Postweaning Multisystemic Wasting Syndrome Research Articles

A prospective CSFV-PCV2 bivalent vaccine effectively protects against classical swine fever virus and porcine circovirus type 2 dual challenge and prevents horizontal transmission

Classical swine fever virus (CSFV) infection leading to CSF outbreaks is among the most devastating swine diseases in the pig industry. Porcine circovirus type 2 (PCV2) infection, resulting in porcine circovirus-associated disease (PCVAD), is also a highly contagious disease affecting pig health worldwide. To prevent and control disease occurrence, multiple-vaccine immunization is necessary in contaminated areas or countries. In this study, a novel CSFV-PCV2 bivalent vaccine was constructed and demonstrated to be capable of eliciting humoral and cellular immune responses against CSFV and PCV2, respectively. Moreover, a CSFV-PCV2 dual-challenge trial was conducted on specific-pathogen-free (SPF) pigs to evaluate vaccine efficacy. All of the vaccinated pigs survived and showed no clinical signs of infection throughout the experimental period. In contrast, placebo-vaccinated pigs exhibited severe clinical signs of infection and steeply increased viremia levels of CSFV and PCV2 after virus challenge. Additionally, neither clinical signs nor viral detections were noted in the sentinel pigs when cohabitated with vaccinated-challenged pigs at three days post-inoculation of CSFV, indicating that the CSFV-PCV2 bivalent vaccine completely prevents horizontal transmission of CSFV. Furthermore, conventional pigs were utilized to evaluate the application of the CSFV-PCV2 bivalent vaccine in field farms. An adequate CSFV antibody response and a significant decrease in PCV2 viral load in the peripheral lymph nodes were observed in immunized conventional pigs, suggesting its potential for clinical application. Overall, this study demonstrated that the CSFV-PCV2 bivalent vaccine effectively elicited protective immune responses and the ability to prevent horizontal transmission, which could be a prospective strategy for controlling both CSF and PCVAD in commercial herds.

Interaction between host cell proteins and open reading frames of porcine circovirus type 2.

Postweaning multisystemic wasting syndrome (PMWS) is caused by a systemic inflammation after porcine circovirus type 2 (PCV2) infection. It was one of the most economically important pathogens affecting pig production worldwide before PCV2 vaccine was first introduced in 2006. After the development of a vaccine against PCV2a type, pig farms gradually restored enormous economic losses from PMWS. However, vaccine against PCV2a type could not be fully effective against several different PCV2 genotypes (PCV2b - PCV2h). In addition, PCV2a vaccine itself could generate antigenic drift of PCV2 capsid. Therefore, PCV2 infection still threats pig industry worldwide. PCV2 infection was initially found in local tissues including reproductive, respiratory, and digestive tracks. However, PCV2 infection often leads to a systemic inflammation which can cause severe immunosuppression by depleting peripheral lymphocytes in secondary lymphoid tissues. Subsequently, a secondary infection with other microorganisms can cause PMWS. Eleven putative open reading frames (ORFs) have been predicted to encode PCV2 genome. Among them, gene products of six ORFs from ORF1 to ORF6 have been identified and characterized to estimate its functional role during PCV2 infection. Acquiring knowledge about the specific interaction between each PCV2 ORF protein and host protein might be a key to develop preventive or therapeutic tools to control PCV2 infection. In this article, we reviewed current understanding of how each ORF of PCV2 manipulates host cell signaling related to immune suppression caused by PCV2.

Immunogenicity Analysis of PCV3 Recombinant Capsid Protein Virus-like Particles and Their Application in Antibodies Detection.

Porcine circovirus type 3 is a newly emerging pathogen of porcine circovirus associated disease (PCVAD). Currently, there is no commercially available vaccine, resulting in huge economic losses to the pig industry. Porcine circovirus type 3 capsid protein (Cap) can self-assemble into virus-like particles (VLPs). Therefore, the expression of the recombinant Cap protein is of great significance for the prevention, diagnosis and control of porcine circovirus type 3 associated diseases. In this study, the recombinant Cap protein was successfully expressed in Escherichia coli by deleting the nuclear localization sequence (NLS). The VLPs were observed by transmission electron microscopy. To evaluate the immunogenicity of the recombinant Cap protein, mice were immunized. As a result, the recombinant Cap protein can induce higher levels of humoral and cellular immune responses. A VLP-based ELISA method was developed for the detection of antibodies. The established ELISA method has good sensitivity, specificity, repeatability and clinical applicability. These results demonstrate the successful expression of the PCV3 recombinant Cap protein and the preparation of recombinant Cap protein VLPs, which can be used for the preparation of subunit vaccines. Meanwhile, the established I-ELISA method lays a foundation for the development of the commercial PCV3 serological antibody detection kit.

