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1.
We report the development of a competitive enzyme-linked immunosorbent assay (c-ELISA) for the detection of antibodies to porcine circovirus type 2 (PCV2), the agent associated with the recently described postweaning multisystemic wasting syndrome in pigs. At present, no method has been published describing a c-ELISA for the detection of antibodies to PCV2, and currently employed tests are impractical for use in some laboratories. The assay described here uses a cell culture isolate of porcine circovirus type 2 as antigen and a PCV2-specific monoclonal antibody as the competing reagent. Evaluation of the ELISA was performed by comparison with results obtained using an indirect immunofluorescent test on 484 sera from pig herds in the United Kingdom, Canada, France, and the USA and serial bleeds from pigs experimentally infected with porcine circoviruses. The sensitivity and specificity of the ELISA were determined as 99.58% and 97.14%, respectively, at 2 standard deviations (SD) from the mean or 95.81% and 100% at 3 SD from the mean. Using this ELISA, a serologic survey of 461 sera collected from commercial pig herds in Northern Ireland between 1973 and 1999 was undertaken. Analysis of the results of this survey demonstrated that the number of ELISA-positive sera detected in an individual year during this period ranged from 55% to 100%. This c-ELISA has applications for large-scale rapid diagnosis of PCV2 infection in pig populations worldwide and for immunoscreening of sera from other species for antibodies to PCV2.  相似文献   

2.
The genome of porcine circovirus type 2 (PCV2) contains two major open reading frames, which have been shown to encode the virus capsid and replication-associated proteins. The capsid protein is a major structural protein of the virus; it can be a suitable target antigen for detecting PCV2-specific antibodies to monitor PCV2 infection. To produce the antigen, the capsid protein coding sequence was cloned into a baculovirus transfer vector, and a recombinant capsid (rC) protein of PCV2 was expressed as a combined fusion protein in frame with a C-terminal peptide of six histidines. The affinity-purified rC protein was used as coating antigen to develop an ELISA for detecting the virus-specific antibodies in swine sera. The rC protein-based ELISA (rcELISA) was evaluated by examining a panel of 49 PCV2-positive and 49 PCV2-negative swine sera. In comparative experiments of immunoperoxidase monolayer assay (IPMA) using 102 field sera, there was 89.2% coincidence between data obtained by the rcELISA and IPMA. The rcELISA achieved 88.5% specificity and 89.4% sensitivity for detection of PCV2 antibody in the field sera. The assay showed no cross-reactivity with antibodies to PCV type 1, porcine reproductive and respiratory syndrome virus and porcine parvovirus. The results suggest that the rcELISA is suitable for routine serodiagnosis and epidemiological surveys of PCV2-associated diseases.  相似文献   

3.
The aim of this study was to evaluate the vasculature in porcine circovirus type 2-infected (PCV2-infected) lungs and to identify the PCV2 subtypes involved in porcine pneumonia. Pulmonary samples from 140 pigs, 2 weeks to 7 months of age, from 36 Hungarian commercial herds with clinical signs of respiratory disease were examined for the presence of respiratory pathogens, with bacterial culture, pathologic evaluation, and immunohistochemistry for PCV2, porcine reproductive respiratory syndrome virus, and swine influenza virus. PCV2 was the most commonly identified pathogen (49 cases) among the 74 of 140 cases (53%) with respiratory pathogens. PCV2 was detected immunohistochemically in the wall of 13% to 100% of pulmonary vessels (mean, 89%) in 38 of 49 cases (78%). Detection of PCV2 antigen was positively correlated with the presence of vascular lesions (P < .001, odds ratio [OR]: 159.54). Other pathogens capable of vascular injury in swine were found in 29 of 49 of the PCV2-positive cases (59%). The probability of detecting vascular lesions in PCV2-infected lung was higher than in infection with porcine reproductive respiratory syndrome virus (P < .002, OR: 14.63), Pasteurella multocida infection (P < .001, OR: 5.75), or Streptococcus spp. infection (not significant, OR: 1.45). Sequence analysis of open reading frame 2 amplicons was possible in 6 PCV2-positive cases, from which 5 cases proved to be PCV2b subtype and 1 case, PCV2a subtype. In conclusion, PCV2 antigen was commonly colocalized with pulmonary vascular lesions in pneumonia in Hungarian swine, and PCV2b was the dominant subtype.  相似文献   

