Postweaning Multisystemic Wasting Syndrome Research Articles

Basigin-CyP elevated porcine circovirus type2 replication

Porcine circovirus type2 (PCV2) is a member of the circoviridae family. PCV2 was identified as the main pathogen of postweaning multisystemic wasting syndrome (PMWS) in weaned piglets and causes massive economic loss. Basigin, is a transmembrane glycoprotein belonging to the immunoglobulin superfamily; which is also a receptor for cyclophilins. CyP belongs to the immunophilin family that has peptidyl-prolyl cis-trans isomerase activity. Basigin-CyP interaction affects the replication stages of several viruses. In this study, we found that Basigin could elevate the replication of PCV2, and the Basigin only affected the replication stage rather than adsorption or endocytosis stages. In addition, the ligands of Basigin, CyPA and CyPB also elevated the replication of PCV2. Basigin-CyP interation was necessary for elevating PCV2 replication; At last, CyPs were proved to promote the replication of PCV2 by activating ERK signaling.

Replacing the decoy epitope of PCV2 capsid protein with epitopes of GP3 and/or GP5 of PRRSV enhances the immunogenicity of bivalent vaccines in mice

Porcine circovirus type 2 (PCV2) is the causative agent of postweaning multisystemic wasting syndrome (PMWS), porcine dermatitis and nephropathy syndrome (PDNS), and reproductive failure and causes economic losses in the domestic swine industry. The decoy epitope (169–180 amino acid (aa)) of the PCV2 capsid (Cap) protein is an immunodominant epitope and diverts the immune response away from protective epitopes. The mixed infection of PCV2 and porcine reproductive and respiratory syndrome virus (PRRSV) is one of the most common co-infections in the pig industry and shows more severe clinical symptoms. Linear B-cell antigenic epitopes of PRRSV GP3 epitope Ⅰ (61–72aa) and PRRSV GP5 epitope Ⅳ (187–200aa) efficiently elicited neutralizing antibodies against PRRSV. The recombinant baculovirus expressing the Cap protein (Bac-Cap) was modified by replacing the decoy epitope of the Cap protein with either the PRRSV GP3 epitope Ⅰ, the PRRSV GP5 epitope Ⅳ, or the PRRSV GP3 epitope Ⅰ- GP5 epitope Ⅳ to produce the recombinant baculoviruses Bac-Cap-GP3, Bac-Cap-GP5 and Bac-Cap-GP35. The four recombinant baculoviruses were successfully established and characterized as demonstrated with western blot analysis and immunofluorescence assay. Immunogenicities of the four recombinant baculoviruses in mice were tested in sera harvested at 21 and 42 days post-primary immunization. The titers of antibodies in the sera were determined by a PCV2-specific enzyme-linked immunosorbent assay (ELISA) and a serum neutralization assay. The serum IFN-γ levels were measured by indirect ELISA. The results showed that Bac-Cap-GP3, Bac-Cap-GP5, and Bac-Cap-GP35 elicited higher GP3/GP5 and Cap antibody titers than the Bac-Cap. Virus neutralization test also confirmed that the serum from the Bac-Cap-GP3 immunized mice had high levels of the both PCV2 and PRRSV neutralization antibodies. These findings collectively demonstrated that substituting the decoy epitope of the PCV2 capsid substituted with PRRSV epitopes could be developed into an effective vaccine against PCV2.

Cross-Sectional Study on the Sero- and Viral Dynamics of Porcine Circovirus Type 2 in the Field.

Porcine circovirus-associated diseases (PCVADs) cause considerable economic losses in industrial pork production in the field. To minimize the economic losses due to PCVAD, porcine circovirus type 2 (PCV2) vaccines have been developed, and there is widespread vaccination worldwide today. However, limited information is available concerning the current status of PCV2 infection in the field on the Asian continent. The present study aimed to assess sero- and viral dynamics of PCV2 from 12 PCV2-contaminated pig herds with vaccination against PCV2 in Southern and Central Taiwan. In particular, the level of PCV2 load during the window period for seroconversion using real-time polymerase chain reaction and a commercial enzyme-linked immunosorbent assay (ELISA) kit. Our results revealed that pig herds showed slight or no seroconversion after three to four weeks post-PCV2 immunization. The presence of PCV2 was observed during the window period for seroconversion in all herds. In conclusion, natural exposure of PCV2 occurs in the growing to fattening period, and viremia can last until slaughter. Additionally, our findings indicate that using ELISA showed the level of antibodies and aided in the understanding and surveillance of the current PCV2 status in the field.

