[1]吴 昊,牛伟杰,王久红,等.三基因缺失重组弱毒株(PRV-HNX-TK-/gE-/gG-+PCV2 ORF2)不同途径免疫的效果分析[J].中国预防兽医学报,2019,(12):1261-1267.[doi:0.3969/j.issn.1008-0589.201902004]
 WU Hao,NIU Wei-jie,WANG Jiu-hong,et al.Evaluation of different immune pathways for porcine pseudorabies virus three gene deletion attenuated recombinant strain(PRV-HNX-TK-/gE-/gG-+PCV2 ORF2)[J].Chinese journal of preventive veterinary medicine,2019,(12):1261-1267.[doi:0.3969/j.issn.1008-0589.201902004]
点击复制

三基因缺失重组弱毒株(PRV-HNX-TK-/gE-/gG-+
PCV2 ORF2)不同途径免疫的效果分析
()
分享到:

《中国预防兽医学报》[ISSN:1008-0589/CN:23-1417/S]

卷:
期数:
2019年12
页码:
1261-1267
栏目:
出版日期:
2020-01-25

文章信息/Info

Title:
Evaluation of different immune pathways for porcine pseudorabies virus three gene deletion attenuated recombinant strain(PRV-HNX-TK-/gE-/gG-+PCV2 ORF2)
文章编号:
1008-0589(2019)12-1261-07
作者:
 

吴 昊123牛伟杰23王久红23胡耀方23姚 伦23库旭钢3何启盖123*

(1. 华中农业大学 农业微生物学国家重点实验室,湖北 武汉 430070;2. 华中农业大学 动物医学学院,湖北 武汉 430070;
3. 生猪健康养殖协同创新中心,湖北 武汉 430070)
Author(s):
 

WU Hao123 NIU Wei-jie23 WANG Jiu-hong23 HU Yao-fang23 YAO Lun23 KU Xu-gang3 HE Qi-gai123*

 (1. State Key Laboratory of Agricultural Microbiology, Huazhong Agricaltural University, Wuhan 430070, China;
2. College of veterinary Medicine Huazhong Agricultural University, Wuhan 430070, China;
关键词:
 PRV缺失弱毒株口服免疫免疫效果评价
Keywords:
PRV three gene deletion recombinant attenuated vaccine  oral immunization  immune effect evaluation
分类号:
S852.65
DOI:
0.3969/j.issn.1008-0589.201902004
文献标志码:
A
摘要:
为评价口服免疫猪伪狂犬病毒三基因缺失重组弱毒株 PRV-HNX-TK-/gE-/gG-+PCV2 ORF2对母源抗体阳性保育猪的保护效果,本研究将该重组弱毒株通过口服和颈部肌肉注射两种不同途径免疫母源抗体阳性的断奶仔猪,通过ELISA方法测定血清PRV gB抗体、PCV2 Cap抗体和IFN-γ水平,利用固定病毒-稀释血清的中和试验方法检测PRV中和抗体水平;免疫结束后攻毒,记录各组猪临床表现和病理变化、测定其鼻拭子和肛拭子中病毒排毒时间、病毒载量等指标。结果显示,整个免疫试验过程中,口服组与注射组猪的PRV gB抗体均维持阳性、血清中PRV中和抗体效价均高于1∶4、IFN-γ含量在免疫后均能达到300 pg/mL以上,PCV2抗体阳性率则出现下降;而未免疫组猪在第7 d后gB抗体转阴,中和抗体效价低于1:4、IFN-γ含量维持在约250 pg/mL,PCV2抗体阳性率(母源抗体)持续到二免后28 d才开始下降。免疫后攻毒,检测结果显示,口服组与注射组比对照组猪提前2 d体温下降,提前7 d恢复采食,而对照组猪出现高温、打喷嚏、咳嗽、流鼻涕、食欲不振、精神沉郁和神经症状并持续至攻毒后第8 d,并于攻毒后第12 d开始采食;口服组与注射组猪排毒高峰在攻毒后第2 d~6 d,第12 d后均停止排毒,对照组猪排毒持续期大于14 d;脑组织病理切片观察可见,口服组与注射组猪脑组织中仅少量小胶质细胞和淋巴细胞增多,而对照组猪脑组织中出现胶质细胞结节、血管袖套现象、卫星现象、噬神经元现象等病理变化,对照组猪扁桃体隐窝腔内含有大量脱落的隐窝上皮细胞、白细胞和炎性渗出液,并有燕麦细胞增多。以上结果表明口服方式免疫该三基因缺失重组弱毒株能够使保育猪产生针对PRV的有效免疫应答,但并不能诱导高水平的PCV2抗体。本研究为PRV疫苗研发带来了新的方向,也为免疫途径提供了更多选择,同时首次证实了该缺失弱毒株口服和肌注均能提供相似的免疫保护效果。
  