4.
To obtain information about the porcine circovirus type 2 (PCV2) infection status of pigs in Cuba and the probable association of PCV2 with other porcine viruses, tissue samples collected from ill pigs were evaluated using polymerase chain reaction (PCR). The PCR analysis showed that 67.7% of the samples (23/34) from seven swine herds of six different geographic regions were detected to be positive for PCV2. Ten of the 23 PCV2 positive samples (43.5%) shown a concurrent infection with porcine parvovirus (PPV) and 17 of 23 PCV2 positive samples (73.9%) exhibited a concomitant infection with classical swine fever virus (CSFV). This study is the first report of PCV2 infecting pigs with different clinical conditions in Cuban swine herds and provides evidence of PCV2 co-infection with PPV and CSFV in the field.  相似文献   

5.
山东德州某猪场发生猪高热、呼吸系统疾病甚至死亡的疫情。采集病料提取病变组织总DNA或RNA进行猪繁殖与呼吸综合征病毒、猪圆环病毒2型、猪细小病毒、猪伪狂犬病病毒、猪瘟病毒的PCR或RT-PCR检测。PCR扩增出353 bp的猪圆环病毒2型特异性条带。同时进行细菌分离培养、生化鉴定等试验,诊断为猪圆环病毒2型和大肠杆菌、溶血葡萄球菌混合感染。  相似文献   

6.
猪圆环病毒2型IgM抗体间接ELISA的建立   总被引：1，自引：0，他引：1  
猪圆环病毒2型(PCV2)是断奶仔猪多系统衰竭综合症的重要原发性病原,其感染的早期检测有助于及时采取防控措施。以重组衣壳蛋白(Cap蛋白)作为包被抗原,通过对反应条件的优化,建立了PCV2 IgM抗体间接ELISA,其最适抗原包被浓度为1.25 μg／mL,最适血清稀释度为1∶100,临界值为0.35。该检测方法与其他5种常见猪疫病阳性血清无交叉反应,特异性良好。批内、批间重复试验的变异系数均在2%~6%,重复性好。对100份临床样本的检测结果表明,该检测方法与国外同类试剂盒相比,敏感性为90.3%,特异性为92.8%,总符合率为92%。试验结果表明,所建立的PCV2 IgM间接ELISA特异性好和敏感性高,适用于PCV2感染早期的流行病学调查。  相似文献   

7.
A serological survey of selected pathogens in wild boar in Slovenia   总被引：6，自引：0，他引：6  
Serum samples collected from 178 shot wild boars (Sus scrofa) were tested for the presence of antibodies against classical swine fever virus, Aujeszky's disease virus (ADV), porcine reproductive and respiratory syndrome virus, porcine respiratory coronavirus (PRCV), transmissible gastroenteritis virus, swine influenza virus, porcine parvovirus (PPV), swine vesicular disease virus, Actinobacillus pleuropneumoniae (APP), Mycoplasma hyopneumoniae, Salmonella spp., Brucella spp. and Haemophilus parasuis (HPS) throughout Slovenia during the hunting season 2003/2004. The number of samples corresponds to 3% of the total hunting bag. By enzyme-linked immunosorbent assay (ELISA) antibodies against ADV were detected in 55 sera (31%), against PRCV in five sera (3%), PPV in 87 sera (49%), APP in 93 sera (52%), M. hyopneumoniae in 38 sera (21%), Salmonella spp. in 85 sera (47%) and HPS in 33 sera (18%).  相似文献   