Comparison of Multiplex Real-Time PCR and PCR-Reverse Blot Hybridization Assays for the Direct and Rapid Detection of Porcine Circovirus Type 2 Genotypes

Porcine circovirus type 2 (PCV2), the causative agent of porcine circovirus-associated diseases (PCVAD), poses a serious economic threat for the swine industry. Currently, PCV2 is classified into five major genotypes: PCV2a, PCV2b, PCV2c, PCV2d, and PCV2e. The aim of this study is to evaluate the performance of two commercially available methods, multiplex real-time PCR assay and PCR-reverse blot hybridization assay (REBA), for the rapid detection of PCV2 and direct identification of PCV2 genotypes from clinical samples as well as to compare the results with that of sequence analysis. Molecular diagnostic methods were used to evaluate a total of 180 samples, including tissues and blood samples from pigs that were suspected of PCVAD infection. The results of this study showed that the detection rate for positive PCV2 was 48.3% (n = 87) in both multiplex real-time PCR and PCR-REBA methods. Using sequence analysis, which is the gold standard, and multiplex real time PCR assay, the sensitivity, specificity, positive predictive value, and negative predictive value of PCV2 genotyping were found to be 97.1% (n = 67, 95% CI 0.894–0.998, p < 0.001), 100% (n = 93, 95% CI 0.966–1.000, p < 0.001), 100% (95% CI 0.953–1.000, p < 0.001), 97.9% (95% CI 0.921–0.998, p < 0.001), respectively. The results of PCR-REBA were found to be consistent with those of sequence analysis for all the samples and showed good agreement (κ = 1). The most prevalent genotypes detected in this study were PCV2d (n = 53, 60.9%), followed by PCV2a (n = 17, 19.5%), PCV2b (n = 14, 16.1%), and PCV2a/b co-infection (n = 3, 3.5%). Both the methods required ~3 h for completion. Therefore, we conclude that two molecular methods are rapid and reliable for the characterization of the causative pathogen with PCV2 genotypes.

Reduction of Postweaning Multisystemic Wasting Syndrome-Associated Clinical Symptoms by Virus-Like Particle Vaccine Against Porcine Parvovirus and Porcine Circovirus Type 2.

The porcine circovirus type 2 (PCV2) capsid (Cap) protein and porcine parvovirus (PPV) VP2 protein have been studied in vaccines to control postweaning multisystemic wasting syndrome (PMWS). Virus-like particle (VLP) vaccines are nonreplicative vectors that deliver epitopes and induce immune responses. However, most VLP vaccines are recombinant proteins expressed in eukaryotic systems and are expensive and complex. In this study, the full-length PCV2-Cap and PPV-VP2 proteins were expressed in Escherichia coli, which self-assembled into VLPs. The highly soluble proteins were purified using Ni-chelating affinity chromatography. The proteins self-assembled into VLPs of ∼20 nm (Cap VLP) and 25 nm (VP2 VLP) in diameter. The immunogenicities of Cap VLP and VP2 VLP were determined in piglets coinfected with PPV and PCV2 postimmunization. The results suggested that Cap VLP and VP2 VLP did not antagonize each other. The combined vaccine induced stronger humoral and cellular immune responses and provided the best protection against PPV and PCV2 coinfection. On a farm containing PMWS-infected pigs, the combined Cap VLP and VP2 VLP vaccine significantly improved piglet growth indices; the average daily weight gains were significantly higher than those of the Cap VLP vaccine and nonimmunized groups. Thus, Cap and VP2 protein expression in E. coli is feasible for large-scale VLP vaccine production. The combined vaccine may be a promising candidate vaccine for better preventing PMWS-associated diseases coinfected with PCV2 and PPV.

A retrospective study of DNA prevalence of porcine parvoviruses in Mexico and its relationship with porcine circovirus associated disease.