Abstract:
Abstract:  In order to evaluate the immune effect of porcine pseudorabies virus (PRV) three gene deletion attenuated strain (PRV-HNX-TK-/gE-/gG-+PCV2 ORF2), maternal antibody-positive nursery pigs were vaccinated through oral and neck intramuscular immunization. Serum PRV gB antibody, PCV2 Cap antibody and IFN-γ levels were determined by ELISA, and PRV neutralizing antibody levels were detected by neutralization assay. After challenge, clinical manifestations and pathological changes were observed and recorded, and virus shedding in nasal swabs and rectal swabs was measured by fluorescence quantitative PCR. Duration of virus shedding from each group was also examined. Throughout the whole immunization experiment, the gB antibody in the oral and intramuscular groups remained positive, the PRV neutralizing antibody titer was higher than 1:4, and IFN-γ reached above 300 pg/mL, and PCV2 antibody positive rate decreased. While in the control group, gB antibody became negative  after 35 days of age; the PRV neutralizing antibody titer was lower than 1∶4, and  the levels of IFN-γ were maintained at around 250 pg/mL with no significant change. After the challenge, the clinical symptoms of the oral group and the injection group were alleviated from the fourth day, and the body temperature began to decrease 2 days earlier than the control group, and feeding began 7 days earlier than the control group. The control group showed symptoms such as elevated body temperature, sneezing, coughing, runny nose, loss of appetite, mental depression and neurological symptoms, which continued until the eighth day, and feeding began the twelfth day; virus shedding peak in each group ranged from day 2 to day 6, after day 12 two vaccinated groups stopped shedding; however, the duration of viral shedding in the control group was longer than 14 days. In the pathological section of the brain tissue, only a small amount of microglia and lymphocytes appeared in both oral group and the injection group; on the contrary, glial cell nodules, vascular cuff, satellite and neuronophagia phenomena were observed in the brain tissue of the control group. The tonsillar recess in the control group contained a large number of exfoliated crypt epithelial cells, white blood cells and inflammatory exudates and there was a growing oat cell. These results indicate that this recombinant attenuated strain orally immunized can produce an effective immune response against PRV, but does not induce high levels of PCV2 antibodies. This study provides a new direction for  PRV vaccine development and more options for immune pathway.

参考文献/References:

[1]Mettenleiter T C. Immunobiology of pseudorabies (Aujeszky’s Disease) [J]. Vet Immunol Immunopathol, 1996, 54(1-4): 221- 229.
[2]陈焕春,何启盖主编. 伪狂犬病[M]. 第1版. 北京:中国农业出版社,2015.
[3]童武,郑浩,单同领,等. 伪狂犬病毒变异株(JS-2012)对仔猪的致病性研究[J]. 中国动物传染病学报,2014,(5):10-14.
[4]张明辉,刘洋,库旭钢,等. 伪狂犬病病毒HNX株增殖特性及其致病性的研究[J]. 中国兽医科学,2015,45(12):1247-1253.
[5]郝飞,汤德元,李春燕,等. 伪狂犬病病毒Guizhou-DY株的分离鉴定及致病性研究[J]. 畜牧兽医学报,2013,44(11):1851-1856.
[6]Wu Rui, Bai Chao-yong, Sun Jin-zhong, et al. Emergence of virulent pseudorabies virus infection in northern China [J]. J Vet Sci, 2013, 14(3): 363-365.
[7]Maresch C, Lange E, Teifke J P, et al. Oral immunization of wild boar and domestic pigs with attenuated live vaccine protects against Pseudorabies virus infection [J]. Vet Microbiol, 2012, 161(1-2): 20-25.
[8]Monger V R, Stegeman J A, Dukpa K, et al. Evaluation of Oral Bait Vaccine Efficacy Against Classical Swine Fever in Village Backyard Pig Farms in Bhutan [J]. Transbound Emerg Dis, 2016, 63(6): e211.
[9]Liang Xun, Sun Le-qiang, Yu Teng, et al. A CRISPR/Cas9 and Cre/Lox system-based express vaccine development strategy against re-emerging Pseudorabies virus [J]. Sci Rep, 2016, 6:19176.
[10]张明辉,库旭钢,余腾,等. 猪伪狂犬病病毒gD基因荧光定量PCR检测方法的建立[C]. 中国畜牧兽医学会2014年学术年会,2014.
[11]何启盖,童光志,杨汉春,等. 猪伪狂犬病流行病学特征、净化技术及其应用示范[J]. 中国畜牧杂志,2015,51(24):68-74.
[12]谭鑫,库旭钢,于学祥,等. IFN-γ-ELISpot方法用于评价猪伪狂犬病疫苗免疫效果[J]. 畜牧兽医学报,2017,48(10):1939-1948.
(本文编辑:彭永刚;英文编辑:王玉娥)

相似文献/References:

