,中国兽医药品监察所,国家/OIE猪瘟参考实验室,北京 100081The Impact of Classical Swine Fever and African Swine Fever on Pig Industry
WANG Qin,National/OIE Reference Laboratory for Classical Swine Fever, China Institute of Veterinary Drug Control, Beijing 100081通讯作者:
责任编辑: 林鉴非
收稿日期:2018-10-24接受日期:2018-10-25网络出版日期:2018-11-01
| 基金资助: |
Received:2018-10-24Accepted:2018-10-25Online:2018-11-01
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王琴. 猪瘟与非洲猪瘟对养猪业的重大冲击[J]. 中国农业科学, 2018, 51(21): 4143-4145 doi:10.3864/j.issn.0578-1752.2018.21.012
WANG Qin.
猪瘟（classical swine fever,CSF）是由猪瘟病毒（classical swine fever virus,CSFV）引起的猪的一种高度接触性传染性疾病,传染性强、死亡率高,是严重危害养猪业的重大传染病,是世界动物卫生组织（OIE）规定的必需报告的疫病,我国将其列为一类动物疫病,也是我国政府计划消灭的重大动物传染病之一。CSF呈全球分布,目前流行于大部分亚洲（包括中国）、中美洲和南美洲、部分东欧国家、加勒比和非洲国家,偶尔散发于部分中欧和西欧国家。根据OIE截止到2018年5月的报告,已有35个国家、巴西部分区域和哥伦比亚部分区域宣布为无CSF的国家和地区,但是今年的10月8日和9日分别在日本和巴西重新爆发猪瘟[1],根除CSF的国家主要是通过政府立法、阶段性的综合防控技术措施来根除CSF,特别是欧美国家CSF的根除技术为全世界动物疫病的根除树立了典范[1]。
猪和野猪是能感染CSFV的唯一宿主,CSFV属于黄病毒科（Flaviviridae）瘟病毒属（Pestivirus）的成员之一,病毒基因组大小约为12.3kb,含有一个大的开放阅读框架（ORF）,编码一个由3 898个氨基酸残基组成的多聚蛋白,此多聚蛋白经病毒和宿主细胞的酶作用,形成4个成熟的结构蛋白（C、Erns、E1、E2）以及至少8个非结构蛋白（Npro、P7、NS2、NS3、NS4A、NS4B、NS5A、NS5B）,各蛋白的排列顺序从N端到C端依次为NH2-Npro-C-Erns-E1- E2-P7-NS2- NS3-NS4A-NS4B-NS5A-NS5B-COOH[2]。现已明确了CSFV大部分基因片段的分子结构与功能,其中5′结构蛋白编码区的基因定位已比较清楚。在病毒复制过程中,CSFV基因组的5′端非编码区（5′non-translated region, 5′NTR）对病毒基因组复制是必需的,并且该区域能够抑制内部核糖体进入位点（internal ribosome entry site, IRES）介导的病毒蛋白翻译,其余非结构蛋白Npro、P7、NS2、NS3、NS4A、NS4B、NS5A、NS5B也是病毒复制所必需的[3]。通过嵌合CSFV致病性研究,证实3'UTR12nt的插入对病毒复制和毒力有重要影响,与减毒有重要关系,揭示了我国猪瘟兔化弱毒疫苗致弱的分子机制[4]。
CSFV分子流行病学研究对阐明病毒的遗传变异规律、掌握流行现状、追溯疫情来源、监测疫苗有效性具有十分重要的科学意义。国际上将CSFV分为3个基因型（Ⅰ型、Ⅱ型和Ⅲ型）,11个基因亚型（1.1、1.2、1.3、1.4、2.1、2.2、2.3、3.1、3.2、3.3和3.4）。通过研究我国自1979年到2016年的875条（包括台湾省14条）E2基因主要抗原区域序列（190 bp）,中国流行的CSFV分属为Ⅰ、Ⅱ和Ⅲ三个基因型,1.1、2.1、2.2、2.3 和3.4五个基因亚型,其中74.24%的流行毒株属于基因Ⅱ型,为优势基因型,查清了中国CSFV分子流行病学的本底;研究还表明中国CSFV流行株基因组在38年间处于稳定状态,结合动物免疫保护试验证实了现用C-株疫苗对我国流行毒株（Ⅰ和Ⅱ型）能有效保护,来自英国APHA的数据显示C-株疫苗对基因Ⅲ型也能提供完全保护[5],否定了多年认为“由野毒株变异引起免疫失败”的学术争端,为我国继续使用C-株疫苗进行全面免疫提供了重要科学依据;研究还表明,到目前为止,中国大陆一直没有监测到基因Ⅲ型的传入与流行,但周边国家及我国台湾省存在基因Ⅲ型。