Worldwide, many emerging porcine parvoviruses (PPVs) have been linked to porcine circovirus-2 (PCV2) associated disease (PCVAD), which includes post-weaning multi-systemic wasting syndrome (PMWS), PCV2-related reproductive failure (PCV2-RF), as well as other syndromes. To determine the DNA prevalence of PPVs and their relationship with PMWS and PCV2-RF in Mexico, 170 formalin-fixed paraffin-embedded tissues were selected from archival collections to detect PPVs using a nested polymerase chain reaction. The tissues were composed of 50 PMWS cases, 20 age-matched tissues from healthy pigs, 56 PCV2-related reproductive failure (PCV2+ -RF) cases, and 44 PCV2- -RF cases. Overall, PPV2 and PPV6 were the most prevalent species (90.0% and 74.7%, respectively). In 8-11 week old pigs, the highest prevalence was for PPV6 and PPV3. Concerning reproductive failure, the PCV2-affected farms had a significantly higher prevalence for PPV6 (61.6%) and PPV5 (36.4%) than the PCV2-unaffected farms (35.0% and 5.0%, respectively). The concurrent infection rate was high, being significant for PPV2/PPV4 and PPV1/PPV5 within the PMWS cases and for PPV6/PPV5 among the PCV2+ -RF tissues. PPV5 showed a significant relationship with PMWS, whereas PPV5 and PPV6 were significant for PCVAD. The prevalence and coinfection rate of PPVs in Mexico were markedly higher than that described in other countries, denoting that PPV5 and PPV6 might have a potential role in PCVAD in Mexico. It is concluded that it is likely that the density population of pigs in Mexico is contributing to high PPV inter-species and PCV2 coinfections which might lead to a different pathogenic outcome.

PCV2 co-infection does not impact PRRSV MLV1 safety but enhances virulence of a PRRSV MLV1-like strain in infected SPF pigs

Co-infection by a type 1 modified live vaccine-like strain (MLV1-like) of porcine reproductive and respiratory syndrome virus (PRRSV) and a type 2 porcine circovirus (PCV2) was identified on a French pig farm with post-weaning multisystemic wasting syndrome (PMWS). An in vivo experiment was set up to characterize the virulence level of the MLV1-like strain compared with the parental MLV1 strain, and to assess the impact of PCV2 co-infection on the pathogenicity of both PRRSV strains. Six groups of six pigs each were inoculated only with either one of the two PRRSV strains or with PCV2, or co-inoculated with PCV2 and MLV1 or PCV2 and MLV1-like strains. Six contact pigs were added to each inoculated group to assess viral transmission. The animals were monitored daily for 35 days post-inoculation for clinical symptoms. Blood and nasal swabs were sampled twice a week, and tissue samples were collected during necropsy for viral quantification. Compared to MLV1-infected pigs, animals infected with the MLV1-like strain had increased PRRSV viremia and nasal shedding, a higher viral load in the tonsils, and lymph node hypertrophy at microscopic level. PCV2 co-infection did not influence clinical, virologic or transmission parameters for MLV1, but co-infected MLV1-like/PCV2 pigs had the most severe lung lesions, the highest viremia in contact animals and the highest transmission rate. Our study demonstrated that the MLV1 strain tested was safe when co-inoculated with PCV2 in piglets. However, co-infection by the MLV1-like strain and PCV2 resulted in increased virulence compared with that due to a single infection.

Molecular epidemiology and genetic variation analyses of porcine circovirus type 2 isolated from Yunnan Province in China from 2016-2019

BackgroundPorcine circovirus type 2 (PCV2) is the causative agent of porcine circovirus-associated disease (PCVAD). Its prevalence in swine herds was first reported in China in 2000. PCV2 infection causes immunosuppression that leads to multiple diseases, causing serious economic problems for the swine industry in China. Since information on the genetic variation of PCV2 in Yunnan province is limited, this study aims to investigate the molecular epidemiological and evolutionary characteristics of PCV2 from 2016 to 2019.MethodsA total of 279 clinical samples were collected from different regions of Yunnan between 2016 to 2019, and PCV2 was detected by PCR. We then amplified full genomes from the positive samples, and the sequences were analysed for homology and genetic evolution.ResultsOverall, 60.93% (170/279) of the screened swine herd samples were positive for PCV2. We sequenced 15 Yunnan province PCV2 strains from positive samples. Analyses of the complete genomes and Cap genes led to the classification of the 15 Yunnan PCV2 strains into PCV2a (2 of 15), PCV2b (1of 15) and PCV2d (12 of 15). All strains shared 94.3–99.9% of their identities with the nucleotide sequences of complete genomes in this study and shared 94.2–99.9% identity with the reference sequences. All strains share 89.4–100% and 86.8–100% of their identities with the nucleotide and amino acid (aa) sequences of Cap, respectively.ConclusionsThe results of this study provide evidence that PCV2a, PCV2b and PCV2d genotypes coexisted in Yunnan Province from 2016 to 2019, and the priority prevalence genotype was PCV2d. The data provide evidence for the increased genetic diversity and insights into the molecular epidemiology of PCV2. This study also provides basic data for the Yunnan province PCV2 molecular epidemiological survey and accumulates effective materials for the development of PCV2 vaccines.