Crystal structure of the dimerized of porcine circovirus type II replication-related protein Rep'.

Porcine circovirus type 2 (PCV2) can cause porcine circovirus-associated disease (PCVAD), which causes significant economic losses to the global pig industry annually. There are no effective antiviral drugs used to control and treat PCV2, and prevention is mainly obtained through vaccination. PCV2 genome replicates through the rolling circle replication (RCR) mechanism involving Rep and Rep', so analyzing the holistic structure of Rep and Rep' will help us better understand the replication process of PCV2. However, there are no reports on the integral structure of Rep' and Rep, which seriously hinders the research of the viral replication. By using the x-ray diffraction method, the structure of the Rep' dimer was resolved by us in this study. Structural analysis revealed that Rep' is a dimer formed by the interaction of the C-terminal domain. The two Rep' form a positively charged groove, which may play an essential role in the viral binding of dsDNA. Together, this study help to understand the replication process of the virus and may also provide new insights into the development of antiviral drugs.

Pathogenic ecological characteristics of PCV2 in large-scale pig farms in China affected by African swine fever in the surroundings from 2018 to 2021.

Porcine circovirus type 2 (PCV2) has been identified as the causal agent of postweaning multisystemic wasting syndrome (PMWS), an economically important multifactorial disease of the swine industry worldwide. This research designed a dual nested polymerase chain reaction (PCR) detection method to simultaneously monitor porcine circovirus type 2 (PCV2) and PCV3. The limit of detection (LoD) of sensitivity for PCV2 and PCV3 was ten copies/mL for both viruses. There was no cross-reaction with any other porcine pathogens tested and no non-specific amplification. The coincidence and repetition rates were both 100%. Through the systematic and clinical sampling, 15,130 samples collected from 30 large-scale pig farms in eight provinces in China (including Hubei, Hunan, Henan, Jiangxi, Shanxi, Guangdong, Hainan, and Heilongjiang) were subjected to early warning surveillance and/or clinical diagnosis. These results revealed that the overall positive rates of PCV3 and PCV2 were 0 and 28.29%, respectively, with the lowest level recorded in Jiangxi province. The highest carrying rate was observed in Hainan province. Pigs at different ages displayed varying carrying rates for PCV: fattening pigs and gilts had the highest and the lowest carrying rates for PCV, respectively. In addition, the excretion rates for PCV of colostrum, semen, and nasal, anal, and vulval swabs were tested. The colostrum, anal swabs, and semen had higher excretion rates for PCV; these were followed by the vulval and nasal swabs that had excretion rates for PCV. Furthermore, a high blood virus-carrying rate was detected in moribund pigs, especially in pigs with fever and red skin. As to the virus-carrying rate in the pig organs received from clinical necropsy, the highest rate was found in placental tissue, followed by the kidneys, and the virus also was detected in lymphoid organs, liver, stomach, and intestines. The PCV2-positive samples were sequenced to reveal the molecular epidemic dynamics of PCV2. The results indicated four major branches, namely, PCV2a, PCV2b, PCV2c, and PCV2d, concerning PCV2 molecular epidemiology in China, with PCV2a, PCV2b, and PCV2d dominating. In conclusion, the results obtained in this study elucidated the molecular epidemiology, transmission, and positive blood samples of PCV and provided new ideas for developing comprehensive PCV control technologies to begin eliminating the disease caused by PCV by cleaning pig farms.

Visual and label-free ASFV and PCV2 detection by CRISPR-Cas12a combined with G-quadruplex.

