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新型鸭呼肠孤病毒弱毒S株在雏番鸭体内的增殖规律

胥焯然 程晓霞 郑敏 朱小丽 董慧 肖世峰 刘鸿威 曾显成 陈少莺 陈仕 龙

胥焯然,程晓霞,郑敏,等. 新型鸭呼肠孤病毒弱毒S株在雏番鸭体内的增殖规律 [J]. 福建农业学报,2023,38(9):1011−1016 doi: 10.19303/j.issn.1008-0384.2023.09.001
引用本文: 胥焯然,程晓霞,郑敏,等. 新型鸭呼肠孤病毒弱毒S株在雏番鸭体内的增殖规律 [J]. 福建农业学报,2023,38(9):1011−1016 doi: 10.19303/j.issn.1008-0384.2023.09.001
XU Z R, CHENG X X, ZHENG M, et al. Replication of Attenuated Strain S of Novel Duck Reovirus in Muscovy Ducklings [J]. Fujian Journal of Agricultural Sciences,2023,38(9):1011−1016 doi: 10.19303/j.issn.1008-0384.2023.09.001
Citation: XU Z R, CHENG X X, ZHENG M, et al. Replication of Attenuated Strain S of Novel Duck Reovirus in Muscovy Ducklings [J]. Fujian Journal of Agricultural Sciences,2023,38(9):1011−1016 doi: 10.19303/j.issn.1008-0384.2023.09.001

新型鸭呼肠孤病毒弱毒S株在雏番鸭体内的增殖规律

doi: 10.19303/j.issn.1008-0384.2023.09.001
基金项目: 福建省农业高质量发展超越“5511”协同创新工程项目(XTCXGC2021018、XTCXGC2021012);福建省农业科学院科技创新团队建设项目(CXTD2021034)
详细信息
    作者简介:

    胥焯然(1999 —),女,硕士研究生,主要从事病毒免疫学研究,E-mail:530147013@qq.com

    通讯作者:

    陈少莺(1962 —),女,研究员,主要从事兽医传染病防控研究,E-mail:chensy58@163.com

    陈仕龙(1979 —),男,博士,研究员,主要从事动物病毒病研究,E-mail:CSL6116@163.com

  • 中图分类号: S852.65

Replication of Attenuated Strain S of Novel Duck Reovirus in Muscovy Ducklings

  • 摘要:   目的  研究新型鸭呼肠孤病毒(Novel duck reovirus,NDRV)弱毒S株C30在雏番鸭血液中的病毒血症,口咽、泄殖腔的排毒规律及靶器官的带毒时间,了解弱毒S株在雏番鸭体内的增殖规律和致弱机理。  方法  设计特异性引物,建立检测NDRV核酸的RT-qPCR方法;新型鸭呼肠孤病毒弱毒S株第30代(NDRV-S-C30)腿部肌肉注射2日龄雏番鸭,在免疫后1、2、3、4、6、8、10、12、14 d(Days post vaccination,dpv)采集其血液、肝脏、脾脏、泄殖腔分泌液和口咽液,用RT-qPCR 方法检测NDRV在血液、口咽、泄殖腔及靶器官的增殖能力及带毒时间。  结果  建立了检测NDRV 核酸的特异性RT-qPCR方法,使用该方法检测NDRV弱毒攻毒雏鸭的口咽和泄殖腔排毒时间分别为2~8 d和2~10 d,排毒高峰期为4~6 d;病毒血症时间为1~4 d,其中1~2 d 为病毒血症高峰期;弱毒在肝脏的带毒时间为3~6 d,在脾脏的带毒时间为1~8 d。  结论  NDRV-S-C30弱毒株免疫雏番鸭后可通过口腔和泄殖腔向外界排毒,仅能引起较弱的病毒血症反应,且在肝脏中的增殖能力弱,丧失了对易感靶器官造成病理损伤的能力。明确了NDRV 弱毒S株的排毒规律及病毒血症时间,为建立弱毒免疫效力评价方法奠定了基础。
  • 图  1  qPCR引物特异性验证

    A:RT-PCR 扩增电泳图;M为DNA DL 2000 相对分子质量标准,1为pMD18T-NDRV质粒,2为NDRV-S-C30,3~9分别为GPV、MDPV、MDRV、GRV、DHAV-3、DAdV-B2和DTMUV,10:DEPC水。B:RT-qPCR扩增曲线;1为pMD18T-NDRV质粒,2为NDRV-S-C30,3~9分别为GPV、MDPV、MDRV、GRV、DHV-3、DAdV-B2和DTMUV,10:DEPC水。

    Figure  1.  Specificity of qPCR primers

    A: RT-PCR amplification; M: DL2000 DNA marker; 1: pMD18T-NDRV plasmid; 2: NDRV-S-C30; 3–9: GPV, MDPV, MDRV, GRV, DHV-3, DAdV-B2, and DTMUV, respectively; 10: DEPC water; B: RT-qPCR amplification; 1: pMD18T-NDRV plasmid; 2: NDRV-S-C30; 3–9: GPV, MDPV, MDRV, GRV, DHV-3, DAdV-B2, and DTMUV, respectively; 10: DEPC water.