Short communication: a modified Vaccinia virus Ankara-based Porcine circovirus 2 vaccine elicits strong antibody response upon prime-boost homologous immunization in a preclinical model.

Porcine circovirus 2 (PCV2) infections are related to a number of syndromes and clinical manifestations, generally known as Porcine circovirus-associated diseases, which are related to losses in the swine industry. There are commercially available vaccines and new vaccines being tested, however, persistency of the PCV2 as an important pig pathogen, and the growing number of affected farms in different countries have suggested that there is room for vaccine improvement. In this study, we describe the construction and testing of a recombinant live vaccine based on a modified Vaccinia virus Ankara (MVA) vector expressing the PCV2b capsid protein (CAP). Using a two-dose homologous vaccination regimen, in mice, we demonstrated that the vaccine induced high titers of anti-PCV2 antibodies. The vaccine is stable upon lyophilization, and, together with the good immunogenicity potential observed, the results support further evaluation of the MVA-CAP vaccine in the target species.

Porcine Circovirus Type 2 ORF5 Protein Induces Autophagy to Promote Viral Replication via the PERK-eIF2α-ATF4 and mTOR-ERK1/2-AMPK Signaling Pathways in PK-15 Cells.

Porcine circovirus type 2 (PCV2) is the primary causative agent that causing porcine circovirus-associated disease (PCVAD). The open reading frame 5 (ORF5) protein is a newly discovered non-structural protein in PCV2, which the function in viral pathogenesis remains unknown. The aim of this study was to investigate the mechanism of PCV2 ORF5 protein on autophagy and viral replication. The pEGFP-tagged ORF5 gene was ectopic expressed in PK-15 cells and an ORF5-deficient PCV2 mutant strain (PCV2ΔORF5) were used to infected PK-15 cells. This study demonstrated that the ORF5 is essential for the of PCV2-induced autophagy. The ORF5 protein triggers the phosphorylation of PERK, eIF2α and the expression of downstream transcription factor ATF4. In addition, ORF5 protein activated the AMPK-ERK1/2-mTOR signaling pathways. These findings suggest that ORF5 play essential roles in the induction of autophagy by PCV2. We further revealed that PCV2 ORF5 promotes viral replication through PERK-eIF2α-ATF4 and AMPK-ERK1/2-mTOR pathways. In conclusion, we showed that PCV2 ORF5 induces autophagy to promote virus replication in PK-15 cells.

Porcine circovirus type 2 exploits JNK-mediated disruption of tight junctions to facilitate Streptococcus suis translocation across the tracheal epithelium

Porcine circovirus type 2 (PCV2) is considered as the primary pathogen of porcine circovirus-associated disease (PCVAD), which results in significant economic losses worldwide. Clinically, PCV2 often causes disease through coinfection with other bacterial pathogens, including Streptococcus suis (S. suis), and especially the highly prevalent S. suis serotype 2 (SS2). The present study determined that continuous PCV2 infection in piglets down-regulates tight junction proteins (TJ) ZO-1 and occludin in the lungs. Swine tracheal epithelial cells (STEC) were used to explore the mechanisms and consequences of disruption of TJ, and an in vitro tracheal epithelial barrier model was established. Our results show that PCV2 infection in STEC decreases the expression levels of ZO-1 and occludin and increases the permeability of the tracheal epithelial barrier, resulting in easier translocation of SS2. Moreover, Western blot analysis indicates that PCV2 infection activates the JNK/MAPK pathway. The disruption of TJ in SETC and increased permeability of the epithelial barrier induced by PCV2 could be alleviated by inhibition of JNK phosphorylation, which indicates that the JNK/MAPK pathway regulates the expression of ZO-1 and occludin during PCV2 infection. This study allows us to better understand the mechanisms of PCV2 coinfection with bacterial pathogens and provides new insight into controlling the occurrence of PCVAD.