African swine fever (ASF) and postweaning multisystemic wasting syndrome (PMWS) are acute infectious diseases caused by the African swine fever virus (ASFV) and porcine circovirus type 2 (PCV2). At present, there are no effective vaccines for the prevention of ASFV. PMWS, which is harmful to the domestic and even the world pig industry, is difficult to cure and has a high mortality. So, developing simple, inexpensive, and accurate analytical methods to detect and effectively diagnose ASFV and PCV2 can be conducive to avoid ASFV and PCV2 infection. CRISPR has become a potentially rapid diagnostic tool due to recent discoveries of the trans-cleavage properties of CRISPR type V effectors. Herein, we report the visual detection based on CRISPR-Cas12a (cpf1), which is more convenient than fluorescence detection. Through in vitro cleavage target DNA activation, Cas12a can trans-cleavage ssDNA G-quadruplex. TMB/H2O2 and Hemin cannot be catalyzed by cleavaged G-DNA to produce green color products. This protocol is useful for the detection of ASFV and PCV2 with high sensitivity. This method can enable the development of visual and label-free ASFV and PCV2 detection and can be carried out in the field without relying on instruments or power. This method can complete nucleic acid detection at 37 °C without using other instruments or energy. Our research has expanded the application of Cas12a and laid the foundation for the field's rapid detection of viral nucleic acid in future.

Immunogenicity and Immunoprotection of PCV2 Virus-like Particles Incorporating Dominant T and B Cell Antigenic Epitopes Paired with CD154 Molecules in Piglets and Mice.

Porcine circovirus type 2 (PCV2) is capable of causing porcine circovirus-associated disease (PCVAD) and is one of the major threats to the global pig industry. The nucleocapsid protein Cap encoded by the PCV2 ORF2 gene is an ideal antigen for the development of PCV2 subunit vaccines, and its N-terminal nuclear localization sequence (NLS) structural domain is essential for the formation of self-assembling VLPs. In the present study, we systematically expressed and characterized full-length PCV2 Cap proteins fused to dominant T and B cell antigenic epitopes and porcine-derived CD154 molecules using baculovirus and found that the Cap proteins fusing epitopes were still capable of forming virus-like particles (VLPs). Both piglet and mice experiments showed that the Cap proteins fusing epitopes or paired with the molecular adjuvant CD154 were able to induce higher levels of humoral and cellular responses, particularly the secretion of PCV2-specific IFN-γ and IL-4. In addition, vaccination significantly reduced clinical signs and the viral load of PCV2 in the blood and tissues of challenged piglets. The results of the study provide new ideas for the development of a more efficient, safe and broad-spectrum next-generation PCV2 subunit vaccine.

LAMP Coupled CRISPR-Cas12a Module for Rapid, Sensitive and Visual Detection of Porcine Circovirus 2.

Simple SummaryIn this study, we aimed to establish a visual, rapid, low-cost, sensitive, specific, and portable nucleic acid detection method for PCV2 through coupling LAMP with CRISPR/Cas12a. All the results of LAMP-CRISPR detection, including a low detectable limit of 1 copy/μL, no cross-reaction with main porcine DNA or RNA viruses, and a 100% coincidence rate with qPCR detection, demonstrated that this method was reliable. It has laid the foundation for developing a PCV2 detection kit based on this LAMP-CRISPR method.Porcine circovirus 2 (PCV2) is the main pathogen of porcine circovirus-associated disease (PCVAD), which can cause considerable economic loss to the pig industry. The diagnosis of PCVAD is complicated and requires a series of clinical, pathological, and virological methods. Therefore, a rapid, highly sensitive, on-site, and visual diagnostic approach would facilitate dealing with the spread of PCV2. In this study, we intended to establish a new and effective PCV2 detection method through combining the no specific equipment requirement advantage of loop-mediated isothermal amplification (LAMP) with the property of clustered regular interspaced short palindromic repeats (CRISPR)/Cas12a system possessing the huLbCas12a collateral cleavage activity able to cleave single-stranded DNA fluorophore quencher probe sensor (designed as LAPM-CRISPR). Following a series of optimizations of its reaction conditions, this LAMP-CRISPR-based PCV2 detection could be conducted in constant temperature equipment, with the result reflected in a direct visual readout way. This established PCV2 detection approach presented fine sensitivity, rapidity, specificity, and reliability, as demonstrated by a low detectable limit of 1 copy/μL, completed within an hour, no cross-reaction with main porcine DNA or RNA viruses like PCV1, PCV3, and PEDV, and a 100% coincidence rate with that of the quantitative PCR (qPCR) method in the evaluation of 30 clinical blood samples, respectively. Therefore, this novel method makes rapid, on-site, visual, highly sensitive, and specific detection of PCV2 possible, facilitating the prevention of this pathogen in the field.

