Mechanism of thrombocytopenia in African swine fever

Pigs were inoculated with an African swine fever (ASF) isolate of moderate virulence, and the changes in the number of circulating blood platelets during infection were correlated with the appearance of antiviral antibody and fluctuations in total plasma hemolytic complement concentrations. Thrombocytopenia was detected by postinoculation days (PID) 7 and 8, and antiviral antibody was detected by PID 7, using an indirect immunofluorescence technique. The total hemolytic complement concentration was moderately and transiently decreased from PID 5 to 9, but was consistently low from PID 18 to 26. Pigs inoculated with an ASF virus isolate of greater virulence had a decrease in platelet counts on PID 6 and 7, and the total plasma hemolytic complement levels decreased in all pigs by PID 6 to 7. Antibody to ASF virus was not detected in pigs inoculated with the more virulent isolate. Pigs sensitized to ASF viral antigen with an inactivated-virus vaccine or by previous infection with ASF were challenge exposed. Sensitized pigs became clinically ill and thrombocytopenic by 24 to 72 hours earlier than did inoculated, nonsensitized pigs. Vaccinated pigs inoculated with homologous virus had lower blood virus concentrations than did nonvaccinated pigs. African swine fever virus-sensitized pigs inoculated with heterologous virus had a higher fatality rate than did nonsensitized pigs, and the pigs died peracutely, with only a few gross lesions in evidence. In vitro experiments demonstrated that ASF virus antigen induced platelet aggregation in platelet-rich plasma from recovered, nonviremic pigs. Viral antigen, antibody, or complement was not demonstrable on the surface of platelets from pigs inoculated with ASF virus isolate, by direct immunofluorescence testing.     

非洲猪瘟背景下的三元商品母猪养殖策略

非洲猪瘟病毒(African swine fever virus,ASFV)是一种病毒分子巨大且复杂的DNA病毒,在无有效疫苗的情况下,会引起高度传染性且通常致命的猪瘟。目前非洲猪瘟(African swine fever,ASF)导致我国母猪存栏大幅度下降,猪场引种面临了巨大的生物安全风险,采用三元商品母猪留种成为了一种较好的选择。文章结合生产实践,旨在为三元商品母猪的饲养管理和营养调控提供指导意见。     

某养殖场的病死野猪ASFV实时荧光PCR检测

为了进一步做好非洲猪瘟疫情防控工作,研究用实时荧光PCR法对武威市某养殖场病死野猪的猪耳朵和猪鼻拭子样品进行了非洲猪瘟病毒检测。结果显示该养殖场病死猪耳样品和猪鼻拭子样品均无Ct值,线形为直线,表明所有被检测样品均为ASFV抗原阴性。研究为严防非洲猪瘟进入威武市奠定了坚实的技术储备,也为威武市非洲猪瘟疫情防控提供了一定参考。     

非洲猪瘟研究进展及防控措施

非洲猪瘟(African swine fever,ASF)是由非洲猪瘟病毒科、非洲猪瘟病毒属的唯一成员非洲猪瘟病毒(African swine fever virus,ASFV)引起的一种高致死率、高抵抗力、严格接触性的病毒性疾病,对所有年龄段的家猪和野猪均具有感染性。我国非洲猪瘟传入疫情的出现,对养猪生产构成严重威胁,必将冲击生猪产业的健康稳定发展。本文介绍ASF的病原、流行病学特点,并总结防控措施,以期为ASF的进一步防控提供参考。     

非洲猪瘟病原学及单克隆抗体制备技术研究进展

非洲猪瘟是由非洲猪瘟病毒引起猪的高度接触性、传染性、出血性以及高死亡率的传染病。20世纪中期以来,已在非洲、欧洲和美洲等数十个国家流行,并在近几年内蔓延至欧亚两洲接壤处的格鲁吉亚、亚美尼亚、阿塞拜疆以及俄罗斯境内,其一旦侵入我国,将会给我国养猪业带来极大的危害。非洲猪瘟病原学研究以及制备相应的单克隆抗体对非洲猪瘟病毒快速诊断技术研究和疫苗研制有着重大的现实意义。主要从病原学和单克隆抗体制备方面对非洲猪瘟的研究进展进行了综述。     

An immunohistochemical study of the tonsils in pigs with acute African swine fever virus infection