因此,要加大对周边国家及地区的新亚型流行毒株传入我国大陆的监测力度[3]。我国是CSF染疫国家,疫苗免疫是控制CSF广泛流行主要手段,多年来我国持续实施CSF强制免疫,遏制了CSF的大规模暴发流行。随之而来便出现了非典型CSF或持续性感染CSF,这是造成我国CSF长期持续存在和散发流行恶性循环的主要根源,这种持续带毒现象对CSF的控制乃至净化产生了严重的影响,使得CSF难以根除。通过构建CSF慢性感染动物模型,系统研究了持续感染CSF的临床症候、病理学、组织学、病毒载量、外周血细胞及免疫相关因子转录的变化。持续性感染中CSFV的主要复制场所为淋巴器官,且对淋巴器官具有持续性的、不可恢复性的损伤,对其他组织的损伤具有可恢复性;扁桃体依然是检测慢性感染猪的最敏感、方便的组织之一;系统分析了CSFV在逃避宿主免疫因子作用和形成特异性抗体中起关键作用细胞因子,为探索细胞因子作用奠定了基础,为阐述CSF持续性感染和免疫的机制提供科学的理论依据[6,7]。
我国政府高度重视CSF的防控并取得了重要成效,根据农业部《兽医公报》2010年到2017年的数据显示,我国CSF新发病例和死亡数已大幅下降至767例和273例。2016年7月农业部财政部联合印发了《关于调整完善动物疫病防控支持政策的通知》,自2017年起从中央层面退出对CSF的财政补助。2017年3月农业部印发《国家猪瘟防治指导意见（2017- 2020年）》,要求各地继续对生猪实施CSF全面免疫,进一步落实各项综合防控措施, 有效控制、逐步消灭CSF,防控政策的改变标志着正式将CSF免疫净化付诸行动,这是我国CSF防控政策历史性的重大转变。
本专题针对我国CSF研究的新进展和防控政策的变革时期,集中报道了采用ViewRNA ISH新技术对CSFV中等致病力毒株感染后免疫器官、消化系统、呼吸系统等各组织中病毒RNA的分布动态研究成果,对阐明持续感染的致病机理具有重要的科学意义[8];针对CSFV致病机制十分复杂的状况,本专题还报道了通过筛选可能与CSFV感染相关的miRNA及其功能研究,从宿主细胞的角度来研究CSFV与细胞相互作用的关系,为研究CSFV的致病机理提供有价值的科学资料[9];还特别介绍了CSF的全球流行态势,探讨了我国CSF净化的重要性及有利和不利条件,深入分析了我国CSF净化的成本及效益,总结了欧盟等国家净化CSF的成功经验,并对我国CSF净化思路和方案进行了探讨[10]。
针对近来首次传入我国,随后在我国13个省爆发的42起非洲猪瘟（African swine fever,ASF）疫情,引发兽医行业高度关注,对我国养猪业构成巨大威胁[11]。因此本专题也报道其相关知识,ASF是由非洲猪瘟病毒（African swine fever virus,ASFV）引起猪的一种急性、热性、高度传染性疾病,病程短,死亡率高。虽然ASF与CSF均不感染人,但均可对养猪业造成毁灭性打击,有着重要的社会经济学意义,与CSF一样,ASF也被OIE列为必须报告的动物疫病,在我国也将其列为一类动物疫病,然而在今年8月3日传入我国之前,ASF被列为外来病[12]。引发ASF的病原是一个与CSFV完全不同的ASFV,它是一个复杂的二十面体DNA病毒,属于ASFV相关病毒科（Asfarviridae）、ASFV病毒属（Asfivirus）的唯一成员,有24个基因型,其基因组是猪瘟病毒基因组的15倍[13,14]。
CSF与ASF均通过与传染源直接接触和间接接触感染所有年龄家猪和野猪,临床症状类似,鉴别难度大,一旦感染并传播均会造成猪只的严重发病。但是脾脏极度肿大至5—10倍,严重梗死,质脆易碎,是ASF与CSF的重要鉴别特征,核酸检测也是鉴别两个病的重要方法[15]。ASF作为外来病首次传入我国,其控制措施与CSF也完全不同。对于CSF的控制,我国拥有享誉世界的免疫原性和安全性都非常优秀的C-株疫苗用于CSF的免疫控制乃至净化;而对于ASF,目前国内外尚无可用的有效疫苗,各国对于该病的处置均采用及时准确的监测、严格有效的封锁和扑杀等措施来进行根除[16]。所以,本专题全面报道了ASF的流行病学、诊断和疫苗等方面最新研究进展及防控挑战的综述,以期为我国非洲猪瘟的防控提供参考[17]。
