Temporal and spatial patterns of African swine fever in Sardinia

Temporal patterns and spatial distribution of African swine fever (ASF) were studied through the analysis of routinely collected data in the ASF-endemic area of the Province of Nuoro, Sardinia. During 1993–1996, ASF outbreaks were reported from 45 out of the 82 municipalities of the study area. Overall farm-level incidence rate (IR) was 1.3 outbreaks per 100 farms-year. ASF peaked in 1995 (IR=1.8) and declined in 1996 (IR=0.82). Significant (P<0.05) spring peaks of ASF outbreaks and affected municipalities were detected using statistical methods for circular distributions. Spatial clustering of ASF-affected municipalities, as evaluated by join-count statistics, was significant in 1993 (Z_jc=−3.0, P<0.01) and 1994 (Z_jc=−3.2, P<0.01) but not in 1995 (Z_jc=−0.6, P=0.55) and 1996 (Z_jc=−1.2, P=0.23). Extensive pig farming and ASF were spatially co-distributed (κ=0.51, 95% CI=0.33–0.70).     

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.     

Contribution of market value chain to the control of African swine fever in Zambia

Tropical Animal Health and Production - African swine fever (ASF) is a worldwide disease of pigs endemic in most sub-Saharan African countries. Zambia has been experiencing outbreaks of ASF for...     

African swine fever among slaughter pigs in Mubende district, Uganda

Owing to frequent reports of suspected outbreaks and the presence of reservoir hosts and vectors (warthogs, bushpigs and O. moubata ticks), African swine fever (ASF) is believed to be an endemic disease in Uganda. There have, however, been very few studies carried out to confirm its existence in Uganda. This study was carried out to describe the prevalence of ASF based on pathologic lesions and analysis of serum samples from slaughtered pigs during a suspected outbreak in the Mubende district of Uganda. The study was based on visits to 22 slaughterhouses where individual pigs were randomly selected for a detailed ante-mortem and post-mortem inspections. Sera were also collected for laboratory analysis. A total of 997 pigs (53.7% male and 46.3% female) were examined for lesions suggestive of ASF and sero-positivity of sera for ASF antibodies. The sera were tested using enzyme-linked immunosorbent assay (ELISA) and positive samples were further confirmed with an immunoblot assay. The results showed that 3.8% (38/997) of the pigs examined had clinical signs and post-mortem lesions suggestive of ASF. Two of 997 (0.2%) sera analysed were positive for ASF antibodies. Of the sub-counties investigated, Bagezza (12%) and Kiyuni (11%) had the highest prevalence of lesions suggestive of ASF based on ante- and post-mortem examination results, while Mubende town council (1.7%) had the lowest. This study found a low number of pigs (3.8%) with lesions suggestive of ASF at slaughter and an even lower number of pigs (0.2%) that were seropositive at slaughter, however a significantly higher number of pigs were slaughtered during the outbreak as a strategy for farmers to avoid losses associated with mortality.     

Pathological lesions and presence of viral antigens in four surviving pigs in African swine fever outbreak farms in Vietnam

Investigation of the role of animals that have recovered and survived from African swine fever (ASF) in carrying the ASF virus is currently intense and ongoing. However, no clear definition of the carrier stage has been established. The aim of the present study was to establish criteria to elucidate a clear status of survival in naturally ASF-infected domestic pigs in Vietnam. Seroconversion from previous infection was confirmed by serological assay, and the absence of the viral genome in various organs was also assured by molecular analysis of a partial p72 gene. We recognized that histopathological evidence could benefit from further insights into the status and role of the surviving animals; therefore, we performed a histopathological study on four pigs from farms with a history of ASF outbreak. We found fibrotic changes in the reparative process as the main finding in all four pigs. Immunohistochemical detection of viral protein revealed an interesting result. Despite the negative result from viral genome detection, the p30 protein gave a positive signal in the tonsils, lung, and stomach. This raises the possibility of stress-induced viral reactivation in long-term survivors and the risk of further outbreaks from human handling of contaminated carcasses.     

Comparison of a commercial H1N1 enzyme-linked immunosorbent assay and hemagglutination inhibition test in detecting serum antibody against swine influenza viruses.