Porcine Circovirus Type 2 Vaccines: Commercial Application and Research Advances.

Porcine circovirus type 2 (PCV2) infection can lead to porcine circovirus-associated disease (PCVAD), causing great economic losses to the global swine industry. Conventional vaccination programs are a major measure in the prevention and control of this disease. Currently, there are 5 commercially available PCV2 vaccines in the international market and 10 kinds commercially available PCV2 vaccines in the Chinese market that confer good efficacy against this virus by alleviating clinicopathological manifestations and enhancing growth performance in pigs. In addition, diverse experimental PCV2 vaccines with protective efficiency have been developed, including attenuated chimeric, nucleic acid, subunit, multivalent, and viral-vectored vaccines. These experimental vaccines have been shown to be relatively effective in improving the efficiency of pig production and simplifying prevention procedures. Adjuvants can be used to promote vaccines with higher protective immunity. Herein, we review the application of multiple commercial vaccines over the years and research advances in experimental vaccines, which provide the possibility for the development of superior vaccines to successfully prevent and control PCV2 infection in the future.

Pathological Evaluation of Porcine Circovirus 2d (PCV2d) Strain and Comparative Evaluation of PCV2d and PCV2b Inactivated Vaccines against PCV2d Infection in a Specific Pathogen-Free (SPF) Yucatan Miniature Pig Model.

Porcine circovirus type 2 (PCV2) is an economically important swine pathogen that causes porcine circovirus-associated diseases (PCVADs). The objective of this study was to evaluate the use of specific pathogen-free Yucatan miniature pigs (YMPs) as an experimental model for PCV2d challenge and vaccine assessment because PCV2-negative pigs are extremely rare in conventional swine herds in Korea. In the first experiment, every three pigs were subjected to PCV2d field isolate or mock challenge. During three weeks of experiments, the PCV2d infection group exhibited clinical outcomes of PCVAD with high viral loads, lymphoid depletion, and detection of PCV2d antigens in lymphoid organs by immunohistochemistry. In the second experiment, three groups of pigs were challenged with PCV2d after immunization for three weeks: a nonvaccinated group (three pigs), a PCV2b-Vac group vaccinated with a commercial PCV2b-based inactivated vaccine SuiShot® Circo-ONE (five pigs), and a PCV2d-Vac group vaccinated with an experimental PCV2d-based inactivated vaccine (five pigs). During the three weeks of the challenge period, nonvaccinated pigs showed similar clinical outcomes to those observed in the PCV2d infection group from the first experiment. In contrast, both the PCV2b and PCV2d vaccinations produced good levels of protection against PCV2d challenge, as evidenced by reduced viral loads, improved growth performance, high virus-neutralizing antibody titers, and less development of PCV2-associated pathological lesions. Taken together, these data suggest that YMPs could be an alternative model for PCV2 challenge experiments, and these animals displayed typical clinical and pathological features and characteristics of protective immunity induced by the vaccines that were consistent with those resulting from PCV2 infections in conventional pigs.

Pathomorphology, immunohistochemical, and molecular detection of an atypical porcine dermatitis and nephropathy syndrome (PDNS) due to PCV-2d-2 in naturally affected grower pigs of India