参考文献 原文顺序
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被引期刊影响因子
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DOI:10.1002/(SICI)1096-9071(199911)59:3<412::AID-JMV23>3.0.CO;2-LURLPMID:10516051 [本文引用: 1]
Defective interfering particles (DIs) of bovine viral diarrhea virus (BVDV) have been identified and shown to be cytopathogenic (cp) in the presence of noncytopathogenic (noncp) helper virus. Moreover, a subgenomic (sg) RNA corresponding in its genome structure to one of those BVDV DIs (DI9) was replication competent in the absence of helper virus. We report here that an sg BVDV replicon which encodes from the viral proteins only the first three amino acids of the autoprotease N(pro) in addition to nonstructural (NS) proteins NS3 to NS5B replicates autonomously and also induces lysis of its host cells. This demonstrates that the presence of a helper virus is not required for the lysis of the host cell. On the basis of two infectious BVDV cDNA clones, namely, BVDV CP7 (cp) and CP7ins- (noncp), bicistronic replicons expressing proteins NS2-3 to NS5B were established. These replicons express, in addition to the viral proteins, the reporter gene encoding beta-glucuronidase; the release of this enzyme from transfected culture cells was used to monitor cell lysis. Applying these tools, we were able to show that the replicon derived from CP7ins- does not induce cell lysis. Accordingly, neither N(pro) nor any of the structural proteins are necessary to maintain the noncp phenotype. Furthermore, these sg RNAs represent the first pair of cp and noncp replicons which mimic complete BVDV CP7 and CP7ins- with respect to cytopathogenicity. These replicons will facilitate future studies aimed at the determination of the molecular basis for the cytopathogenicity of BVDV.