Recently a commercial enzyme-linked immunosorbent assay (ELISA) kit for detecting antibody against H1N1 swine influenza virus (SIV) has been made available to diagnosticians and veterinary practitioners. Because the hemagglutination inhibition (HI) test has been considered the standard test for SIV serology, diagnostic performance of the new ELISA was evaluated using positive (n = 60) and negative (n = 188) serum samples from young pigs with known status of SIV infection and compared with that of the HI test. Both ELISA and HI test identified all negative animals correctly. None of the serum samples (n = 64) from pigs inoculated with H3N2 SIV was positive by ELISA for SIV antibody. The H1N1 SIV antibody detectable by ELISA appears to develop more slowly in comparison with antibody detectable by HI test. Although antibody was detected by HI test in all inoculated animals (n = 20) by day 7 postinoculation (PI), antibody was detected by ELISA in 0%, 75%, and 100% of the inoculated animals on days 7, 14, and 28 PI, respectively. Discrepancy in test results between the 2 serologic tests appeared to be because of differences in antibody isotypes detected by each test. Enzyme-linked immunosorbent assay mainly detected IgG antibody, whereas the HI test detects IgM antibody very efficiently as well as IgG antibody. Collectively, the commercial ELISA is highly specific for antibody to H1N1 SIV but may not identify positive animals at the early stage of infection as effectively as the HI test, particularly when SIV is introduced to a na?ve swine population.     

Review of African swine fever outbreaks history in South Africa: From 1926 to 2018

The article reviews the outbreaks and distribution of African swine fever (ASF) in South Africa since the first probable outbreak that occurred in the Koedoesrand Ward in 1926. Retrospective data on the ASF outbreaks in South Africa were obtained from the World Organisation for Animal Health (OIE) disease database and the South African veterinary services annual reports in addition to published articles and online sources. South Africa has experienced many outbreaks that can be divided into 2 time periods: the period before the development of the OIE diseases database (1993) and the period after. More than 141 outbreaks of ASF were reported during the first period. Since the development of OIE disease database, 72 outbreaks directly involving 2968 cases, 2187 dead and 2358 killed pigs mainly in smallholder pig farms were reported. The median number of cases for a given ASF outbreak is 17, but in 50% of outbreaks no pigs were killed for prevention. The most important ASF outbreak was reported in April 2014 in the Greater Zeerust district (North West province) involving 326 cases and 1462 killed pigs. However, the outbreak with highest mortality involving 250 pigs was reported in 2016 (Free State province). According to phylogenetic analysis, nine p72 genotypes (I, III, IV, VII, VIII, XIX, XX, XXI and XXII) have been identified in South Africa. Season-wise, more outbreaks were recorded during summer. It was also observed that the OIE disease database could contain errors that would have been introduced through compiled forms at country level. Spatiotemporal studies on ASF outbreaks in South Africa are therefore required in order to assess statistically and quantitatively the clustering of outbreaks over space and time.     

Establishment of an efficient enzyme-linked immunosorbent assay for the detection of Eperythrozoon sius antibody in swine

The Eperythrozoon suis (E. suis) antigen was purified using a Sephadex G-200 chromatograph, and thereby, a high-affinity, specific E. suis antigen was collected and confirmed with Western blotting. Using this antigen, an enzyme-linked immunosorbent assay (ELISA) system to detect the antibody against E. suis in swine was established. There was no cross-reaction with swine sera, which were affected with Mycoplasmal pneumonia, swine fever, swine colibacillosis, or toxoplasmosis. A comparison of this ELISA system with an indirect hemagglutination (IHA) test using 78 swine samples revealed that the ELISA system significantly improved the sensitivity, specificity, and stability for the serodiagnosis of swine E. suis.     

Benefit-cost analysis of the current African swine fever eradication program in Spain and of an accelerated program

We predicted the rate of spread and economic losses due to African swine fever (ASF) infections in Spanish swine for a 20 year period using current epidemiological data. Two possibilities for eradication were examined: continuation of the eradication program presently in use, assuming continued funding at present levels, and an accelerated eradication program using more frequent testing to identify seropositive carrier animals. Both programs theoretically would result in the eradication of African swine fever. The calculated benefit-cost ratios were 1.23 and 1.47 respectively. A reduction in current yearly funding would result in a benefit-cost ratio of 0.97, making the program unprofitable according to our model.

