Postweaning multisystemic wasting syndrome (PMWS) is an important swine disease that is closely associated with porcine circovirus type 2 (PCV2). The capsid protein (Cap protein) is a major structural protein that has...Postweaning multisystemic wasting syndrome (PMWS) is an important swine disease that is closely associated with porcine circovirus type 2 (PCV2). The capsid protein (Cap protein) is a major structural protein that has at least three immunoreactive regions, and it can be a suitable candidate antigen for detecting the specific antibodies of a PCV2 infection. In the present study, an indirect enzyme-linked immunosorbent assay (TcELISA) based on a truncated soluble Cap protein produced in Escherichia coli (E.coli) was established and validated for the diagnostic PCV2 antibodies in swine. The TcELISA was validated by comparison with an indirect immunofluorescence assay (IIFA). The diagnostic sensitivity (DSN), specificity (DSP), and accuracy of the TcELISA were 88.6%, 90.7% and 89.4%, respectively. The agreement rate was 89.38% between results obtained with TcELISA and IIFA on 113 field sera. A cross-reactivity assay showed that the method was PCV2-specific by comparison with other sera of viral disease. Therefore ,the TcELISA will be helpful for the development of a reliable serology diagnostic test for large scale detection of PCV2 antibodies and for the evaluation of vaccine against PCV2 in swine.展开更多
Objective] The passing rates of porcine reproductive and respiratory syn-drome virus （PRRSV） vaccine and classical swine fever virus （CSFV） vaccine were investigated in this paper. The PRRSV vaccine was injected i...Objective] The passing rates of porcine reproductive and respiratory syn-drome virus （PRRSV） vaccine and classical swine fever virus （CSFV） vaccine were investigated in this paper. The PRRSV vaccine was injected in 14-d-old piglets and the CSFV vaccine was injected in 25-d-old piglets. [Method] The antibody expres-sion amounts were detected with the PRRSV antibody ELlSA detection kit and the CSFV antibody ELlSA detection kit. [Result] The antibody positive rates were 6.7%, 26.7%, 85% and 100% 3, 7, 14 and 30 d after the vaccination of PRRSV vaccine. While for the CSFV vaccine, 13%, 40%, 58.3% and 83.3% of the immune individu-als were antibody positive 3, 7, 14 and 30 d after the vaccination. [Conclusion] The PRRSV vaccine showed the lowest antibody tilter 3 d after the inoculation, but had a great vaccination effect 30 d after the inoculation. The piglets were not immune enough against viral invasion 3 d after the inoculation of CSFV vaccine. Although the antibody positive rate of PRRSV and CSFV vaccines al reached the required level by the Ministry of Agriculture （70%） 30 d after the inoculation, the vaccination of CSFV vaccine could not achieve the ideal effect of 100%.展开更多
In this study, the coding region of type O FMDV capsid protein VP1 and a series of codon optimized DNA sequences coding for VP1 amino acid residues 141-160 （epitopel）, tandem repeat 200-213 （epitope2 （＋2）） and ...In this study, the coding region of type O FMDV capsid protein VP1 and a series of codon optimized DNA sequences coding for VP1 amino acid residues 141-160 （epitopel）, tandem repeat 200-213 （epitope2 （＋2）） and the combination of two epitopes （epitopel-2） was genetically cloned into the prokaryotic expression vector PPRoExHTb and pGEX4T-1, respectively. VP1 and the fused epitopes GST-E1, GST-E2 （＋2） and GST-E1-2 were successfully solubly expressed in the cytoplasm of Escherichia coli and Western blot analysis demonstrated they retained antigenicity. Indirect VP1-ELISA and epitope ELISAs were subsequently developed to screen a panel of 80 field pig sera using LPB-ELISA as a standard test. For VP1-ELISA and all the epitope ELISAs, there were clear distinctions between the FMDV-positive and the FMDV-negative samples. Cross-reactions with pig sera positive to the viruses of swine vesicular disease virus that produce clinically indistinguishable syndromes in pigs or guinea pig antisera to FMDV strains of type A, C and Asia1 did not occur. The relative sensitivity and specificity for the GST-E1 ELISA, GST-E2 （＋2）, GST-E1-2 ELISA and VP1-ELISA in comparison with LPB-ELISA were 93.3% and 85.0%, 95.0% and 90%, 100% and 81.8%, 96.6% and 80.9% respectively. This study shows the potential use of the aforementioned epitopes as alternatives to the complex antigens used in current detection for antibody to FMDV structural proteins.展开更多
