新城疫病毒M蛋白核定位信号突变降低病毒的致病性

doi:10.11843/j.issn.0366-6964.2018.12.018

摘要/Abstract

摘要：

本研究旨在探讨M蛋白核定位信号（NLS）突变对新城疫病毒（NDV）致病性的影响。将M蛋白NLS突变体病毒和亲本病毒分别感染4周龄SPF鸡，观察鸡的临床症状和病理变化，检测鸡喉头和泄殖腔排毒情况以及不同组织中的病毒含量，并对免疫器官进行显微病变观察，以及分析免疫器官中M蛋白的表达量和相关细胞因子的表达变化。结果表明，鸡感染亲本病毒后产生典型的新城疫临床症状和病理变化，喉头和泄殖腔持续排毒且排毒量高，试验鸡的存活率为0%；而鸡感染突变体病毒后的临床症状和病理变化轻微，喉头和泄殖腔排毒时间延迟且排毒量低，试验鸡的存活率为70%。组织病毒含量测定结果表明亲本病毒能在不同组织中复制，尤其以免疫器官（脾、胸腺和法氏囊）和气管中的病毒含量高；而突变体病毒仅在免疫器官和气管中复制，且病毒含量低。病理组织学观察和Western blotting检测结果表明，亲本病毒能造成严重的免疫器官损伤且M蛋白表达量高，而突变体病毒则未引起明显的病理变化且M蛋白表达量低。此外，突变体病毒感染引起的免疫器官中IL-1β、IL-6、IL-10、IFN-β和IFN-λ细胞因子的表达水平要明显低于亲本病毒，说明M蛋白NLS突变降低了NDV诱发的细胞免疫应答反应。本研究首次证实M蛋白NLS突变可显著降低NDV对鸡的致病力，这为深入研究M蛋白细胞核定位在NDV致病性中的作用奠定了基础。

Abstract:

The aim of this study was to investigate the effect of nuclear localization signal (NLS) mutation in the M protein on the pathogenicity of Newcastle disease virus (NDV). The parental virus and the M/NLS mutant virus were used to infect 4-week-old SPF chickens. The clinical symptom and pathological change, the virus shedding in larynx and cloaca, the virus titers in the tissues, the microscopic lesions, the expression change of M protein and the cytokine genes in immune organs of virus-infected chickens were examined. The results showed that chickens infected with the parental virus exhibited typical ND clinical symptom and pathological change, and had continuous and high virus shedding in the larynx and cloaca. By contrast, chicken infected with the mutant virus showed slight clinical symptom and pathological change, and had delayed and low virus shedding. The survival rate of parental and mutant virus-infected group was 0% and 70%, respectively. Compared to the replication ability and virus titers of the parental virus in different tissues, the mutant virus only replicated in the immune organs and trachea and simultaneously had very low virus titers. In addition, the mutant virus caused unconspicuous pathological change and low expression of M protein in the immune organs in comparison to the parental virus, and also reduced expression level of IL-1β, IL-6, IL-10, IFN-β and IFN-λ in immune organs, indicating that M/NLS mutation significantly reduced the cellular immunologic response induced by NDV. This study revealed for the first time the attenuated pathogenicity of M/NLS mutant NDV, and provided useful information for further investigating the nuclear localization function of M protein in NDV pathogenicity.

中图分类号:

S852.659.5

段志强, 邓珊珊, 袁超, 高洪波, 嵇辛勤, 赵佳福, 阮涌. 新城疫病毒M蛋白核定位信号突变降低病毒的致病性[J]. 畜牧兽医学报, 2018, 49(12): 2690-2697.

DUAN Zhi-qiang, DENG Shan-shan, YUAN Chao, GAO Hong-bo, JI Xin-qin, ZHAO Jia-fu, RUAN Yong. Nuclear Localization Signal Mutation in the M Protein Attenuates the Pathogenicity of Newcastle Disease Virus[J]. ACTA VETERINARIA ET ZOOTECHNICA SINICA, 2018, 49(12): 2690-2697.