The projected year for the eradication of outbreaks is 1996 using provinces as units in the model. Similarly, the projected year for the eradication of seropositive animals is 2001.     

非洲猪瘟对我国养猪业的影响与防控建议

非洲猪瘟是由非洲猪瘟病毒引起的一种高度接触传染性、广泛出血性猪病毒病,死亡率可高达100%,无商品化疫苗可用于其预防。自8月初我国爆发首例非洲猪瘟疫情以来,截至11月5日,短短三个月时间里,已有14个省累计爆发58起疫情,扑杀生猪近50万头,直接经济损失数十亿元,给我国养猪业造成了巨大冲击。面对前所未有的非洲猪瘟疫情,我国政府的应对是果断有力的,但也面临种种挑战。本文对我国非洲猪瘟疫情形势及其对相关行业的影响进行了分析,总结了非洲猪瘟防控的重点和难点,并针对目前的形势,提出了遏制非洲猪瘟疫情进一步蔓延的建议和对策。     

Control of post-vaccinal immunity against rinderpest after the 1989 and 1990 vaccination campaigns in Ivory Coast]

The Central Laboratory of Animal Pathology of Bingerville (C?te-d'Ivoire) made an evaluation of post vaccinal immunity against rinderpest in 1990 and 1991 after the 1989 and 1990 national vaccination campaigns. The random sampling method was chosen to collect 6,020 sera in 255 places in 1990 and 3,301 sera in 158 places in 1991. ELISA analysis gave a standard positive rate of 82.39% +/- 0.08 in 1990 and 88.26 +/- 0.06% in 1991.     

非洲猪瘟病毒的分子病原学及致病机理研究进展

非洲猪瘟（African swine fever,ASF）是由非洲猪瘟病毒（African swine fever virus,ASFV）引起的一种烈性传染病,具有急性、高热、高病死率等特征,主要暴发于非洲、东欧国家、俄罗斯及高加索地区。目前,该病缺乏有效的疫苗和治疗方法,给病情爆发地区的养猪业造成严重的影响。ASFV的主要靶细胞是网状内皮细胞和单核-巨噬细胞,造成细胞凋亡,影响宿主的免疫系统,进而表现出相应的疫病特征。ASFV具有基因组较大、基因型较多且易变异等特征。文章主要从分子病原学和致病机理方面对ASFV的研究情况进行综述,为ASF的防控提供理论依据。     

Research Progress on Molecular Etiology and Pathogenesis for African Swine Fever Virus

African swine fever (ASF) is a devastating disease caused by African swine fever virus (ASFV), characterized by acute feature, high fever, high mortality and other characteristics. ASF mainly outbreaks in African, Eastern Europe countries, Russia and Caucasus region. At present, there are no vaccine available and effective control strategies against ASFV spread, therefore, ASF has a serious impact on the pig industry in the affected countries. The major target cells of the virus are swine reticuloendothelial cells and monocyte-macrophage cells. ASFV can result in apoptosis and affect the host's immune system, and then show the characteristics of the corresponding disease. ASFV has the characteristics of large genome, more genotype and more variability. In this paper, the molecular etiology and pathogenesis of ASFV are reviewed, so as to provide theoretical basis for prevention and control of ASF.     

African swine fever outbreak on a medium-sized farm in Uganda: biosecurity breaches and within-farm virus contamination

In Uganda, a low-income country in east Africa, African swine fever (ASF) is endemic with yearly outbreaks. In the prevailing smallholder subsistence farming systems, farm biosecurity is largely non-existent. Outbreaks of ASF, particularly in smallholder farms, often go unreported, creating significant epidemiological knowledge gaps. The continuous circulation of ASF in smallholder settings also creates biosecurity challenges for larger farms. In this study, an on-going outbreak of ASF in an endemic area was investigated on farm level, including analyses of on-farm environmental virus contamination. The study was carried out on a medium-sized pig farm with 35 adult pigs and 103 piglets or growers at the onset of the outbreak. Within 3 months, all pigs had died or were slaughtered. The study included interviews with farm representatives as well as biological and environmental sampling. ASF was confirmed by the presence of ASF virus (ASFV) genomic material in biological (blood, serum) and environmental (soil, water, feed, manure) samples by real-time PCR. The ASFV-positive biological samples confirmed the clinical assessment and were consistent with known virus characteristics. Most environmental samples were found to be positive. Assessment of farm biosecurity, interviews, and the results from the biological and environmental samples revealed that breaches and non-compliance with biosecurity protocols most likely led to the introduction and within-farm spread of the virus. The information derived from this study provides valuable insight regarding the implementation of biosecurity measures, particularly in endemic areas.     