Full-genome sequences of porcine circovirus 3 (PCV3) and high prevalence in mummified fetuses from commercial farms in Brazil

The clinical implications of recently discovered porcine circovirus 3 (PCV3) infections are still unknown. The potential role of this emerging virus in reproductive loss in swine has been described. Herein, we report a high prevalence of PCV3 in mummified fetuses from sows maintained in modern farms in Rio Grande do Sul, Santa Catarina, Paraná, Goiás, and Mato Grosso do Sul states, Brazil. For this analysis, 276 mummified fetuses from 11 commercial swine farms were included. The presence of PCV3 DNA was confirmed using PCR, and the complete sequence of five different viral strains was obtained. Sequences of PCV3 genomes available on GenBank were then used for phylogenetic tree construction. Of the 276 mummified fetuses examined, 270 (nearly 97%) were positive for PCV3. In 93.1% of the fetuses, co-infections with at least one of the following agents were identified: porcine parvovirus (PPV), porcine circovirus 2 (PCV2) and Leptospira spp. Twelve fetuses were positive for PCV3 alone. The amino acid sequence of the capsid gene for the five viral strains shared 98–100% homology among them. Analysis of the DNA sequence indicates that the viruses identified in this study belong to the PCV3a1 subgroup. In summary, PCV3 DNA was detected in mummified fetuses at a surprisingly high rate. The role of PCV3 in porcine circovirus-associated disease (PCVAD) is still uncertain. However, considering that PCV3 has been detected in a variety of conditions, even in healthy animals, the present results confirm the need to investigate PCV3 as a causative agent of fetal mummification in swine.

Phylogenetic analysis of porcine circovirus type 2 (PCV2) between 2015 and 2018 in Henan Province, China

BackgroundPorcine circovirus type 2 (PCV2) is the pathogen of porcine circovirus associated diseases (PCVAD) and one of the main pathogens in the global pig industry, which has brought huge economic losses to the pig industry. In recent years, there has been limited research on the prevalence of PCV2 in Henan Province. This study investigated the genotype and evolution of PCV2 in this area.ResultsWe collected 117 clinical samples from different regions of Henan Province from 2015 to 2018. Here, we found that the PCV2 infection rate of PCV2 was 62.4%. Thirty-seven positive clinical samples were selected to amplify the complete genome of PCV2 and were sequenced. Based on the phylogenetic analysis of PCV2 ORF2 and complete genome, it was found that the 37 newly detected strains belonged to PCV2a (3 of 37), PCV2b (21 of 37) and PCV2d (13 of 37), indicating the predominant prevalence of PCV2b and PCV2d strains. In addition, we compared the amino acid sequences and found several amino acid mutation sites among different genotypes. Furthermore, the results of selective pressure analysis showed that there were 5 positive selection sites.ConclusionsThis study indicated the genetic diversity, molecular epidemiology and evolution of PCV2 genotypes in Henan Province during 2015–2018.

МОНИТОРИНГ ЦИРКОВИРУСНЫХ БОЛЕЗНЕЙ СВИНЕЙ В УСЛОВИЯХ ПРОМЫШЛЕННЫХ СВИНОКОМПЛЕКСОВ

Porcine circovirus associated diseases (PCVAD) - one of the most economically significant diseases of swine. No one of the PCVAD has any specific clinical symptoms. Causative agent is a porcine circovirus type 2 which belongs to Circovirus genus Circoviridae family. Nowadays PCV-2 is ubiquitous. Vaccination against PCVAD is widely using in most of swine farms. Vaccine application against PCVAD does not elicit development of sterile immunity. These fact is making conduction od diagnostic investigations with using of materials from animals of different ages/technological groups are necessary. The article is describing results of investigation about PCVAD diagnosis with applying of longitudinal method of monitoring. Application of the ELISA test-kits for detection PCV- 2 specific immunoglobulins IgG/IgM and polymerase chain reaction for the DNA of PCV-2 detection in blood serum allowed to set significant differences in serology and virology profile in examined farms. It was founded that viremia level in different technological groups is ranged from 0 to 40%; seroprevalence level - from 0 to 100 and from 10 to 50% for specific IgG and IgM, respectively. Application of ELISA test-kit for detection of immunoglobulins IgM to PCV-2 allows to detect a contact animals with PCV-2 (or vaccination) earlier than in case with immunoglobulins IgG. The obtained data reveals possibility of application scientific-based approach in detection of major pathogen in case of systemic disease development such as porcine respiratory syndrome complex (PRDC). In addition, monitoring investigations might be helpful in case of study of a control measures and specific prevention of PCVAD.

