inoculated with avian nephritis virus passaged by five different methods.

The G-4260 strain of avian nephritis virus (ANV) was passaged using five different methods as follows: method 1, passage three times in chorioallantoic membrane (CAM) of 11-day-old embryonated eggs (CAM3); method 2, passage twice in CAM and further passage once in yolk sac (YS) of 6-day-old embryonated eggs (CAM2-YS1); method 3, passage 11 times in CAM (CAM11); method 4, passage 10 times in CAM and further passage twice in YS (CAM10-YS2); method 5, passage as in Method 4 and then passage three times in chicken kidney cell culture (CAM10-YS2-CK3). CAM11 and CAM10-YS2 were each inoculated orally into 25 one-day-old specific-pathogen-free (SPF) chicks. Seven chicks in the CAM11-inoculated group and six chicks in the CAM10-YS2-inoculated group died or were killed because they were moribund; all had either nephrosis or urate deposition. CAM3, CAM2-YS1, CAM10-YS2, and CAM10-YS2-CK3 were each inoculated intraperitoneally into 15 one-day-old SPF chicks. No chicks inoculated with CAM3 or CAM2-YS1 died, but wo chicks inoculated with CAM10-YS2 and three inoculated with CAM10-YS2-CK3 died with urate deposition. At 14 postinoculation, plasma urate values of the CAM10-YS2- and CAM10-YS2-CK3-inoculated chicks were significantly higher than those of CAM3- and CAM2-YS1-inoculated chicks and control chicks (P less than 0.01). However, interstitial nephritis was observed in most of the ANV-inoculated birds.     

不同来源传染性支气管炎病毒诱导SPF鸡发病的免疫机制研究

试验旨在探讨不同来源的传染性支气管炎病毒(Infectious bronchitis virus,IBV)诱导SPF鸡发病的免疫机制。选用140只1日龄SPF白来航鸡,随机分为4组,3组攻毒组通过滴鼻点眼途径分别接种鸡源IBV强毒株、鸡源IBV弱毒株和野鸡源IBV毒株3个毒株,对照组以同种方式接种等量灭菌的磷酸盐缓冲液。在感染后12 h、36 h、72 h、7 d和14 d,每组随机选取5只进行剖检,并分别采集法氏囊、肾脏和气管组织,剩余鸡用于观察临床症状、发病及死亡情况。应用实时荧光定量PCR检测攻毒后不同时间点采集的各组织中IBV的病毒载量、Toll样受体(Toll-like receptors,TLRs)及部分细胞因子(白细胞介素(interleukin,IL)和干扰素(interferon,IFN))表达量的变化。结果显示,感染不同来源IBV毒株之后仅鸡源IBV强毒株感染组SPF鸡出现抑郁、翅膀下垂、甩头等典型的临床症状,且在感染后5~10 d共有7只死亡,死亡率为20%。病理剖检发现,感染鸡源IBV强毒株的鸡肾脏肿大、尿酸盐沉积和有花斑样病变,而感染野鸡源IBV毒株、鸡源IBV弱毒株和对照组的鸡无明显的眼观病变。实时荧光定量PCR结果显示,在鸡源IBV强毒株组的法氏囊、肾脏和气管3个组织中均检测到病毒。对照组和野鸡源IBV毒株组中均未检测到病毒,鸡源IBV弱毒株组只在部分组织中检测到病毒。在感染后72 h,鸡源IBV强毒株组与其他各组相比,TLR1、TLR2、TLR3、TLR5、TLR7和TLR15基因在法氏囊中的表达量均显著升高(P<0.05),IL-6和IFN-β参与更强烈的抗病毒免疫反应;在感染后7 d,鸡源IBV弱毒株组与其他各组相比,肾脏中TLR2、TLR3、TLR15、TLR21、IL-6和IL-18基因表达量均显著升高(P<0.05)。野鸡源IBV感染后36 h法氏囊组织中IFN-γ基因表达量显著上调(P<0.05)。综上所述,3个IBV毒株中仅鸡源IBV强毒感染引起SPF鸡典型临床发病症状与可视组织病变,且可提高SPF鸡组织中免疫相关因子的基因表达量。本研究结果揭示,不同来源的IBV对SPF鸡的不同致病性与其感染诱导的免疫反应不同有关。     

鸡白痢沙门氏菌鸡胚感染模型建立方法的筛选

为筛选出理想的鸡白痢沙门氏菌宿主感染模型的建立方法,本研究通过选择不同接种方式配合不同菌液浓度接种不同胚龄鸡胚建立感染模型,将90枚SPF鸡胚随机分成9组,每组10枚,分别为A、B、C、D、E、F、G、H、I组,A组为空白对照组,其余各组为模型组,模型组分别以气室接种、蛋壳接触接种的方式对不同胚龄（11和14胚龄）SPF鸡胚接种不同浓度（1×10³、3×10³、9×10³、3×10⁵、9×10⁵ CFU/mL）的鸡白痢沙门氏菌C79-3菌株。每天观察情况直至出雏,待雏鸡孵出后按组分笼饲养,每隔12 h观察1次,连续观察7 d,观察各组雏鸡的临床症状、死亡情况,观察期结束对死亡和存活雏鸡进行剖检、组织病理学检查及鸡白痢沙门氏菌的分离鉴定。结果显示,各模型组均有雏鸡出现明显的鸡白痢病症状并死亡,相较于其他模型组,以11胚龄蛋壳接触感染方式接种9×10⁵ CFU/mL的C79-3菌液的F组和14胚龄气室感染方式接种3×10³ CFU/mL C79-3菌液0.1 mL的H组,鸡胚出壳率较高,可分别达到80%和90%,出壳雏鸡呈现明显鸡白痢病特征,剖检可见肝脏出血、盲肠膨大、卵黄囊吸收不良;组织病理切片可观察到肝脏、盲肠、心脏各有不同程度的损伤,发病率分别100%和88.8%,并且鸡白痢沙门氏菌阳性检出率分别达70%和90%。本研究结果表明,以11胚龄蛋壳接触感染方式接种0.1 mL 9×10⁵ CFU/mL的C79-3菌液和14胚龄以气室感染方式接种0.1 mL 3×10³ CFU/mL的C79-3菌液的两种方法均可用于建立稳定的鸡白痢沙门氏菌宿主感染模型。     

Pathogenicity and molecular characteristics of infectious bronchitis virus (IBV) strains isolated from broilers showing diarrhoea and respiratory disease

1. The possibility that infectious bronchitis virus (IBV) variants isolated from broilers with enteric and respiratory problems have a different tropism and pathological outcome from those IBV strains causing classical respiratory disease was investigated.

2. IBV variants were isolated from broiler flocks with enteric and respiratory problems in two regions of Brazil. The USP-10 isolate, of enteric origin, was inoculated via the oral oroculonasal routes into IBV-antibody-free broilers and specific pathogen-free (SPF) chickens to determine tissue tropism and pathogenicity and compared with an IBV variant (USP-50) isolated from chickens showing signs of respiratory disease only.

3. Both USP-10 and USP-50 strains caused similar pathological patterns by either route of inoculation. Both variants were detected in respiratory and non-respiratory tissues, including the kidney, intestine and testis.

4. Broilers were more susceptible to infection than SPF chickens, and seroconversion was detected in all of the chicks.