Establishment and application of an indirect ELISA for porcine circovirus 3

Deng, Junhua; Li, Xiangdong; Zheng, Dingding; Wang, Yanwei; Chen, Lingyan; Song, Huanhuan; Wang, Tongyan; Huang, Yuxin; Pang, Wei; Tian, Kegong

doi:10.1007/s00705-017-3607-7

Cited by 43 publications

(50 citation statements)

References 8 publications

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“…Fu et al (2018) reported that PCV3 was positive in 26.7% of their samples [24]. By seroprevalence, Deng et al (2018) reported that the PCV3 positive rate increased from 22.3% in 2015 to 51.8% in 2017 [26]. A recent US report revealed that 56.6% of their serum samples were positive [7].…”

Section: Discussionmentioning

confidence: 99%

“…Variations of the ELISA method used, particularly the purity of the coating antigen could have significant impact on determination of the cutoff values to differentiate positive samples from negative ones. Deng et al (2018) set the cutoff value at 0.31 with average absorbance of 0.29 from 20 serum samples from cesarean-derived and colostrum-deprived pigs used as negative controls [26]. Palinski et al reported the cutoff value at 0.87 (with mean OD at 0.49 of 18 serum samples from PCV3negative SPF pigs [7].…”

Section: Discussionmentioning

confidence: 99%

“…To detect the PCV3 antibodies in serum samples, indirect ELISA was employed as previously described [26] using overnight coating, at 4°C, of the purified capsid protein coating (2 μg/well) on EIA/RIA high binding plates (Corning, USA). The plates were then blocked with 0.1% casein for 2 h at 37°C.…”

Section: Indirect Elisa For Detection Of Pcv3 Antibodiesmentioning

confidence: 99%

“…The plates were measured at OD 450nm on the microplate reader (Epoch, Biotek, USA). The cutoff value differentiating positive and negative serum samples was set at 0.53 OD 450nm , that is, three units of standard deviation above the mean of the negative control sera [7,26] (average OD 450nm = 0.284 tested on 40 serum samples from clinically healthy pigs with standard deviation of 0.083). The coating antigen PCV3 cap was also tested on PCV2-positive serum samples to examine the specificity of closely related virus infection using the established ELISA protocol.…”

Section: Indirect Elisa For Detection Of Pcv3 Antibodiesmentioning

confidence: 99%

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Prevalence of porcine circovirus type 3 in pigs in the southeastern Chinese province of Zhejiang

et al. 2019

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Background Porcine circovirus type 3 (PCV3) was first reported in US in 2016. The virus was also identified later in China. Prevalence of PCV3 in Zhejiang province in southeastern China is not clear though it has been reported in many parts of China. Results PCV3 infection and its co-infection with other swine viral pathogens in pig herds of Zhejiang province were retrospectively investigated by quantitative PCR (qPCR) and its sero-prevalence by indirect ELISA. PCV3 was found positive in 67.1% of the 283 clinical samples taken from 2014 to 2017 as shown by qPCR. Single infection with PCV3 accounted for only one-third of the samples, and majority were of co-infections, predominantly with PEDV (41.6%) but generally low with other swine viruses. Indirect ELISA using the PCV3 capsid protein as the coating antigen revealed an average sero-positive rate of 52.6% (40.8 to 60.8%) in 2345 serum samples from 2011 to 2017, with earliest yet high positive findings in samples taken in 2012. Of 203 serum samples, the qPCR method showed more positive findings than ELISA (81.3% vs 56.2%). With 89 serum samples negative by ELISA, vast majority ( n = 81) were found positive by qPCR. There was negative correlation in levels of PCV3 DNA and anti-capsid antibody response. ORF2-based phylogenetic analysis revealed three major groups (PCV3a, PCV3b and PCV3c) of the 200 strains, 38 from this study and 162 reference strains from GenBank. Most of the strains from this study were clustered into PCV3c. Of the putative signature residues of the capsid protein (aa 24, 27, 77 and 150) relative to the three groups, only the PCV3a group strains showed a distinct pattern of residues VKSI (95% of the strains), while the other two groups did not have such a ‘signature’ pattern. Conclusions Results from this study provided further evidence that the novel virus PCV3 was widely distributed in China and might have emerged in Zhejiang province before 2014, most probably back in 2012 when there was high PCV3 sero-prevalence. PCV3 might be viremic in pigs and could spread by fecal shedding. Electronic supplementary material The online version of this article (10.1186/s12917-019-1977-7) contains supplementary material, which is available to authorized users.

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Indirect Elisa For Detection Of Pcv3 Antibodiesmentioning

confidence: 99%

Section: Indirect Elisa For Detection Of Pcv3 Antibodiesmentioning

confidence: 99%

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Prevalence of porcine circovirus type 3 in pigs in the southeastern Chinese province of Zhejiang

et al. 2019

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“…For example, PCV2 VLPs are used as coating antigens to be able to detect serum neutralizing antibodies (SNAbs) in ELISA (Nainys et al 2014;Zhang et al 2016). Currently, PCV3 Cap proteins harvested from insect cells or E. coli have been used as antigens to detect PCV3-specific antibodies in an ELISA (Deng et al 2018;Zhang et al 2019). There is still lacking a VLP-based serological diagnosis method for PCV3.…”

Section: Introductionmentioning

confidence: 99%

Self-assembly into virus–like particles of the recombinant capsid protein of porcine circovirus type 3 and its application on antibodies detection

Wang

Duan

et al. 2020

AMB Expr

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PCV3 capsid protein (Cap) is an important antigen for diagnosis and vaccine development. To achieve high-level expression of recombinant PCV3 Cap in Escherichia coli (E. coli), the gene of wild-type entire Cap (wt-eCap) was amplified from clinical samples, and three optimized entire Cap (opti-eCap) and one optimized Cap deleted nuclear location signal (NLS) (opti-dCap) gene fragments encoding the same amino acid sequence with wt-eCap were synthesized based on the codon bias of E. coli. Those gene fragments were inserted into the pET30a expression vector. One recombinant strain with the highest expressed soluble eCap from four entire Cap (one wt-eCap and three opti-eCap) and one recombinant strain expressed opti-dCap were selected for further purification. The purified eCap and dCap were identified by transmission electron microscopy (TEM), a large number of round hollow particles with a diameter of 10 nm virus-like particles (VLPs) were observed in eCap, whereas irregular aggregation of proteins observed in dCap. After formation the VLPs were applied as a coating antigen to establish an indirect ELISA (I-ELISA) for detection of PCV3-specific antibody in swine serum. 373 clinical swine serum samples from China collected in 2019 were tested utilizing the VLP-based I-ELISA method under optimized conditions. To the best of our knowledge, this is the first report of self-assembly into VLPs of PCV3 recombinant Cap. Our results demonstrated that the VLP-based I-ELISA will be a valuable tool for detecting the presence of PCV3 antibodies in serum samples and will facilitate screening of large numbers of swine serum for clinical purposes.

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Fine mapping of linear B cell epitopes on capsid protein of porcine circovirus 3

Jiang

Guo

Zhang

et al. 2020

Appl Microbiol Biotechnol

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Establishment and application of an indirect ELISA for porcine circovirus 3

Cited by 43 publications

References 8 publications

Prevalence of porcine circovirus type 3 in pigs in the southeastern Chinese province of Zhejiang

Prevalence of porcine circovirus type 3 in pigs in the southeastern Chinese province of Zhejiang

Self-assembly into virus–like particles of the recombinant capsid protein of porcine circovirus type 3 and its application on antibodies detection

Fine mapping of linear B cell epitopes on capsid protein of porcine circovirus 3

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