云南省不同宿主H9N2感染调查和NA分子进化分析

2006年6月-2007年10月,从云南省16个地州随机采集的各种家禽、家猪的喉气管和口腔棉拭子样品7 901份,应用RT-PCR结合血凝、血凝抑制试验和抗原捕捉ELISA,检测H9N2亚型禽流感感染情况,选择具有代表性的阳性样品,克隆NA全基因并进行基因序列分和推导氨基酸的糖基化位点分析。结果表明,云南省2006-2007年H9N2亚型禽流感病毒在全省14个地州均有流行,感染宿主包括鸡、鸭、鹅和猪;分离的19个毒株中的13个鸡源和1个鹅源毒珠的NA全基因长1 407bp,编码469个氨基酸,同源性为96.1%～99.6%,是目前云南省流行的优势亚群;3个鸭源毒株、1个鹅源和1个猪源毒株NA基因长1 398bp,编码466个氨基酸,5个非鸡源毒株的NA基因的同源性为99%,构成云南目前流行的另一分支,推导氨基酸第61～63位3个氨基酸的缺失是与优势分支的特征性区别;2个分支间NA基因同源性为90%,与国内外其他毒株比较神经氨酸酶基因序列具有多态性。     

Co-circulation of two genetically distinct viruses in an outbreak of African swine fever in Mozambique: no evidence for individual co-infection

In 1998, domestic pigs originating from villages within a 40 km radius of Ulongwe in the northern Tete Province of Mozambique were held in a quarantine facility for a 3-month period prior to their importation into South Africa. Eight of a total of 25 pigs died within the first 3 weeks of quarantine of what appeared clinically and on post mortem examination to be African swine fever (ASF). Organs were collected and preserved in formol-glycerosaline and the presence of ASF virus in these specimens was confirmed by three independent polymerase chain reaction (PCR) tests. Two gene regions were characterised, namely the C-terminus end of the major immunodominant protein VP72 and the central variable region (CVR) of the 9RL open reading frame (ORF). Results confirmed the presence of two genetically distinct viruses circulating simultaneously within a single outbreak focus. However, despite the pigs being housed within the same facility, no evidence of co-infection was observed within individual animals. Comparison of the two 1998 virus variants with viruses causing historical outbreaks of the disease in Mozambique revealed that these viruses belong to two distinct genotypes which are unrelated to viruses causing outbreaks between 1960 and 1994. In addition, the CVR and p72 gene regions of one of the 1998 Mozambique virus variants (variant-40) was shown to be identical to the virus recovered from an ASF outbreak in Madagascar in the same year, whilst the other (variant-92) was identical to a 1988 pig isolate from Zambia.     

Evaluation of ELISA for the detection of Toxoplasma antibodies in swine sera

Enzyme-linked immunosorbent assay (ELISA) is compared with the indirect fluorescent antibody test (IFAT), the indirect haemagglutination test (IHAT) and the latex agglutination (LA) test for the detection of toxoplasma antibodies in swine sera. The 100 swine sera examined represent ELISA values from greater than 0 to 154 EIU. The agreement was highest (0.67) between ELISA and IFAT with an ELISA cut-off value of 30 EIU, and between ELISA and the LA test with an ELISA cut-off value of 50 EIU (0.74). All sera giving less than 10 EIU were negative in the other tests, and all those with greater than 70 EIU were positive in 1, 2 or all of the reference tests. In order to avoid false positive results with ELISA, all sera giving 10-70 EIU should be confirmed with a test which has a good specificity, e.g. IFAT. ELISA is a sensitive test and is highly suitable for the screening of large amounts of samples, but it may be too complicated for screening toxoplasma antibodies in the laboratories of abattoirs.     