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DOI:10.1016/j.virol.2008.01.008URLPMID:18279903 [本文引用: 1]
We report here the discovery of an attenuation mechanism of classic swine fever virus (CSFV) induced by introduction of a continuous 12-nt (CUUUUUUCUUUU) insertion in viral 3′ UTR. The 12-nt insertion sequence was first found in one attenuated vaccine strain HCLV (Hog Cholera Lapinized Virus) which did not exist in other CSFV strains. To address the function of the 12-nt insertion in viral replication and attenuation, we constructed and analyzed two chimeras stemmed from a highly virulent strain Shimen either with introduction of the 12-nt insertion in 3′ UTR or the replacement of viral 3′ UTR by the 3′ UTR of HCLV. We found that the two chimeras' maximum titers declined approximately 100-fold than their parental strain Shimen in PK15 cells. An animal experiment showed that the two chimeras were both dramatically attenuated in pigs. All the chimera-infected pigs survived infection and remained clinically normal with the exception of a transient fever while the 100% mortality was observed for the Shimen-infected pigs. In addition, the two chimeras can induce neutralization antibody to completely protect the pigs against lethal challenge with highly virulent CSFV, which was similar to the vaccine strain HCLV. These data demonstrate that the 12-nt insertion in 3′ UTR is sufficient for the attenuation of CSFV. Taken together, a novel attenuation mechanism of CSFV is found and may pave a way to further research for new attenuated vaccine.
DOI:10.1371/journal.pone.0029310URLPMID:3250419 [本文引用: 1]
Pre-emptive culling is becoming increasingly questioned as a means of controlling animal diseases, including classical swine fever (CSF). This has prompted discussions on the use of emergency vaccination to control future CSF outbreaks in domestic pigs. Despite a long history of safe use in endemic areas, there is a paucity of data on aspects important to emergency strategies, such as how rapidly CSFV vaccines would protect against transmission, and if this protection is equivalent for all viral genotypes, including highly divergent genotype 3 strains. To evaluate these questions, pigs were vaccinated with the Riemser C-strain vaccine at 1, 3 and 5 days prior to challenge with genotype 2.1 and 3.3 challenge strains. The vaccine provided equivalent protection against clinical disease caused by for the two challenge strains and, as expected, protection was complete at 5 days post-vaccination. Substantial protection was achieved after 3 days, which was sufficient to prevent transmission of the 3.3 strain to animals in direct contact. Even by one day post-vaccination approximately half the animals were partially protected, and were able to control the infection, indicating that a reduction of the infectious potential is achieved very rapidly after vaccination. There was a close temporal correlation between T cell IFN- responses and protection. Interestingly, compared to responses of animals challenged 5 days after vaccination, challenge of animals 3 or 1 days post-vaccination resulted in impaired vaccine-induced T cell responses. This, together with the failure to detect a T cell IFN- response in unprotected and unvaccinated animals, indicates that virulent CSFV can inhibit the potent antiviral host defences primed by C-strain in the early period post vaccination.
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DOI:10.3864/j.issn.0578-1752.2016.12.015URL [本文引用: 1]