[1]梁化春,齐景伟,刘淑英*,等.绵羊肺腺瘤病的病理学及RT-PCR诊断 [J].中国预防兽医学报,2009,(06):443.
 LIANG Hua-chun,QI Jing-wei,LIU Shu-ying*,et al.The pathological and RT-PCR diagnosis ofsheep pulmonary adenomatosis [J].Chinese journal of preventive veterinary medicine,2009,(12):443.
[2]刘合义,孙留霞,王进轶,等.牛冠状病毒重组N蛋白间接ELISA检测方法的建立 [J].中国预防兽医学报,2009,(08):618.
 LIU He-yi,SUN Liu-xia,WANG Jin-yi,et al.Development of an indirect ELISA for the detectionof Bovine coronavirus using recombinant N protein [J].Chinese journal of preventive veterinary medicine,2009,(12):618.
[3]秦玉寅,王雪峰,韦华冕,等.马传染性贫血病毒弱毒疫苗gp90多样性及其对病毒体外增殖的影响[J].中国预防兽医学报,2014,(03):182.[doi:10.3969/j.issn.1008-0589.2014.03.04]
 QIN Yu-yin,WANG Xue-feng,WEI Hua-mian,et al.gp90 genetic diversity of EIAV attenuated vaccine and its impact on EIAV’s proliferation in vitro[J].Chinese journal of preventive veterinary medicine,2014,(12):182.[doi:10.3969/j.issn.1008-0589.2014.03.04]
[4]刘 畅,李 磊,于立新,等.稳定表达绵羊肺腺瘤病毒受体Hyal-2蛋白细胞系的建立[J].中国预防兽医学报,2014,(03):187.[doi:10.3969/j.issn.1008-0589.2014.03.05]
 LIU Chang,LI Lei,YU Li-xin,et al. Development of a cell line stably expressing Jaagsiekte retrovirus receptor Hyal-2 protein[J].Chinese journal of preventive veterinary medicine,2014,(12):187.[doi:10.3969/j.issn.1008-0589.2014.03.05]
[5]冷青文,李志远,鲁海富,等.盘羊体内绵羊肺炎支原体的分离和鉴定[J].中国预防兽医学报,2014,(03):197.[doi:10.3969/j.issn.1008-0589.2014.03.07]
 LENG Qing-wen,LI Zhi-yuan,LU Hai-fu,et al.Isolation and identification of Mycoplasma ovipneumoniae in Argali[J].Chinese journal of preventive veterinary medicine,2014,(12):197.[doi:10.3969/j.issn.1008-0589.2014.03.07]
[6]程子英,高 洋,王 铭,等.弓形虫酵母双杂交cDNA文库构建及与AMA1羧基端相互作用蛋白的筛选[J].中国预防兽医学报,2014,(03):201.[doi:10.3969/j.issn.1008-0589.2014.03.08]
 CHENG Zi-ying,GAO Yang,WANG Ming,et al. Construction of yeast two-hybrid cDNA library of Toxoplasma gondii and screen of AMA1 c-terminal interacting proteins[J].Chinese journal of preventive veterinary medicine,2014,(12):201.[doi:10.3969/j.issn.1008-0589.2014.03.08]
[7]韩启灿,霍光华*,罗桂祥,等.野生冬菇产病原拮抗物的工艺参数确定[J].中国预防兽医学报,2014,(03):208.[doi:10.3969/j.issn.1008-0589.2014.03.10]
 HAN Qi-can,HUO Guang-hua*,LUO Gui-xiang,et al.The fermentation process of anti-pathogenic bacteria substances produced by a wild Flammulina ssp[J].Chinese journal of preventive veterinary medicine,2014,(12):208.[doi:10.3969/j.issn.1008-0589.2014.03.10]
[8]安 伟,肖 雨*,张崇文,等.鳜鱼传染性脾肾坏死病毒TaqMan荧光定量PCR检测方法的建立[J].中国预防兽医学报,2014,(03):214.[doi:10.3969/j.issn.1008-0589.2014.03.11]
 AN Wei,XIAO Yu*,ZHANG Chong-wen,et al. Establishment of a TaqMan qPCR assay for detecting the infectious spleen and kidney necrosis virus of Siniperca chuatsi[J].Chinese journal of preventive veterinary medicine,2014,(12):214.[doi:10.3969/j.issn.1008-0589.2014.03.11]
[9]胡 骑,何于雯,信爱国*,等.口蹄疫病毒实时定量RT-PCR方法的建立及初步应用[J].中国预防兽医学报,2014,(03):218.[doi:10.3969/j.issn.1008-0589.2014.03.12]
 HU Qi,HE Yu-wen,XIN Ai-guo*,et al.Development and application of a real-time RT-PCR assay for detection of foot-and-mouth disease virus[J].Chinese journal of preventive veterinary medicine,2014,(12):218.[doi:10.3969/j.issn.1008-0589.2014.03.12]
[10]周 兵,李天鹤,李 宁,等.抗鸡传染性法氏囊病病毒单链抗体库的构建及其中和抗体的筛选[J].中国预防兽医学报,2014,(03):227.[doi:10.3969/j.issn.1008-0589.2014.03.14]
 ZHOU Bing,LI Tian-he,LI Ning,et al.Construction and screening of an anti-IBDV scFv library[J].Chinese journal of preventive veterinary medicine,2014,(12):227.[doi:10.3969/j.issn.1008-0589.2014.03.14]

更新日期/Last Update: 2020-01-19