Generation and Immunogenicity of a Recombinant Pseudorabies Virus Co-Expressing Classical Swine Fever Virus E2 Protein and Porcine Circovirus Type 2 Capsid Protein Based on Fosmid Library Platform.

Pseudorabies (PR), classical swine fever (CSF), and porcine circovirus type 2 (PCV2)-associated disease (PCVAD) are economically important infectious diseases of pigs. Co-infections of these diseases often occur in the field, posing significant threat to the swine industry worldwide. gE/gI/TK-gene-deleted vaccines are safe and capable of providing full protection against PR. Classical swine fever virus (CSFV) E2 glycoprotein is mainly used in the development of CSF vaccines. PCV2 capsid (Cap) protein is the major antigen targeted for developing PCV2 subunit vaccines. Multivalent vaccines, and especially virus-vectored vaccines expressing foreign proteins, are attractive strategies to fight co-infections for various swine diseases. The gene-deleted pseudorabies virus (PRV) can be used to develop promising and economical multivalent live virus-vectored vaccines. Herein, we constructed a gE/gI/TK-gene-deleted PRV co-expressing E2 of CSFV and Cap of PCV2 by fosmid library platform established for PRV, and the expression of E2 and Cap proteins was confirmed using immunofluorescence assay and western blotting. The recombinant virus propagated in porcine kidney 15 (PK-15) cells for 20 passages was genetically stable. The evaluation results in rabbits and pigs demonstrate that rPRVTJ-delgE/gI/TK-E2-Cap elicited detectable anti-PRV antibodies, but not anti-PCV2 or anti-CSFV antibodies. These findings provide insights that rPRVTJ-delgE/gI/TK-E2-Cap needs to be optimally engineered as a promising trivalent vaccine candidate against PRV, PCV2 and CSFV co-infections in future.

PCV2 Regulates Cellular Inflammatory Responses through Dysregulating Cellular miRNA-mRNA Networks.

Porcine circovirus type 2 (PCV2) is closely linked to postweaning multisystemic wasting syndrome (PMWS) and other PCV-associated diseases (PCVADs), which influence the global pig industry. MicroRNAs (miRNAs) are evolutionarily conserved classes of endogenous small non-coding RNA that regulate almost every cellular process. According to our previous transcription study, PCV2 infection causes up-regulation of genes related to inflammation. To reveal the function of miRNAs in PCV2 infection and PCV2-encoded miRNAs, next generation sequencing and data analysis was performed to explore miRNA expression in PCV2-infected cells and non-infected cells. Data analysis found some small RNAs matched the PCV2 genome but PCV2 does not express miRNAs in an in vitro infection (PK-15 cells). More than 297 known and 427 novel miRNAs were identified, of which 44 miRNAs were differently expressed (DE). The pathways of inflammation mediated by chemokine and cytokine signaling pathway (P00031), were more perturbed in PCV2-infected cells than in mock controls. DE miRNAs and DE mRNA interaction network clearly revealed that PCV2 regulates the cellular inflammatory response through dysregulating the cellular miRNA-mRNA network. MiRNA overexpression and down-expression results demonstrated that miRNA dysregulation could affect PCV2 infection-induced cellular inflammatory responses. Our study revealed that host miRNA can be dysregulated by PCV2 infection and play an important role in PCV2-modulated inflammation.

Early antibody responses map to non-protective, PCV2 capsid protein epitopes

Porcine circovirus type 2 (PCV2) is an economically important cause of post-weaning multisystemic wasting syndrome (PMWS) in weanling piglets. Current commercial vaccines against PCV2 are highly effective. Yet, a recurring emergence of new genotypes in vaccinated herds necessitates a better understanding of protective immunity. The study objectives were to identify previously unrecognized decoy epitopes in the PCV2 capsid and test the hypothesis that early antibody responses would map to decoy epitopes and vice versa. Using a peptide library spanning the PCV2a capsid and weekly sera collections from PCV2a infected animals, three major immunodominant regions mapping the early responses to decoy epitopes were identified. Regions with potential decoy activity were mapped using peptide blocking fluorescent focus inhibition assays to residues 55 YTVKATTVRTPSWAVDMM 72, 106 WPCSPITQGDRGVGSTAV 123 and 124 ILDDNFVTKATALTYDPY 141. Post-vaccination responses largely recognized these same three identified regions and dominated the antibody responses to PCV2 in both infection and vaccination.