Porcine circovirus type 2 (PCV-2) is of great economic significance to porcine industry worldwide. PCV-2 variants and genotypes, alternating world over, are the etiological agent of several clinical syndromes such as porcine dermatitis and nephropathy syndrome (PDNS), post-weaning multi-systemic wasting syndrome (PMWS) and others in pigs. This study is reporting an atypical manifestation of PDNS in twelve grower pigs, 3- to - 4.5 months age and either sex, died of the disease, with predominant lesions of nephropathy and no obvious clinical lesions in skin. Necropsy examination of pigs showed lesions of petechial -to- ecchymotic hemorrhages in the kidneys and in the right auricular musculature of the hearts. Microscopic lesions in H & E sections of the kidneys showed acute glomerulonephritis, interstitial nephritis, and vasculitis, but the skin morphology and architecture remained unaltered in contrast to the pathognomonic lesions of PDNS described in the literature. Other syndromic associations of PDNS in these cases included-perimyocarditis, interstitial pneumonia, depleted lymphoid tissues, tonsillitis, enteritis, and meningo-encephalitis. The lesional sites in duplicate paraffin tissue sections of kidneys, heart, lungs, spleen, lymph nodes, intestine, and brain demonstrated PCV-2 antigen in the cytoplasm of cells as highlighted by the intense immunolabeling on IHC staining. The PCV-2 positive organs reconfirmed by PCR, targeting ORF2 gene, which yielded 481bp size of products. The sequencing results of 481bp products on phylogenetic analysis showed 94% similarity with that of PCV-2 sequences in the database that grouped into PCV2d-2 genotype. The present report confirms, probably for the first time, the atypical PDNS cases due to PCV2d-2 genotype in naturally affected grower pigs of India.

Detection of Porcine Circovirus 1/2/3 and Genetic Analysis of Porcine Circovirus 2 in Wild Boar from Jiangxi Province of China.

Simple SummarySince PCV2 is currently the primary viral agent in intensive pigs, the cost of immune suppression and preventative measures has resulted in severe economic losses. It has been claimed that PCVs infect wild pigs in a number of nations, including South Korea, Germany, and Italy. Jiangxi province in China is a heavily populated area with an extremely intensive pig production industry, but data on the transmission of PCVs in wild boar are poor. In order to ascertain the incidence of PCV1/2/3 and the genetic characteristics of PCV2 circulating in wild boar in Jiangxi province, an epidemiological investigation was conducted. We discovered a medium prevalence of PCVs, notably PCV2, which indicated that wild boar in Jiangxi province were exposed to PCVs. Our findings highlight the need for the pig sector to actively prevent the contact between livestock and wild boar, which have a high risk of PCVs infection.A number of disorders that harm pig production are linked to porcine circoviruses, including PCV2. PCV2 infection is a substantial contributor to porcine-circovirus-associated illnesses (PCAS) and the post-weaning multi-systemic wasting syndrome (PMWS), which have a significant negative economic impact on pig production. Additionally, PCV infection has been labeled as a global concern to cattle and wildlife. This study’s objectives were to examine the prevalence of PCV1/2/3 in Jiangxi Province, China, and to clarify the epidemiological significance of wild boar in PCV epidemiology. The 2020 hunting seasons resulted in the collection of 138 wild boar samples for PCV1/2/3 detection, which was followed by the genetic clarification of PCV2 strains. According to our data, 21.7% (30/138) of the population had PCV1 positivity, 22.5% (31/138) had PCV2 positivity, and 5.8% (8/138) had PCV3 positivity. Additionally, 10 out of 138 wild boar samples had PCV1 and PCV2 co-infections, while 5 out of 138 wild boar samples had PCV2 and PC3 co-infections. Nineteen full-length PCV2 genomes measuring 1767 nt were recovered from various animal tissues using conventional PCR. Eighteen out of nineteen PCV2 strains were identified as PCV2b by phylogenetic tree analysis, which was completed by the reference strain HLJ2015 obtained from domestic pigs in 2015. Additionally, one genotype of PCV2d JX11-2020 (MW889021) shared a sub-branch with the referenced strain TJ (AY181946), which was isolated in domestic pigs in 2002. This finding raises the possibility that domestic pigs could contract PCV2 strains from wild boar, posing a serious threat to the Jiangxi province of China’s pig production industry.

Prevalence and genetic diversity of porcine circovirus type 2 in northern Guangdong Province during 2016–2021

The emergence and widespread of porcine circovirus-associated diseases (PCVADs), mainly caused by porcine circovirus type 2 (PCV2), threatens the Chinese swine industry. In this study, to investigate the recent prevalence of PCV2 in northern Guangdong Province of China, 573 tissue samples from 132 pig farms were collected during 2016–2021 and analyzed via PCR. Overall, 51.38% (297/573, 95%CI 47.74–55.92) samples were tested PCV2 positive. The detection rate of PCV2 was significantly lower in samples collected before 2016-2018 than after the outbreak of African Swine Fever (2019-2021), being 59.85% (158/264, 95%CI 53.94–65.76) and 41.47% (141/340, 95%CI 36.43–46.71), respectively. On the other end, the genetic characteristics of 26 PCV2 strains were further analyzed. These PCV2 strains belonged to three genotypes, including PCV2a, PCV2b, and PCV2d. Specifically, the predominant genotype prevalent during two periods (2016–2018 and 2019–2021) wasPCV2b (81.82%, 9/11) and PCV2d (80.0%, 12/15), respectively. The results above illustrated the high prevalence and the genetic evolution feature of PCV2 in Guangdong Province in recent years.