8.
Porcine circovirus (PCV) was recently divided into 2 antigenically distinct types that differ (65% amino acid identity) in the protein encoded by open reading frame 2 (ORF2). Porcine circovirus 1 is apparently non-pathogenic and, in contrast, PCV2 is associated with porcine multisystemic wasting syndrome (PMWS). Our objective was to determine the extent of exposure of normal pigs in Canada and Costa Rica to PCV2. Recombinant DNA techniques were used to produce an antigen from ORF2 of PCV2 that was suitable for the detection of antibody in swine sera. The presence of PCV2 nucleotide sequences was detected using polymerase chain reaction (PCR) techniques. Using these tests, specific antibody and nucleotide sequences were demonstrated in sera from a cohort of pigs during a PMWS outbreak. Antibody was detected in normal, healthy hogs slaughtered in Canada (82.4% of 386) and in Costa Rica (14.6% of 322). This is the first report indicating the presence of PCV2 in Latin America. More than 50% of these sera also contained PCV2 nucleotide sequence. Although these hogs were healthy when slaughtered, they were infected with PCV2 and may have previously been ill. The widespread occurrence of PCV2 in swine suggests that this virus is adapted to replication in porcine tissue.  相似文献   

9.
The objective of this work was to investigate the susceptibility of rabbits and mice experimentally inoculated with porcine circoviruses type 1 (PCV1) and type 2 (PCV2) to infection and development of disease and/or lesions. Forty six New Zealand rabbits and 50 ICR-CDI mice were both divided into two groups comprising PCVI and PCV2 inoculated animals, and a third group inoculated with non-infected cell culture medium. Rabbits were inoculated intranasally while mice were inoculated intraperitoneally. Clinical signs and body weights were recorded at the start of the experiment and at necropsy. Animals were bled, euthanised and necropsied at days 0, 3, 7, 10, 14 and 20 post-inoculation and samples were collected for histopathological, serological, in situ hybridisation and PCR analysis. No clinical signs or gross and microscopic lesions compatible with PCV2 infections such as those seen in pigs were observed. No presence of PCV2 nucleic acid was detected in rabbits and mice by in situ hybridisation. Only one mouse inoculated with PCV1 seroconverted on day 20 P1. PCV1 and PCV2 genome was detected in serum by PCR in mice inoculated with each porcine circovirus, while rabbits were negative for both viral types. These studies indicated that porcine circoviruses did not cause any disease or microscopic lesions in inoculated rabbits and mice during the experimental period. However, intraperitoneally inoculated mice might have harboured PCV2 in circulation without evidence of viral replication.  相似文献   

10.
A cross-sectional serological study was conducted in Bhutan between October 2011 and February 2012 to determine the prevalence of antibodies to classical swine fever virus (CSFV), porcine reproductive and respiratory syndrome virus (PRRSV), porcine circovirus type 2 (PCV2), swine influenza virus (SIV) subtype H1N1 and Aujeszky's disease virus (ADV). Furthermore, risk factors for the seropositive status were investigated.  相似文献   

11.
根据GenBank登录的猪伪狂犬病毒（PRV）、猪圆环病毒2型（PCV2）和猪繁殖与呼吸综合征病毒（PRRSV）的参考基因序列,设计3对引物分别用于扩增PCV2的ORF2基因、PRV的gE基因、PRRSV的N基因的目的片段,通过优化反应中各个影响因素,建立了PRV、PCV2、PRRSV的多重PCR（mPCR）检测方法。敏感性和特异性的结果表明,该方法对这3种病毒的最低核酸检出量分别为32.5（PRV）、25.2（PCV2）、35.9pg（PRRSV）。该方法对猪流感病毒（SIV）、猪圆环病毒1型（PCV1）、大肠杆菌、猪瘟病毒（CSFV）、猪流行性腹泻病毒（TGE）等病毒的检测结果均为阴性。200份临床样品的多重PCR结果表明,PCV2感染率为80%（160/200）,PRV感染率为21%（42/200）,PRRSV的感染率为78%（156/200）。200份临床样品主要为PCV2和PRRSV混合感染,阳性率达56.0%（112/200）。该方法的建立对这3种病毒病的早期快速检测和指导临床实践具有十分重要的意义。  相似文献   

12.
本文利用PCR或RT-PCR技术连续两年对规模猪场采集的猪全血分别进行猪瘟病毒（CSFV）、猪繁殖障碍综合征病毒（PRRSV）、猪伪狂犬病病毒（PRV）、猪圆环病毒2型（PCV2）、牛病毒性腹泻病毒（BVDV）和日本乙型脑炎病毒（JEV）的检测,结果发现2008年猪全血中CSFV、PRRSV、PRV、PCV2的感染率为30.0%、50.0%、4.0%和30.0%,2009年猪全血中CSFV、PRRSV、PRV、PCV2、BVDV和JEV的感染率分别为19.8%、13.7%、1.1%、31.9%、0.7%和2.7%,这表明用猪的全血可进行猪瘟等6种疫病的监测和流行,为更深入研究疫病的流行病学奠定了基础。  相似文献   