An immunohistochemical study of the tonsils was carried out to gain further insight in the pathogenesis of acute African swine fever (ASF). Twenty-one pigs were inoculated by intramuscular route with a highly virulent isolate of ASF virus and painlessly killed at 1-7dpi. Viral antigen was highly distributed in the tonsil from 3 to 4dpi and an increase in the number of monocyte-macrophages was very evident at the same days post inoculation. This phenomenon was observed together with an increase of the expression of proinflammatory cytokines (Tumour necrosis factor alpha and Interleukin-1 alpha) and the apoptosis of lymphocytes studied by the terminal deoxynucleotidyltransferase-mediated dUTP nick end labelling (TUNEL) technique and haemorrhages. With these results, we can conclude that the tonsil is suffering similar lesions than those observed in other lymphoid organs in acute African swine fever, even when the route of inoculation is the intramuscular and not oral-nasal.     

浅谈非洲猪瘟背景下的后备母猪初配标准和改善措施

非洲猪瘟进入我国已2年有余,目前全国生猪产能已经恢复到非瘟发生前的88％以上.但是,猪场在快速恢复产能过程中诸多问题会对猪场生产成绩造成持续影响,例如如何确立后备母猪初配标准,亟待建立和规范.文章从后备母猪初配标准的重要性、关键指标和改善措施3个方面进行深入剖析和探讨,旨在为猪场建立最佳的后备母猪初配条件提供参考.     

非洲猪瘟常态下猪的日粮营养与饲料安全

随着非洲猪瘟疫情的暴发,我国的生猪养殖业遭受了前所未有的毁灭性打击,使得中国猪肉的自给能力也面临着巨大的威胁与挑战。由于非洲猪瘟病毒结构的复杂性,安全有效的疫苗开发道路也是曲折且漫长。对此,文章主要围绕母仔猪的日粮营养和饲料安全两方面展开讨论,旨在浅析非洲猪瘟常态下,如何维持和提高猪场的稳定与发展,促进我国养猪生产水平的迅速恢复与猪肉供给能力,稳定终端消费市场。     

非洲猪瘟病毒与宿主细胞相互作用研究进展

非洲猪瘟(ASF)自2018年8月在我国暴发,对养猪业构成巨大威胁.该病是由非洲猪瘟病毒(ASFV)引起的家养猪高热、急性和高死亡率的出血症和淋巴组织坏死症;认识和了解病原ASFV与宿主的相互作用是理解致病机制的基础和疫病防控的前提.本文对ASFV与宿主细胞受体和内体系统的互作、基因转录和蛋白合成系统的互作、细胞凋亡和内质网应激系统的互作、天然免疫干扰素应答系统的互作、抗原递呈细胞的互作五个方面现有研究进展进行综述;并试图在此基础上理解ASFV的免疫保护和免疫逃逸作用,思考ASFV与宿主互作中需要回答的问题,为研制保护性疫苗和深入认识ASF致病机制提供线索.     

Approaches to the identification of non-essential genes of African swine fever virus.

It is poorly understood why vaccines could not be developed for the control and prevention of African swine fever (ASF) virus infection. The aim of our study was to identify genes non-essential for ASF virus replication because there were indications that certain viral gene products, which apparently are non-essential for viral replication, conferred protection from death due to ASF. A cosmid library representing the genome of ASF virus strain France 64 was established and characterized. Then, in order to inactivate viral genes by insertion, the beta-galactosidase (beta-gal) gene was introduced either randomly or at specific locations of selected cloned DNA fragments. These constructions were transfected into cells which had been previously infected with a cell-culture-adapted viral strain in order to allow the generation of recombinant progeny virus. Viable recombinant progeny was identified by at least one of the following means: (1) expression of beta-gal; (2) detection of beta-gal specific DNA by plaque hybridization, and (3) absence of a functional product of the inactivated gene. Presently, we are characterizing a recombinant virus with an insertionally inactivated thymidine kinase gene.     

A highly sensitive and specific gel-based multiplex RT-PCR assay for the simultaneous and differential diagnosis of African swine fever and Classical swine fever in clinical samples

The development and standardisation of a novel, highly sensitive and specific one-step hot start multiplex RT-PCR assay is presented for the simultaneous and differential diagnosis of African swine fever (ASF) and Classical swine fever (CSF). The method uses two primer sets, each one specific for the corresponding virus, amplifying DNA fragments different in length, allowing a gel-based differential detection of the PCR products. Universal detection of ASF and CSF virus strains was achieved through selection of primers in conserved viral genome regions. The detection range was confirmed by analysis of a large collection of isolates of the two viruses. The high specificity of the assay was proven by testing related viruses, uninfected cell line cultures and healthy pig tissues. Additional confirmatory tests of the ASF and CSF virus amplicon specificity, based on restriction endonuclease analysis with BsmA I or Ban II, respectively, are also described. The analysis of whole blood and serum samples from experimentally infected animals proved the usefulness of the method for an early diagnosis of both diseases, even before the appearance of the first clinical signs. A study of 150 positive field samples from several ASF and CSF outbreaks showed the suitability of this method for a rapid (less than five hours), sensitive and specific differential diagnosis in clinical samples. In addition, a highly sensitive and specific uniplex RT-PCR for CSFV was also developed and standardised as a powerful tool for fast and early diagnosis of the disease.     