PDZK1 upregulates nitric oxide production through the PI3K/ERK2 pathway to inhibit porcine circovirus type 2 replication

Porcine circovirus type 2 (PCV2) is the causative agent of porcine circovirus-associated disease. Changes in host cell gene expression are induced by PCV2 infection. Here, we showed that porcine PDZ Domain-Containing 1 (PDZK1) expression was enhanced during PCV2 infection and that overexpression of PDZK1 inhibited the expression of PCV2 Cap protein. PCV2 genomic DNA copy number and viral titers were decreased in PDZK1-overexpressing PK-15B6 cells. PDZK1 knockdown enhanced the replication of PCV2. Overexpression of PDZK1 activated the phosphoinositide 3-kinase (PI3K)/ERK2 signaling pathway to enhance nitric oxide (NO) levels, while PDZK1 knockdown had the opposite effects. A PI3K inhibitor (LY294002) and a NO synthase inhibitor (L-NAME hydrochloride) decreased the activity of PDZK1 in restricting PCV2 replication. ERK2 knockdown enhanced the proliferation of PCV2 by decreasing levels of NO. Levels of interleukin (IL)− 4 mRNA were reduced in PDZK1 knockdown and ERK2 knockdown PK-15B6 cells. Increased IL-4 mRNA levels were unable to decrease NO production in PDZK1-overexpressing cells. Thus, we conclude that PDZK1 affected PCV2 replication by regulating NO production via PI3K/ERK2 signaling. PDZK1 affected IL-4 expression through the PI3K/ERK2 pathway, but PDZK1 modulation of PCV2 replication occurred independently of IL-4. Our results contribute to understanding the biological functions of PDZK1 and provide a theoretical basis for the pathogenic mechanisms of PCV2.

Pathology of PMWS Associated with Classical Swine Fever Virus, Porcine Parvo Virus and Multidrug Resistant Enteric Escherichia coli Infections in an Organized Swine Farm, Rajasthan, India

Background: Postweaning multisystemic wasting syndrome (PMWS) is caused by porcine circovirus type 2 (PCV2). It is a globally emerging clinical condition which affects nursing piglets and growing pigs. PCV2 requires various cofactors for fully development of PMWS. The present report describes the pathology of PMWS associated with classical swine fever virus (CSFV), porcine parvo virus (PPV) and multidrug resistant enteric Escherichia coli infections in an organized swine farm, Rajasthan. In India, this is the first report on pathology of PMWS associated with all the three viruses and multidrug resistant enteric colibacillosis. Methods: Detailed necropsy was conducted on pig carcass submitted to Post-mortem Facility, Division of Pathology, ICAR-Indian Veterinary Research Institute (IVRI), Izatnagar in the year 2017 from an organized swine farm, Rajasthan, India. Heart blood and diarrhoeic intestinal contents were collected aseptically and stored separately at 4°C for bacteriological examination and antibiotic sensitivity test. Tissue samples were collected in sterile container and stored at -40°C for DNA and RNA extraction and polymerase chain reaction (PCR) and also collected in 10% neutral buffered saline for histopathological processing. Result: The organized farm was having history of 33% (200/600) of mortality among 2 months to 6 months old piglets. Gross lesions were wasting, lymphadenopathy, non-collapsed lungs, hydropericardium, hepatomegaly, icterus, enteritis and pale kidneys, button shaped ulcers in the caecum and colon etc. Histopathologically, lymphoid depletion, reticuloendothelial cell proliferation in lymphoid tissue, fibrin thrombi in spleen, severe hepatitis, interstitial pneumonia, interstitial nephritis, severe necrosis in large intestine, meningitis, perivascular cuffing (PVC), gliosis, neuronal degeneration in cerebrum etc were observed. Molecular diagnosis and bacteriological examination confirmed presence of PCV2, PPV, CSFV and multidrug resistant Escherichia coli.