13.
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14.
猪圆环病毒分子生物学研究进展   总被引：1，自引：0，他引：1  
猪圆环病毒（PCV）是单链环状DNA病毒。它是近年来阻碍养猪业发展的重要传染病,因而成为各国学者的研究热点。本文就猪圆环病毒的基因组结构与功能,主要编码的蛋白质及其功能,分子生物学诊断方法等的最新研究进展作一综述。  相似文献   

15.
Serum samples from 1,133 dairy cows (187 herds), 3,712 ewes (103 flocks) and 1,317 adult pigs (877 herds), were tested for neutralizing antibodies against the NADL strain of bovine virus diarrhoea virus. The prevalence rate of seropositive animals was 18.5% in cattle, 4.5% in sheep and 2.2% in pigs, such seroreactors being found in 28% of the cattle herds and 18% of the sheep flocks. In all three species the rate showed considerable herd and geographical variation. In cattle the seroreactor rate was similar in herds with normal reproduction and in 62 herds with problems of repeat breeding. Of 31 pig sera containing antibodies against the NADL strain, 27 were also positive in a neutralization test for antibodies against swine fever virus (Baker strain). However, all sera showed a higher titre of antibodies against the bovine strain than against the swine fever virus. It was concluded that the immune response of the pigs had been induced by ruminant pestivirus, and not by swine fever virus.  相似文献   

16.
为了解浙江省猪主要病毒性传染病的流行情况及变化规律,本试验对2017-2020年浙江省不同地区规模化猪场送检的血清和病料样品进行猪繁殖与呼吸综合征病毒(PRRSV)、猪瘟病毒(CSFV)、猪圆环病毒2型(PCV2)和猪伪狂犬病病毒(PRV)4种疫病病原和抗体检测。抗体检测结果显示：PRRSV、CSFV、PCV2、PRV-gE/gI、PRV-gB的抗体平均阳性率分别为81.4%,83.6%,91.4%,36.1%,94.3%;不同年份中,CSFV和PCV2抗体水平一直呈稳中上升趋势,2020年PRRSV和PRV-gE/gI抗体水平下降明显。病原检测结果显示：PRRSV、CSFV、PCV2和PRV的感染具有普遍性,病原平均阳性率分别为22.9%,2.5%,17.3%,6.6%,其中PRRSV/PCV2双重感染较多,阳性率为9.7%;不同日龄中,PRRSV、PCV2和CSFV保育猪病原阳性率最高,育肥猪次之,而PRV产房仔猪阳性率最高;此外,2019-2020年CSFV、PCV2和PRV的病原阳性率下降明显。  相似文献   

17.
随着规模化养猪业的发展,传染性疾病越来越多,混合感染或多重感染十分普遍,给疾病的诊断和预防带来很大困难。本文将我们实验室2010年分别采自福建、广西、河南、上海、内蒙、浙江、江苏和山西等地发病猪场的185份病料,通过RT-PCR和PCR方法对其进行猪繁殖与呼吸综合征病毒（Porcine reproductive and respiratory syndrome virus,PRRSV）、猪繁殖障碍的猪瘟病毒（Classical swine fever virus,CSFV）、猪圆环病毒2（Porcine circovirus type 2,PCV2）、猪细小病毒（Porcine parvovirus,PPV）、猪伪狂犬病毒（Pseudorabies virus,PRV）、细环病毒2（Torque teno virus 2,TTV2）等病毒检测。结果表明在所检的病料中PRRSV、PCV2和TTV2的阳性率比较高,分别为49.2%、62.2%和95.1%。有些地方TTV2的阳性率高达100%;同时,还存在很普遍的PRRSV与PCV2、PRRSV与TTV2、PCV2与TTV2等混合感染,混合感染率分别为27.6%、45.4%、58.4%;以及PRRSV、PCV2与TTV2的三重感染率为24.9%。同时对部分PRRSV阳性病料的PRRSV GP5和nsp2基因分别进行测序,分析测序结果表明病料中的PRRSV的GP5和nsp2序列与PRRSV HuN4株的相应序列的亲缘关系分别在98.2%和96.2%以上,且在nsp2区域都存在30个不连续氨基酸的缺失,说明现在临床中所流行的PRRSV可能仍是与高致病性PRRSV基因型相似的病毒。通过本文可以及时了解当前养猪场的病毒性疾病的流行情况,为猪场病毒性疾病的防控提供依据。  相似文献   