基金supported by a project from National Key Technology R&D Program in the 11th Fiveyear Plan of China (2006BAD06A12)
文摘Postweaning multisystemic wasting syndrome (PMWS) is an important swine disease that is closely associated with porcine circovirus type 2 (PCV2). The capsid protein (Cap protein) is a major structural protein that has at least three immunoreactive regions, and it can be a suitable candidate antigen for detecting the specific antibodies of a PCV2 infection. In the present study, an indirect enzyme-linked immunosorbent assay (TcELISA) based on a truncated soluble Cap protein produced in Escherichia coli (E.coli) was established and validated for the diagnostic PCV2 antibodies in swine. The TcELISA was validated by comparison with an indirect immunofluorescence assay (IIFA). The diagnostic sensitivity (DSN), specificity (DSP), and accuracy of the TcELISA were 88.6%, 90.7% and 89.4%, respectively. The agreement rate was 89.38% between results obtained with TcELISA and IIFA on 113 field sera. A cross-reactivity assay showed that the method was PCV2-specific by comparison with other sera of viral disease. Therefore ,the TcELISA will be helpful for the development of a reliable serology diagnostic test for large scale detection of PCV2 antibodies and for the evaluation of vaccine against PCV2 in swine.
基金Supported by Agricultural Science and Technology Achievement Transformation Fund of Ministry of Science and Technology(2012GB2A200045)Fund Project of Hebei Province Department of Education(2008449)Special Project of Qinhuangdao Municipal Bureau of Science and Technology(201101A181)~~
文摘Objective] The passing rates of porcine reproductive and respiratory syn-drome virus （PRRSV） vaccine and classical swine fever virus （CSFV） vaccine were investigated in this paper. The PRRSV vaccine was injected in 14-d-old piglets and the CSFV vaccine was injected in 25-d-old piglets. [Method] The antibody expres-sion amounts were detected with the PRRSV antibody ELlSA detection kit and the CSFV antibody ELlSA detection kit. [Result] The antibody positive rates were 6.7%, 26.7%, 85% and 100% 3, 7, 14 and 30 d after the vaccination of PRRSV vaccine. While for the CSFV vaccine, 13%, 40%, 58.3% and 83.3% of the immune individu-als were antibody positive 3, 7, 14 and 30 d after the vaccination. [Conclusion] The PRRSV vaccine showed the lowest antibody tilter 3 d after the inoculation, but had a great vaccination effect 30 d after the inoculation. The piglets were not immune enough against viral invasion 3 d after the inoculation of CSFV vaccine. Although the antibody positive rate of PRRSV and CSFV vaccines al reached the required level by the Ministry of Agriculture （70%） 30 d after the inoculation, the vaccination of CSFV vaccine could not achieve the ideal effect of 100%.
文摘In this study, the coding region of type O FMDV capsid protein VP1 and a series of codon optimized DNA sequences coding for VP1 amino acid residues 141-160 （epitopel）, tandem repeat 200-213 （epitope2 （＋2）） and the combination of two epitopes （epitopel-2） was genetically cloned into the prokaryotic expression vector PPRoExHTb and pGEX4T-1, respectively. VP1 and the fused epitopes GST-E1, GST-E2 （＋2） and GST-E1-2 were successfully solubly expressed in the cytoplasm of Escherichia coli and Western blot analysis demonstrated they retained antigenicity. Indirect VP1-ELISA and epitope ELISAs were subsequently developed to screen a panel of 80 field pig sera using LPB-ELISA as a standard test. For VP1-ELISA and all the epitope ELISAs, there were clear distinctions between the FMDV-positive and the FMDV-negative samples. Cross-reactions with pig sera positive to the viruses of swine vesicular disease virus that produce clinically indistinguishable syndromes in pigs or guinea pig antisera to FMDV strains of type A, C and Asia1 did not occur. The relative sensitivity and specificity for the GST-E1 ELISA, GST-E2 （＋2）, GST-E1-2 ELISA and VP1-ELISA in comparison with LPB-ELISA were 93.3% and 85.0%, 95.0% and 90%, 100% and 81.8%, 96.6% and 80.9% respectively. This study shows the potential use of the aforementioned epitopes as alternatives to the complex antigens used in current detection for antibody to FMDV structural proteins.