Advances in Crosstalk between Porcine Circoviruses and Host.

Porcine circoviruses (PCVs), including PCV1 to PCV4, are non-enveloped DNA viruses with a diameter of about 20 nm, belonging to the genus Circovirus in the family Circoviridae. PCV2 is an important causative agent of porcine circovirus disease or porcine circovirus-associated disease (PCVD/PCVAD), which is highly prevalent in pigs and seriously affects the swine industry globally. Furthermore, PCV2 mainly causes subclinical symptoms and immunosuppression, and PCV3 and PCV4 were detected in healthy pigs, sick pigs, and other animals. Although the pathogenicity of PCV3 and PCV4 in the field is still controversial, the infection rates of PCV3 and PCV4 in pigs are increasing. Moreover, PCV3 and PCV4 rescued from infected clones were pathogenic in vivo. It is worth noting that the interaction between virus and host is crucial to the infection and pathogenicity of the virus. This review discusses the latest research progress on the molecular mechanism of PCVs–host interaction, which may provide a scientific basis for disease prevention and control.

Porcine circovirus-like virus P1 activates pancreatic secretion pathway by interacting with CHRM3 protein

The porcine circovirus-like virus P1, a member of the circovirus family, causes post-weaning multisystemic wasting syndrome (PMWS) in weaned piglets with progressive wasting as the main clinical symptom. The pancreatic secretion pathway induces pancreatic acinar cells to secrete various digestive enzymes and as such is an important signaling pathway for the digestive system and somatic growth. This study examined the effects and mechanism of P1 virus infection on the pancreatic secretion pathway. The experiment was conducted by transfecting double-copy plasmid P1 into PK-15 and 3D4 cells and by infecting cells with the P1 virus. Samples were collected at various times after transfection or infection. The pathway's transcription and translation levels of CHRM3, Gq, PLC-β2, PRKCA, Rab3D, RhoA, Rac1, and amyA proteins were detected by real-time PCR and Western blots; these analyses confirmed that the P1 virus infection could upregulate the expression level of key pancreatic secretion signaling molecules. Then, we confirmed that the VP1 protein of the P1 virus could interact with the pathway initiation protein CHRM3 using Co-IP, pull-downs, and confocal fluorescence microscopy. Finally, we demonstrated that the VP1 protein activates the pancreatic secretory pathway through the CHRM3 protein. In conclusion, this study demonstrated that the P1 virus can interact with the CHRM3 receptor protein to activate the pancreatic secretion pathway and promote the secretion of various digestive enzymes downstream of the pathway, thereby providing a basis for P1 virus pathogenesis.

Pathogenicity of Porcine Circovirus Type 2e in Experimentally Infected Pigs

A new porcine circovirus type 2e (PCV2e) genotype was recently isolated from diseased pigs. To investigate the pathogenicity of PCV2e, groups of conventional pigs were inoculated in one of three ways: with PCV2e only, with Mycoplasma hyopneumoniae and PCV2e, or with PCV2e and porcine reproductive and respiratory syndrome virus (PRRSV). Pigs were examined post mortem at 21 days post inoculation. Pigs in the M. hyopeumoniae/PCV2e group were inoculated intratracheally with M. hyopneumoniae at 4 weeks of age followed by an intranasal inoculation with PCV2e at 6 weeks of age. Pigs in the PCV2e/PRRSV group were inoculated intranasally with PCV2e and PRRSV at 6 weeks of age. Significant differences in PCV2e loads in blood or lymph nodes, or in the severity of lymphoid lesions, were not detected between the M. hyopneumoniae/PCV2e and PCV2e/PRRSV groups. All pigs co-infected with either M. hyopneumoniae/PCV2e or PCV2e/PRRSV developed mild porcine circovirus-associated disease (PCVAD), whereas none of the pigs infected with PCV2e alone developed PCVAD. Production of PCVAD in pigs therefore appears to require PCV2e infection simultaneously with an additional infectious agent such as M. hyopneumoniae or PRRSV for full disease expression in pigs. These results demonstrate that PCV2e is not associated with significant clinical disease as assessed by levels of PCV2e viraemia and severity of lymphoid lesions.