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参考文献

[1] DUAN Z Q, XU H Q, JI X Q, et al. Recombinant Newcastle disease virus-vectored vaccines against human and animal infectious diseases[J]. Future Microbiol, 2015, 10(8):1307-1323.
[2] 段志强,胡顺林,刘秀梵.新城疫病毒与其它副黏病毒基质蛋白功能的比较[J].微生物学报, 2016, 56(7):1070-1078.
DUAN Z Q, HU S L, LIU X F. Function comparison of the matrix protein between Newcastle disease virus and other paramyxoviruses-a review[J]. Acta Microbiologica Sinica, 2016, 56(7):1070-1078. (in Chinese)
[3] HARRISON M S, SAKAGUCHI T, SCHMITT A P. Paramyxovirus assembly and budding:Building particles that transmit infections[J]. Int J Biochem Cell Biol, 2010, 42(9):1416-1429.
[4] COLEMAN N A, PEEPLES M E. The matrix protein of Newcastle disease virus localizes to the nucleus via a bipartite nuclear localization signal[J].Virology, 1993, 195(2):596-607.
[5] DUAN Z Q, XU H X, JI X Q, et al. Importin α5 negatively regulates importin β1-mediated nuclear import of Newcastle disease virus matrix protein and viral replication and pathogenicity in chicken fibroblasts[J]. Virulence, 2018,9(1):783-803.
[6] SANCHEZ A, GUERRERO-JUAREZ C F, RAMIREZ J, et al. Nuclear localized Influenza nucleoprotein N-terminal deletion mutant is deficient in functional vRNP formation[J]. Virol J, 2014,11:155.
[7] LI C X, WEI H C, YU L P, et al. Nuclear localization of the p17 protein of avian reovirus is correlated with autophagy induction and an increase in viral replication[J]. Arch Virol, 2015, 160(12):3001-3010.
[8] CHEN T T, DONG Z Q, HU N, et al. Baculovirus LEF-11 nuclear localization signal is important for viral DNA replication[J]. Virus Res, 2017,238:133-140.
[9] AUDSLEY M D, JANS D A, MOSELEY G W. Roles of nuclear trafficking in infection by cytoplasmic negative-strand RNA viruses:Paramyxoviruses and beyond[J]. J Gen Virol, 2016, 97(10):2463-2481.
[10] REED L J, MUENCH H. A simple method of estimating fifty percent endpoints[J].Am J Epidemiol, 1938, 27(3):493-497.
[11] MORI Y, OKABAYASHI T, YAMASHITA T, et al. Nuclear localization of Japanese encephalitis virus core protein enhances viral replication[J]. J Virol, 2005, 79(6):3448-3458.
[12] LEE C, HODGINS D, CALVERT J G, et al. Mutations within the nuclear localization signal of the porcine reproductive and respiratory syndrome virus nucleocapsid protein attenuate virus replication[J]. Virology, 2006, 346(1):238-250.
[13] ROWE C L, WAGSTAFF K M, OKSAYAN S, et al. Nuclear trafficking of the rabies virus interferon antagonist P-Protein is regulated by an importin-binding nuclear localization sequence in the C-terminal domain[J]. PLoS One, 2016, 11(3):e0150477.
[14] GHILDYAL R, HO A, JANS D A. Central role of the respiratory syncytial virus matrix protein in infection[J]. FEMS Microbiol Rev, 2006, 30(5):692-705.
[15] IWASAKI M, TAKEDA M, SHIROGANE Y, et al. The matrix protein of measles virus regulates viral RNA synthesis and assembly by interacting with the nucleocapsid protein[J]. J Virol, 2009, 83(20):10374-10383.
[16] DUAN Z Q, LI J, ZHU J, et al. A single amino acid mutation,R42A,in the Newcastle disease virus matrix protein abrogates its nuclear localization and attenuates viral replication and pathogenicity[J]. J Gen Virol, 2014, 95(5):1067-1073.
[17] HUANG Z H, PANDA A, ELANKUMARAN S, et al. The hemagglutinin-neuraminidase protein of Newcastle disease virus determines tropism and virulence[J]. J Virol, 2004, 78(8):4176-4184.
[18] KRISTEEN-TEO Y W, YEAP S K, TAN S W, et al. The effects of different velogenic NDV infections on the chicken bursa of Fabricius[J]. BMC Vet Res, 2017, 13(1):151.
[19] ECCO R, BROWN C, SUSTA L, et al. In vivo transcriptional cytokine responses and association with clinical and pathological outcomes in chickens infected with different Newcastle disease virus isolates using formalin-fixed paraffin-embedded samples[J].Vet Immunol Immunopathol, 2011, 141(3-4):221-229.
[20] NAKAMURA K, OHTSU N, NAKAMURA T, et al. Pathologic and immunohistochemical studies of Newcastle disease (ND) in broiler chickens vaccinated with ND:Severe nonpurulent encephalitis and necrotizing pancreatitis[J]. Vet Pathol, 2008, 45(6):928-933.
[21] RUE C A, SUSTA L, CORNAX I, et al. Virulent Newcastle disease virus elicits a strong innate immune response in chickens[J]. J Gen Virol, 2011, 92(4):931-939.
[22] HU Z L, HU J, HU S L, et al. High levels of virus replication and an intense inflammatory response contribute to the severe pathology in lymphoid tissues caused by Newcastle disease virus genotype VⅡd[J]. Arch Virol, 2015, 160(3):639-648.
[23] 程欣. 基因VⅡd亚型新城疫病毒感染LaSota疫苗免疫鸡引起的免疫器官损伤及其对疫苗免疫效果的影响[D]. 扬州:扬州大学, 2016.
CHENG X. Infections with genotype VⅡd Newcastle disease virus induces damages of immuolymphoid organs in chickens vaccinated with LaSota and affects vaccination efficacies thereafter[D]. Yangzhou:Yangzhou University, 2016. (in Chinese)
[24] PENTECOST M, VASHISHT A A, LESTER T, et al. Evidence for ubiquitin-regulated nuclear and subnuclear trafficking among Paramyxovirinae matrix proteins[J]. PLoS Pathog, 2015, 11(3):e1004739.