【目的】为研究CSFV RNA在体外感染细胞中的分布及定位,建立了一种准确、敏感的RNA可视化原位杂交技术。【方法】本研究通过比对Gen Bank中公布的CSFV、BVDV和BDV全序列,避开BVDV和BDV的同源区,设计了CSFV RNA及内参基因β-actin的特异探针。以CSFV中等致病力毒株（He BHH1/95）为参考毒株,在PK15细胞中培养病毒,加入RNA可视化原位杂交的特异探针和相应试剂,采用荧光共聚焦显微镜进行成像观察。通过综合分析观测结果、荧光强度、重复性等因素,采用正交试验优化了对原位杂交过程中具有重要影响的蛋白酶K浓度和甲醛固定时间,建立了CSFV RNA可视化原位杂交技术,并与FAT方法比较该技术灵敏度;用我国目前流行的CSFV 1.1、2.1、2.2、2.3基因亚型及BVDV、PPV、PRV和PCV-2病毒进行特异性试验。最终,以CSFV强致病力毒株（SM）接种PK15细胞,病毒感染后0.5、1、3、6、8、10、14、18、24、36、48、72、96h（hours post inoculation,hpi）取样,每个时间点2个重复,采用CSFV RNA可视化原位杂交技术进行检测。为佐证病毒蛋白在细胞中的定位及分布,同时采用FAT方法对SM株E2蛋白在PK15细胞中的表达情况进行动态研究。【结果】采用该技术在荧光共聚焦显微镜下可观察到CSFV RNA在细胞中的定位;当蛋白酶K浓度为1：1 000、甲醛固定时间为30min时为最优反应条件;灵敏度试验表明该技术对病毒的检测极限为10-8/200μL,比FAT高3.5个数量级;特异性试验结果显示该探针能与CSFV 1.1、2.1、2.2、2.3亚型结合,与BVDV、PPV、PCV-2、PRV无交叉反应。采用该技术对CSFV RNA感染后在靶细胞中的定位与分布研究结果显示：0.5hpi在胞核和胞浆均能检测到RNA,0.5—6hpi RNA主要分布于胞核内并在核内富集;10hpi胞浆内RNA逐渐增多,胞核内RNA逐渐减少,24hpi RNA主要集中在胞浆内细胞核周围;36hpi核外RNA大量聚集增多,72hpi达到峰值;96hpi RNA总量有17
DOI:10.3864/j.issn.0578-1752.2016.12.015URL [本文引用: 1]
【目的】为研究CSFV RNA在体外感染细胞中的分布及定位,建立了一种准确、敏感的RNA可视化原位杂交技术。【方法】本研究通过比对Gen Bank中公布的CSFV、BVDV和BDV全序列,避开BVDV和BDV的同源区,设计了CSFV RNA及内参基因β-actin的特异探针。以CSFV中等致病力毒株（He BHH1/95）为参考毒株,在PK15细胞中培养病毒,加入RNA可视化原位杂交的特异探针和相应试剂,采用荧光共聚焦显微镜进行成像观察。通过综合分析观测结果、荧光强度、重复性等因素,采用正交试验优化了对原位杂交过程中具有重要影响的蛋白酶K浓度和甲醛固定时间,建立了CSFV RNA可视化原位杂交技术,并与FAT方法比较该技术灵敏度;用我国目前流行的CSFV 1.1、2.1、2.2、2.3基因亚型及BVDV、PPV、PRV和PCV-2病毒进行特异性试验。最终,以CSFV强致病力毒株（SM）接种PK15细胞,病毒感染后0.5、1、3、6、8、10、14、18、24、36、48、72、96h（hours post inoculation,hpi）取样,每个时间点2个重复,采用CSFV RNA可视化原位杂交技术进行检测。为佐证病毒蛋白在细胞中的定位及分布,同时采用FAT方法对SM株E2蛋白在PK15细胞中的表达情况进行动态研究。【结果】采用该技术在荧光共聚焦显微镜下可观察到CSFV RNA在细胞中的定位;当蛋白酶K浓度为1：1 000、甲醛固定时间为30min时为最优反应条件;灵敏度试验表明该技术对病毒的检测极限为10-8/200μL,比FAT高3.5个数量级;特异性试验结果显示该探针能与CSFV 1.1、2.1、2.2、2.3亚型结合,与BVDV、PPV、PCV-2、PRV无交叉反应。采用该技术对CSFV RNA感染后在靶细胞中的定位与分布研究结果显示：0.5hpi在胞核和胞浆均能检测到RNA,0.5—6hpi RNA主要分布于胞核内并在核内富集;10hpi胞浆内RNA逐渐增多,胞核内RNA逐渐减少,24hpi RNA主要集中在胞浆内细胞核周围;36hpi核外RNA大量聚集增多,72hpi达到峰值;96hpi RNA总量有17
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DOI:10.1016/j.virusres.2012.10.020URLPMID:23142553 [本文引用: 1]
African swine fever virus (ASFV) is a large icosahedral DNA virus which replicates predominantly in the cytoplasm of infected cells. The ASFV double-stranded DNA genome varies in length from about 170 to 193kbp depending on the isolate and contains between 150 and 167 open reading frames. These are closely spaced and read from both DNA strands. The virus genome termini are covalently closed by imperfectly base-paired hairpin loops that are present in two forms that are complimentary and inverted with respect to each other. Adjacent to the termini are inverted arrays of different tandem repeats. Head to head concatemeric genome replication intermediates have been described. A similar mechanism of replication to Poxviruses has been proposed for ASFV. Virus genome transcription occurs independently of the host RNA polymerase II and virus particles contain all of the enzymes and factors required for early gene transcription. DNA replication begins in perinuclear factory areas about 6h post-infection although an earlier stage of nuclear DNA synthesis has been reported. The virus genome encodes enzymes required for transcription and replication of the virus genome and virion structural proteins. Enzymes that are involved in a base excision repair pathway may be an adaptation to enable virus replication in the oxidative environment of the macrophage cytoplasm. Other ASFV genes encode factors involved in evading host defence systems and modulating host cell function. Variation between the genomes of different ASFV isolates is most commonly due to gain or loss of members of multigene families, MGFs 100, 110, 300, 360, 505/530 and family p22. These are located within the left terminal 40kbp and right terminal 20kbp. ASFV is the only member of the Asfarviridae, which is one of the families within the nucleocytoplasmic large DNA virus superfamily.