18.
The objective of the present study was to determine the presence of recognised abortifacient viruses such as porcine reproductive and respiratory virus (PRRSV), Aujeszky's disease virus (ADV), porcine parvovirus (PPV) and porcine circovirus type 2 (PCV2), in tissues from aborted fetuses and stillborn neonates in cases of late reproductive failure in swine. A total of 293 specimens (fetuses aborted in the last third of gestation and stillborn piglets) from 100 different cases of late-term abortions and premature farrowing from 15 different Spanish provinces were studied. PRRSV was detected in 9/100 cases by RT-PCR. Only 1/100 cases analysed (corresponding to a late-term aborted fetus with a negative PRRSV RT-PCR result) was positive for PCV2 by PCR. Neither ADV (monitored by viral isolation plus antigen detection) nor PPV (monitored by ELISA antigen capture test) infection was identified. The results suggest that PRRSV is one of the most important infectious agents, if not the most relevant one, associated with fetal infection leading to abortion or premature farrowing in Spain. Moreover, other viral pathogens such as ADV, PPV and PCV2 seem to have a minor impact on reproductive disease.  相似文献   

19.
In order to examine an association between porcine circovirus type-2 (PCV2) infection and reproductive failure in pigs, sera (n = 171) from stillborn fetuses were collected from 3 different farms with prolonged histories of reproductive problems. These sera were tested for the presence of antibodies to PCV2 using an immunoperoxidase monolayer assay. Of the 171 sera tested, 28 had PCV2 antibody titers of ≥ 1:16. When these 28 samples were tested by a polymerase chain reaction assay, 13 were found to contain PCV2 viral DNA. Of these 13 samples containing both PCV2 antibodies and viral DNA, 9 yielded PCV2 on virus isolation. Amino acid sequences comprising open reading frame 2 of PCV2 from 2 of these isolates were compared to PCV2 isolates from cases of post-weaning multi-systemic wasting syndrome (PMWS). The amino acid sequences of the 2 isolates from stillborn pigs were shown to be nearly identical to each other, as well as to other PCV2 isolates associated with reproductive failure. When compared with PMWS isolates, the isolates from the stillborn fetuses showed differences of at least 2 amino acids. These results confirm previous findings that transplacental infection of PCV2 occurs in the field and that stillbirths in pigs may be associated with PCV2 infections. At present, the significance of minor differences in amino acid sequences is not known.  相似文献   

20.
5种猪病多重PCR检测方法的建立   总被引：1，自引：0，他引：1  
To establish a method for simultaneous detection of classical swine fever virus (CSFV), porcine reproductive and respiratory syndrome virus (PRRSV), pseudorabies virus (PRV), porcine circovirus type 2 (PCV2) and porcine parvovirus (PPV), a multiplex PCR was developed with a set of specific primers designed based on the conserved sequences of CSFV, PRRSV, PRV, PCV2 and PPV. Under the optimized conditions of multiplex PCR,five special fragments of 167 (CSFV),433 (PRRSV),305 (PRV), 559 (PCV2) and 882 bp (PPV) were amplified with a detection limit of 220, 1.6, 72, 400 and 370 pg, respectively. But the multiplex PCR amplification results of swine influenza virus （SIV）, Japanese encephalitis virus （JEV）, Streptococcus suis （SS） and porcine epidemic diarrhea virus （PEDV） were negative．The results showed that the multiplex PCR method was capable of CSFV, PRV, PRRSV, PCV2, PPV infection of single or mixed clinical samples for rapid diagnosis.  相似文献   

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