    图  2  RT-qPCR和普通RT-PCR的敏感性比较

    A:荧光定量RT-PCR;1~9分别为1.89×109~1.89×101 拷贝·μL−1,10为DEPC水;B:普通RT-PCR;M:DNA DL 2000 相对分子质量标准,1~9分别为1.89×109~1.89×101拷贝·μL−1,10为DEPC水。

    Figure  2.  Sensitivity of conventional PCR and RT-qPCR assays in detecting NDRV

    A: RT-qPCR; 1–9: 1.89×109−1.89×101 copies·μL−1; 10: DEPC water; B: conventional RT-PCR; M: DL2000 DNA marker; 1–9: 1.89×109−1.89×101 copies·μL−1, respectively; 10: DEPC water.

    表  1  RT-qPCR检测NDRV-S-C30免疫雏番鸭的排毒规律

    Table  1.   RT-qPCR detection of viral shedding in Muscovy ducklings after NDRV-S-C30 vaccination

    采集部位
    Sampling site
    检测结果(阳性样品/检测样品)
    Detection result(positive samples/detected samples)
    1 d2 d3 d4 d6 d8 d10 d12 d14 d
    喉 Throat免疫组 Vaccination group0/32/32/33/33/31/30/30/30/3
    对照组 Control group0/20/20/20/20/20/20/20/20/2
    泄殖腔 Cloaca免疫组 Vaccination group0/32/32/33/33/31/31/30/30/3
    对照组 Control group0/20/20/20/20/20/20/20/20/2
    下载: 导出CSV

    表  2  NDRV-S-C30免疫后雏番鸭血液RT-qPCR检测结果

    Table  2.   RT-qPCR detection on blood of Muscovy ducklings after NDRV-S-C30 vaccination

    采集部位
    Sampling site
    处理组
    Treatment group
    检测结果(阳性样品/检测样品)
    Detection result(psositive samples/detected samples)
    1 d2 d3 d4 d6 d8 d10 d12 d14 d
    血清 Serum免疫组 Vaccination group2/32/31/31/30/30/30/30/30/3
    对照组 Control group0/20/20/20/20/20/20/20/20/2
    下载: 导出CSV

    表  3  NDRV-S-C30在雏番鸭肝、脾中的RT-qPCR检测结果

    Table  3.   RT-qPCR detection on liver and spleen of Muscovy ducklings after NDRV-S-C30 vaccination


    采集部位
    Sampling site
    处理组
    Treatment group
    检测结果(阳性样品/检测样品)
    Detection result(positive samples/detected samples)
    1 d2 d3 d4 d6 d8 d10 d12 d14 d
    肝脏 Liver免疫组 Vaccination group0/30/32/31/31/30/30/30/30/3
    对照组 Control group0/20/20/20/20/20/20/20/20/2
    脾脏 Spleen免疫组 Vaccination group2/31/33/33/31/31/30/30/30/3
    对照组 Control group0/20/20/20/20/20/20/20/20/2
    下载: 导出CSV
  • [1] 陈少莺, 陈仕龙, 林锋强, 等. 新型鸭呼肠孤病毒的分离与鉴定 [J]. 病毒学报, 2012, 28(3):224−230.

    CHEN S Y, CHEN S L, LIN F Q, et al. The isolation and identification of novel duck reovirus [J]. Chinese Journal of Virology, 2012, 28(3): 224−230.(in Chinese)
    [2] 陈少莺, 陈仕龙, 林锋强, 等. 一种新的鸭病(暂名鸭出血性坏死性肝炎)病原学研究初报 [J]. 中国农学通报, 2009, 25(16):28−31.

    CHEN S Y, CHEN S L, LIN F Q, et al. The primary study of pathogen of duck hemorrhagic-necrotic hepatitis [J]. Chinese Agricultural Science Bulletin, 2009, 25(16): 28−31.(in Chinese)
    [3] 陆新浩, 陈秋英, 刘鸿, 等. 番鸭出血性坏死性肝炎的诊断 [J]. 中国家禽, 2015, 37(15):65−66.

    LU X H, CHEN Q Y, LIU H, et al. Diagnosis of hemorrhagic necrotizing hepatitis in Muscovy ducks [J]. China Poultry, 2015, 37(15): 65−66.(in Chinese)
    [4] 马丰英, 丛雁方, 于春梅, 等. 新型鸭呼肠孤病毒病的研究进展 [J]. 中国家禽, 2022, 44(9):92−100.