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DOI:10.1186/s40813-015-0013-yURLPMID:5382474 [本文引用: 1]
African Swine Fever (ASF) is an important contagious haemorrhagic viral disease affecting swine whose notification is mandatory due to its high mortality rates and the great sanitary and socioeconomic impact it has on international trade in animal and swine products. This disease only affects porcine species, both wild and domestic, and produces a variety of clinical signs such as fever and functional disorders of the digestive and respiratory systems. Lesions are mainly characterized by congestive-haemorrhagic alterations. ASF epidemiology varies significantly between countries, regions and continents, since it depends on the characteristics of the virus in circulation, the presence of wild hosts and reservoirs, environmental conditions and human social behaviour. Furthermore, a specific host will not necessarily always play the same active role in the spread and maintenance of ASF in a particular area. Currently, ASF is endemic in most sub-Saharan African countries where wild hosts and tick vectors (Ornithodoros) play an important role acting as biological reservoirs for the virus. In Europe, the disease has been endemic since 1978 on the island of Sardinia (Italy) and since 2007, when it was first reported in Georgia, in a number of Eastern European countries. It is also endemic in certain regions of the Russia Federation, where domestic pig and wild boar populations are widely affected. By contrast, in the affected eastern European Union (EU) countries where ASF is currently as epidemic, the on-going spread of the disease affects mainly wild boar populations located in restricted areas and, to a much less extent, domestic pigs. Unlike most livestock diseases, no vaccine or specific treatment is currently available for ASF. Therefore, disease control is mainly based on early detection and the application of strict sanitary and biosecurity measures. Epidemiology of ASF is very complex by the existence of different virus circulating, reservoirs and a number of scenarios, and the on-going spread of the disease through Africa and Europe. Survivor pigs can remain persistently infected for months which may contribute to virus transmission and thus the spread and maintenance of the disease, thereby complicating attempts to control it.
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DOI:10.1016/j.jcpa.2014.09.003URLPMID:25443146 [本文引用: 1]
African swine fever (ASF) is one of the most important infectious diseases of swine and has major negative consequences for affected countries. ASF is present in many sub-Saharan countries, Sardinia and several countries of eastern and central Europe, where its continuous spread has the swine industry on heightened alert. ASF is a complex disease for which no vaccine or treatment is available, so its control is based on early detection and rapid control of spread. For a robust and reliable early detection programme it is essential to be able to recognize the clinical signs and pathological changes of ASF, keeping in mind that in most cases the first introductions don't show high mortality nor characteristic clinical signs or lesions, but fever and some hemorrhagic lymph nodes. Knowledge of the main characteristics of this infection, including its current distribution and routes of transmission, is also essential for preventing and controlling ASF. This review addresses each of these topics and aims to update knowledge of the disease in order to improve early detection of ASF in the field and allow implementation of public health programmes.
[本文引用: 1]
[本文引用: 1]