Development of a Combined Genetic Engineering Vaccine for Porcine Circovirus Type 2 and Mycoplasma Hyopneumoniae by a Baculovirus Expression System.

Mycoplasma hyopneumoniae (Mhp) and porcine circovirus type 2 (PCV2) are the main pathogens for mycoplasmal pneumonia of swine (MPS) and post-weaning multisystemic wasting syndrome (PMWS), respectively. Infection by these pathogens often happens together and causes great economic losses. In this study, a kind of recombinant baculovirus that can display P97R1P46P42 chimeric protein of Mhp and the capsid (Cap) protein of PCV2 was developed, and the protein location was identified. Another recombinant baculovirus was constructed without tag proteins (EGFP, mCherry) and was used to evaluate the immune effect in experiments with BALB/c mice and domestic piglets. Antigen proteins P97R1P46P42 and Cap were expressed successfully; both were anchored on the plasma membrane of cells and the viral envelope. It should be emphasized that in piglet immunization, the recombinant baculovirus vaccine achieved similar immunological effects as the mixed commercial vaccine. Both the piglet and mouse experiments showed that the recombinant baculovirus was able to induce humoral and cellular responses effectively. The results of this study indicate that this recombinant baculovirus is a potential candidate for the further development of more effective combined genetic engineering vaccines against MPS and PMWS. This experiment also provides ideas for vaccine development for other concomitant diseases using the baculovirus expression system.

The impact of porcine circovirus associated diseases on live attenuated classical swine fever vaccine in field farm applications

Porcine circovirus associated diseases (PCVADs) are among the most important diseases affecting the worldwide swine industry. Vaccination against porcine circovirus type 2 (PCV2) infection has been utilized for disease control and effectively reduces clinical signs of PCVADs. To evaluate the efficacy of the PCV2 vaccine in field farms, we conducted a trial using conventional pigs immunized with the subunit PCV2 vaccine followed by PCV2 challenge. Immunized pigs demonstrated lower serum viral loads, less viral antigen staining in lymph nodes, and higher average daily weight gain, confirming the protective efficacy of the vaccine. However, low levels of PCV2 infection were still detected in vaccinated pigs after challenge, suggesting that the PCV2 vaccine was unable to eradicate the virus, which could lead to asymptomatic PCV2 subclinical infection (PCV2-SI) in pig farms. Additionally, PCV2 infection is a risk factor for impaired pig immune response development during the weaning to growth stages, which is a crucial period to receive vaccines against classical swine fever (CSF). Therefore, the impact of PCV2-SI or PCV2-systemic disease (PCV2-SD) on live attenuated CSF vaccine was investigated. After PCV2 challenge, there was no difference in levels of classical swine fever virus (CSFV) neutralizing antibodies (NA) between pigs with PCV2-SD and PCV2-SI, suggesting that the efficacy of CSF vaccine was compromised. Moreover, results of long-term monitoring of CSFV NA titers in PCV2-SI pigs with minimized interference by maternally-derived antibodies suggested that serum PCV2 viral loads greater than 102 copies/mL may compromise the efficacy of CSF vaccine. Overall, a conventional pig model was established to demonstrate the impaired efficacy of the subunit PCV2 vaccine and its impact on the CSF vaccine in vaccination-challenge trials. Additionally, the impaired efficacy of the PCV2 vaccine resulted in increased PCV2-SI, eventually leading to compromised the live attenuated CSF vaccine induced NA response in field farm applications.

Transmission of porcine circovirus genotype type 2 (PCV2) in Russia and genotype association (PCV2d) with porcine dermatitis and nephropathy syndrome (PDNS)

The study of the prevalence of PCV2 genotypes in pig farms of the Russian Federation is presented. It is shown that currently the genotype PCV2d is the most common, which is important in connection with the assumed probability of lack of the immune response to PCV-2d induced by PCV-2a-based vaccines. The main goal of this investigation was to conduct monitoring studies for DNA, immunoglobulins IgG and IgM classes to PCV2 by the polymerase chain reaction (PCR) and enzyme-linked immunosorbent assay (ELISA) assays, respectively, as well as identification of the PCV2 genotype by sequencing of pigs of different ages were carried out at one of the Russian industrial pig farms with PCVAD (Porcine circovirus associated disease) outbreaks that used PCV2a vaccination. The data of this field observation indicates that PCV2d – is a major genotype in Russia. Additionally, it was shown that PCV2d prevalence in the farm correlates with clinical manifestation of PCVAD.