    MA F Y, CONG Y F, YU C M, et al. Research progress on novel duck reovirusis disease [J]. China Poultry, 2022, 44(9): 92−100.(in Chinese)
    [5] VARGA-KUGLER R, MARTON S, THUMA Á, et al. Candidate 'avian orthoreovirus B': An emerging waterfowl pathogen in Europe and Asia? [J]. Transboundary and Emerging Diseases, 2022, 69(5): e3386−e3392.
    [6] 陈仕龙, 陈少莺, 程晓霞, 等. 新型鸭呼肠孤病毒分离株的致病性研究 [J]. 西北农林科技大学学报(自然科学版), 2010, 38(4):14−18.

    CHEN S L, CHEN S Y, CHENG X X, et al. The study on the pathogenicity of new type duck reovirus [J]. Journal of Northwest A & F University (Natural Science Edition), 2010, 38(4): 14−18.(in Chinese)
    [7] LI N, HONG T Q, WANG Y, et al. The pathogenicity of novel duck reovirus in Cherry Valley ducks [J]. Veterinary Microbiology, 2016, 192: 181−185. doi: 10.1016/j.vetmic.2016.07.015
    [8] WANG S, LIN F Q, CHENG X X, et al. The genomic constellation of a novel duck reovirus strain associated with hemorrhagic necrotizing hepatitis and splenitis in Muscovy ducklings in Fujian, China [J]. Molecular and Cellular Probes, 2020, 53: 101604. doi: 10.1016/j.mcp.2020.101604
    [9] ZHANG X L, SHAO J W, LI X W, et al. Molecular characterization of two novel reoviruses isolated from Muscovy ducklings in Guangdong, China [J]. BMC Veterinary Research, 2019, 15(1): 143. doi: 10.1186/s12917-019-1877-x
    [10] WANG S, CHEN S L, CHENG X X, et al. Sequence and phylogenetic analysis of M-class genome segments of novel duck reovirus NP03 [J]. Canadian Journal of Veterinary Research, 2015, 79(2): 147−150.
    [11] KONG J, SHAO G M, ZHANG Y K, et al. Molecular characterization, complete genome sequencing, and pathogenicity of novel duck reovirus from south coastal area in China [J]. Poultry Science, 2023, 102(8): 102776. doi: 10.1016/j.psj.2023.102776
    [12] YANG H H, ZHANG W D, WANG M H, et al. Characterization and pathogenicity evaluation of recombinant novel duck reovirus isolated from Southeast China [J]. Frontiers in Veterinary Science, 2023, 10: 1124999. doi: 10.3389/fvets.2023.1124999
    [13] 华炯钢, 叶伟成, 刘可姝, 等. 新型鸭呼肠孤病毒弱毒株在雏番鸭体内的分布和排毒规律 [J]. 浙江农业科学, 2021, 62(11):2295−2298.

    HUA J G, YE W C, LIU K S, et al. Distribution and shedding pattern of novel duck reovirus attenuated strain in vaccinated Muscovy ducklings [J]. Journal of Zhejiang Agricultural Sciences, 2021, 62(11): 2295−2298.(in Chinese)
    [14] 林锋强, 程晓霞, 陈仕龙, 等. 新型鸭呼肠孤病毒在番鸭体内分布和排毒规律 [J]. 中国家禽, 2018, 40(3):53−55.

    LIN F Q, CHENG X X, CHEN S L, et al. Distribution and detoxification of a new duck reovirus in Muscovy ducks [J]. China Poultry, 2018, 40(3): 53−55.(in Chinese)
    [15] 沈丽雅, 潘群兴, 卢凤英, 等. 基于P18蛋白基因的新型鸭呼肠孤病毒荧光定量RT-PCR检测方法的建立 [J]. 江苏农业学报, 2021, 37(6):1481−1487.

    SHEN L Y, PAN Q X, LU F Y, et al. Establishment of SYBR Green I fluorescence quantitative RT-PCR assay for detection of novel duck reovirus based on the gene encoding P18 protein [J]. Jiangsu Journal of Agricultural Sciences, 2021, 37(6): 1481−1487.(in Chinese)
    [16] WU X, HUANG S, WANG M, et al. A novel live attenuated duck Tembusu virus vaccine targeting N7 methyltransferase protects ducklings against pathogenic strains. Veterinary Research, 2023, 54(1): 47.
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出版历程
  • 收稿日期:  2023-07-07
  • 修回日期:  2023-08-10
  • 网络出版日期:  2023-09-19
  • 刊出日期:  2023-09-28

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