Intervening Effects and Molecular Mechanism of Quercitrin on PCV2-Induced Histone Acetylation, Oxidative Stress and Inflammatory Response in 3D4/2 Cells.

Porcine circovirus type 2 (PCV2) is the main pathogen causing porcine circovirus-associated diseases (PCVD/PCVADs), and infection of the host induces immunosuppression. Since quercitrin (QUE) has anti-inflammatory and antiviral activity, it is worth exploiting in animal diseases. In this study, the interventional effects and the molecular mechanism of QUE on PCV2-induced oxidative stress and inflammatory responses in 3D4/2 cells and the modulation of histone acetylation modifications were investigated. The ROS production was measured by DCFH-DA fluorescent probes. HAT and HDAC enzyme activity were determined by ELISA. Histone acetylation, oxidative stress and inflammation-related gene expression levels were measured by q-PCR. Histone H3 and H4 (AcH3 and AcH4) acetylation, oxidative stress and inflammation-related protein expression levels were measured by Western blot. The results showed that QUE treatment at different concentrations on PCV2-infected 3D4/2 cells was able to attenuate the production of ROS. Moreover, QUE treatment could also intervene in oxidative stress and decrease the enzyme activity of HAT and the mRNA expression level of HAT1, while it increased the enzyme activity of HDAC and HDAC1 mRNA expression levels and downregulated histone H3 and H4 (AcH3 and AcH4) acetylation modification levels. In addition, QUE treatment even downregulated the mRNA expression levels of IL-6, IL-8, IκB, AKT and p38, but upregulated the mRNA expression levels of IL-10, SOD, GPx1, p65, Keap1, Nrf2, HO-1 and NQO1. As to protein expression, QUE treatment downregulated the levels of iNOS, p-p65 and IL-8 as well as the phosphorylation expression of IκB and p38, while it upregulated the levels of HO-1 and NQO1. It was shown that QUE at 25, 50 or 100 μmol/L regulated p38MAPK and PI3K/AKT signaling pathways by downregulating cellular histone acetylation modification levels while inhibiting the NF-κB inflammatory signaling pathway and activating the Nrf2/HO-1 antioxidant signaling pathway, thus regulating the production of inflammatory and antioxidant factors and exerting both anti-inflammatory and antioxidant effects.

Genetic Diversity of Porcine Circovirus 2 in Wild Boar and Domestic Pigs in Ukraine.

Porcine circovirus type 2 (PCV2) is responsible for a number of porcine circovirus-associated diseases (PCVAD) that can severely impact domestic pig herds. For a non-enveloped virus with a small genome (1.7 kb ssDNA), PCV2 is remarkably diverse, with eight genotypes (a–h). New genotypes of PCV2 can spread through the migration of wild boar, which are thought to infect domestic pigs and spread further through the domestic pig trade. Despite a large swine population, the diversity of PCV2 genotypes in Ukraine has been under-sampled, with few PCV2 genome sequences reported in the past decade. To gain a deeper understanding of PCV2 genotype diversity in Ukraine, samples of blood serum were collected from wild boars (n = 107) that were hunted in Ukraine during the November–December 2012 hunting season. We found 34/107 (31.8%) prevalence of PCV2 by diagnostic PCR. For domestic pigs, liver samples (n = 16) were collected from a commercial market near Kharkiv in 2019, of which 6 out of 16 (37%) samples were positive for PCV2. We sequenced the genotyping locus ORF2, a gene encoding the PCV2 viral capsid (Cap), for 11 wild boar and six domestic pig samples in Ukraine using an Oxford Nanopore MinION device. Of 17 samples with resolved genotypes, the PCV2 genotype b was the most common in wild boar samples (10 out of 11, 91%), while the domestic pigs were infected with genotypes b and d. We also detected genotype b/d and b/a co-infections in wild boars and domestic pigs, respectively, and for the first time in Ukraine we detected genotype f in a wild boar from Poltava. Building a maximum-likelihood phylogeny, we identified a sublineage of PCV2 genotype b infections in both wild and domestic swine, suggesting a possible epizootic cluster and an ecological interaction between wild boar and domestic pig populations in northeastern Ukraine.