Detection of African swine fever viral antigens in paraffin-embedded tissues by use of immunohistologic methods and polyclonal antibodies.

Tissues obtained from pigs inoculated with African swine fever virus (ASFV), fixed by vascular perfusion using glutaraldehyde, and embedded in paraffin or araldite were used for an immunohistologic electron microscopic study. To detect ASFV antigens, 4 methods were used on paraffin sections with or without pretreatment of the tissues. Use of biotinylated anti-ASFV antiserum combined with avidin-biotin complex and peroxidase proved to be the most suitable method, and antigen was detected in tissues infected with 2 ASF viruses of different virulence. Use of the glutaraldehyde fixation method should ensure optimal morphologic (structural and ultrastructural) data while allowing an immunohistologic study, and add to knowledge of the pathogenesis of ASF.     

Extraction of viral DNA from erythrocytes of swine with acute African swine fever

The preparation of wild-type African swine fever (ASF) virus DNA from small amounts of viremic blood from acutely febrile pigs is outlined. The extracted DNA is viral and not host-cell DNA, because of specific homology with cell culture grown and purified ASF virus and because no DNA bands are obtained with an equal amount of nonviremic pig blood. Thus, in the absence of suitable serologic methods for strain identification, it is now possible to catalogue wild-type isolates by characteristic DNA restriction patterns. The wild-type virus genome contains terminal single-stranded DNA cross-links and has the largest genome size (180 kilobase pairs) reported for the ASF virus. Experimental passage of the virus in contact-infected pigs and buffy coat cultures appears to confirm the stable nature of the ASF genome in the field.     

Age-related viral load and severity of systemic pathological lesions in acute naturally occurring African swine fever virus genotype II infections

African swine fever (ASF) causes a contagious hemorrhagic disease in all ages of pigs without sex predilections. The objective of this study was to determine the age-related viral loads and severity of systemic pathological lesions among three different swine group ages (weaned pigs, fattening pigs, and sows) during a recent outbreak of acute ASF in Vietnam. Age-related viral loads were determined in 5 major organs (lung, liver, spleen, kidney, and lymph node) by immunohistochemistry as well as in the blood by real-time polymerase chain reaction (PCR). Age-related systemic pathological lesions were analyzed in the listed organs among three age groups.Weaned pigs had significantly (p < 0.05) higher levels of viral loads in their lung, liver, lymph nodes and blood than in those of fattening pigs and sows. Fattening pigs had significantly (p < 0.05) higher scores of macroscopic lung and lymphoid lesions, and microscopic liver lesions compared with those of weaned pigs and sows. The results of this study demonstrated that viral loads were age-related in acute naturally occurring ASF but the severity of pathological lesions was not correlated with the level of viral loads in the five major organs.     

猪场生物安全系统要点

建立生物安全体系是猪场防控疾病的前提,也是最经济有效的疫病防治措施。2018年我国发生非洲猪瘟以来,疫情对我国生猪养殖业带来深刻影响,在未来很长一段时间内该病都将是我国猪场的常态化疾病。非洲猪瘟疫情发生后,我国养猪业创新性地提出了系列生物安全防控技术,建立了能够有效应对疫情常态威胁的生物安全技术体系,两年多来的防控实践证明了这些生物安全防控手段的有效性。文章总结了猪场生物安全体系建设和管理中的关键环节及注意事项,希望可以为猪场非洲猪瘟防控提供参考。     