猪乙型脑炎病毒囊膜E蛋白间接ELISA诊断方法的建立与初步应用

本研究利用纯化的原核表达乙型脑炎囊膜E蛋白作为包被抗原,建立了乙型脑炎间接ELISA诊断方法。对检测的各种条件进行了优化,优化反应条件后确定的抗原最适包被浓度为2μg/mL,抗原最佳包被条件为37℃包被2 h,血清的最适稀释度为1∶160,酶标抗体最适稀释度为1∶5000,最佳封闭条件为1%BSA,阴阳性临界值判定标准为D492 nm=0.254。该方法不与猪瘟、猪繁殖与呼吸综合征、猪圆环病毒2型、猪伪狂犬病毒阳性血清反应,其D492 nm0.254,说明该方法具有良好的特异性。采用该方法对150份疑似乙型脑炎血清样品进行检测,结果显示,与某猪乙型脑炎试剂盒相比符合率为90.77%,表明建立的间接ELISA方法具有较高的敏感性和特异性,因此,本研究成功建立了能特异性检测抗乙型脑炎血清抗体的ELISA检测方法。     

非洲猪瘟诊断方法研究进展

非洲猪瘟(African swine fever,ASF)是由非洲猪瘟病毒(African swine fever virus,ASFV)引起的猪的急性、热性、高度接触性传染病,急性病例的发病率和死亡率几乎达100%。世界动物卫生组织将ASF列为法定报告动物疫病,中国将其列为一类动物疫病。当前中国ASF疫情防控形势十分严峻,且尚无有效的疫苗及其他防治制剂,因此执行严格的卫生措施及监测病原和抗体,进而排查和清除带毒动物是ASF防控的重要措施。笔者就ASFV诊断标识出发,总结了近年来ASF病原检测与血清学监测技术的最新研究进展。抗原检测方面,在PCR基础上发展的多种等温扩增技术如重组酶聚合酶扩增技术、交叉引物扩增技术、环介导等温扩增技术及微滴数字PCR从不同方面克服了常规PCR的缺陷,实时荧光定量PCR检测方法具有速度快、准确、结果客观和可定量基因组含量等优点,新型CRISPR/Cas12a技术无需昂贵的仪器,可为ASFV的检测提供一种更加便携、简单、灵敏和特异性强的新型替代方法;抗体检测方面,基于酶联免疫吸附试验(ELISA)、免疫层析技术和间接免疫荧光试验等的检测方法灵敏度和特异性也有不同程度的提高。     

非洲猪瘟间接ELISA诊断试剂盒的研究

用带有编码非洲猪瘟病毒衣壳蛋白Ｐ７２ 基因的重组杆状病毒（Ｂａｃｐ７２）作载体,在ｓｆ９细胞中表达并得到重组Ｐ７２蛋白,ＳＤＳ－ＰＡＧＥ可得到分子量在 ７２ｋＤａ左右的电泳带。用标准阳性非洲猪瘟血清对 Ｐ７２蛋白进行 ＥＬＩＳＡ检测,证明该蛋白具有生物学活性。用Ｐ７２作为间接法的包被抗原,对ＥＬＩＳＡ反应条件进行了优化。确定最佳包被液为ＰＢＳ（ｐＨ７．２）、最佳封闭液为１％ＰＣＴ、最佳血清稀释液为４％ＰＥＧ６０００／ＰＢＳ、最佳冲洗液为０．５Ｍ ＮａＣｌ／０．５％Ｔｗｅｅｎ－２０／ＰＢＳ（ｐＨ７．２）。本实验反应体系采用５０μｌ的微量法,可节约试剂及抗原。反应在２小时内即可完成,达到了快速诊断的目的。包被了抗原并用封闭液封闭后的酶标板密封后保存于－２０℃的冰箱中,至少可以保存５个月。阻断试验和交叉试验表明ＥＬＩＳＡ法有良好的特异性。间接ＥＬＩＳＡ比Ｄｏｔ－ＥＬＩＳＡ法具有更高的灵敏性。血清学调查没有得到阳性结果,与我国实际情况相符。用Ｂａｃｐ７２表达的非洲猪瘟病毒Ｐ７２蛋白抗原作为间接ＥＬＩＳＡ的检测抗原来检测非洲猪瘟血清具有快速、简单、无感染的特点。本实验为非洲猪瘟ＥＬＩＳＡ检测试剂盒的最终组装提供了实验依据。