The lesional changes and pathogenesis in the kidney in african swine fever

African swine fever is a viral haemorrhagic disease of pigs which has been used as a model for the study of viral haemorrhagic diseases in man. The acute course of the disease is characterized by acute proliferative glomerulonephritis, with viral replication in mesangial cells and occasional focal necrosis of the renal tubular system; hyperplasia of the collecting ducts is associated with evident virus replication. Haemorrhages have been attributed to endothelial dysfunction, aggravated by virus replication in endothelial cells in the final stages of the disease. The renal interstitium displays intense oedema and an infiltrate largely composed of macrophages. Virus replication has also been observed in fibroblasts and in the smooth0muscle cells of arterioles and venules.In subacute-chronic forms of the disease, various types of glomerulonephritis are observed, ranging from mesangial proliferative glomerulonephritis to focal and segmental hyalinosis associated with immune-mediated phenomena. No striking changes are reported in the renal tubular system. Interstitial haemorrhages are associated with diapedesis due to immunologically mediated events. The interstitium has also been found to contain a lymphohistiocytic infiltrate with abundant plasma cells. No evidence has been reported of viral replication in any cell population.Abbreviations ASF African swine fever - ASFV African swine fever virus - C complement component - DNA deoxyribonucleic acid - HAD₅₀ 50% haemoadsorbing doses - Ig immunoglobulin - MPS mononuclear phagocytic system - PAS periodic acid-Schiff reagent     

非洲猪瘟病毒入胞过程相关蛋白及分子机制的研究进展

非洲猪瘟病毒(ASFV)是非洲猪瘟(ASF)的病原,可引发家猪和野猪急性、传染性出血死亡,目前尚无有效的疫苗和抗病毒药物。ASFV是囊膜病毒,其囊膜蛋白的结构和功能可能是影响病毒入侵和细胞嗜性的重要因素。病毒入胞是病毒感染细胞的第一步,通常是通过细胞表面特定的分子与病毒蛋白相结合吸附于宿主细胞表面,对ASFV入胞过程的病毒蛋白或宿主因子为靶点或可有效抑制ASFV的复制。本文从ASFV入胞过程中起重要作用的囊膜蛋白出发,对ASFV的入胞分子机制进行综述,为ASFV入胞深入研究以及治疗性药物和疫苗的研发提供参考。     

非洲猪瘟病毒免疫逃逸分子机制研究进展

非洲猪瘟(African swine fever，ASF)是由非洲猪瘟病毒(African swine fever virus，ASFV)感染猪引起的一种急性、烈性、高度接触性传染病，至今没有研发出安全有效的疫苗，一旦暴发会造成重大经济损失。ASFV在和宿主长期作用过程中，通过抑制干扰素和炎症反应，调节凋亡、自噬及细胞免疫等多种途径逃逸机体免疫反应促进自身复制，但具体的机制仍不完全清楚。ASFV复杂的免疫逃逸机制可能是阻碍有效疫苗研发的关键因素之一。借助生物信息学技术对ASFV的基因组和蛋白质组深入分析，筛选病毒的免疫调控关键基因和保护性抗原表位，将在ASFV免疫逃逸分子机制的研究与疫苗研发中发挥重要作用。本文主要对ASFV感染引起的免疫应答反应及可能的免疫逃逸机制研究进行概述，以期为ASF疫苗研制及综合防控提供思路。     

Inhibitory effect of African swine fever virus on lectin-dependent swine lymphocyte proliferation

The incubation of swine peripheral blood mononuclear cells (PBMC) with African swine fever (ASF) virus preparations strongly inhibited the proliferative response of lymphocytes to PHA and other lectins. The inhibition, which persisted after inactivation of the virus by UV radiation, was dependent upon the dose and the time that virus preparations were present in cultures. When virus preparations were fractionated by ultracentrifugation, the inhibitory activity resulted to be soluble, whereas no activity was found in the sedimented viral fraction. However, the preincubation during 4 days of this sedimented fraction with swine PBMC, before the addition of the mitogen, restored the inhibitory activity. The results obtained suggest that the inhibition is mediated by one or more soluble factors released by swine PBMC after coincubation with ASF virus in a time dependent process. These factors show a molecular weight between 40 and 80 kDa by gel filtration chromatography. The inhibitory activity described in the present paper is an indication of inhibition of lymphocyte function produced by ASF virus which can help to understand how this virus escapes from the host immune system.     

非洲猪瘟的研究进展

非洲猪瘟是猪的一种急性、高度接触传染性疾病。其感染率高, 多种传播途径,死亡率可达100％,现还无有效的疫苗及治疗措施。非洲猪瘟在临床上与猪瘟相似，虽这2种疾病的病原体完全不同，但在诊断时极易误诊，为了进一步了解该病,作者综述了该病的病原、流行病学、临床症状、鉴别诊断及我国现状，从而为诊断该病提供了依据。