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Porcine circoviruses (PCVs) are widely distributed in swine herds. PCV2, the significant swine pathogen, causes infections characterized by growth and development disorders, skin lesions, and respiratory distress. PCV3 has been circulating worldwide and can be associated with various clinical signs and disease developments. Wild boars are the main reservoir of these pathogens in wildlife and can create an alarming threat to pig farming. In Russia, three PCV2 genotypes (PCV2a, PCV2b, and PCV2d) were identified in pig farms. Additionally, PCV3 was observed in pig herds during the monitoring studies in the country. However, data considering the circulation of PCVs in herds of wild boars in Russia is scant. For this purpose, we performed PCR assays of the samples from 30 wild boars hunted in the Moscow Region of Russia in 2021–2023. The ratios of wild boars positive for PCV2, PCV3, or coinfected were 50, 10, and 13.3%, respectively. Additionally, we sequenced 15 PCV2 and four PCV3 complete genomes and conducted phylogenetic analysis, which divided PCV2 isolates into two groups: PCV2d and PCV2b. The study showed a high infection rate of PCV2 among wild boars, with PCV2d dominance. Simultaneously, PCV3 also circulates among wild boars. The obtained results can provide a basis for the development of preventive measures to support infection transmission risks between farm and wild animals. [ABSTRACT FROM AUTHOR]

Porcine circovirus disease (PCV) causes substantial economic losses in the pig industry, primarily from porcine circovirus type 2 (PCV2) and porcine circovirus type 3 (PCV3). Novel vaccines are necessary to prevent and control PCV infections. PCV coat proteins are crucial for eliciting immunogenic proteins that induce the production of antibodies and immune responses. A vaccine platform utilizing Semliki Forest virus RNA replicons expressing vesicular stomatitis virus glycoprotein (VSV-G), was recently developed. This platform generates virus-like vesicles (VLVs) containing VSV-G exclusively, excluding other viral structural proteins. In our study, we developed a novel virus-like vesicle vaccine by constructing recombinant virus-like vesicles (rVLVs) that also express EGFP. These rVLVs were created using the RNA replicon of Venezuelan equine encephalomyelitis (VEEV) and New Jersey serotype VSV-G. The rVLVs underwent characterization and safety evaluation in vitro. Subsequently, rVLVs expressing PCV2d-Cap and PCV3-Cap proteins were constructed. Immunization of C57 mice with these rVLVs led to a significant increase in anti-porcine circovirus type 2 and type 3 capsid protein antibodies in mouse serum. Additionally, a cellular immune response was induced, as evidenced by high production of IFN-γ and IL-4 cytokines. Overall, this study demonstrates the feasibility of developing a novel porcine circovirus disease vaccine based on rVLVs. [ABSTRACT FROM AUTHOR]

Porcine circoviruses (PCVs) are widely distributed in swine herds. PCV2, the significant swine pathogen, causes infections characterized by growth and development disorders, skin lesions, and respiratory distress. PCV3 has been circulating worldwide and can be associated with various clinical signs and disease developments. Wild boars are the main reservoir of these pathogens in wildlife and can create an alarming threat to pig farming. In Russia, three PCV2 genotypes (PCV2a, PCV2b, and PCV2d) were identified in pig farms. Additionally, PCV3 was observed in pig herds during the monitoring studies in the country. However, data considering the circulation of PCVs in herds of wild boars in Russia is scant. For this purpose, we performed PCR assays of the samples from 30 wild boars hunted in the Moscow Region of Russia in 2021–2023. The ratios of wild boars positive for PCV2, PCV3, or coinfected were 50, 10, and 13.3%, respectively. Additionally, we sequenced 15 PCV2 and four PCV3 complete genomes and conducted phylogenetic analysis, which divided PCV2 isolates into two groups: PCV2d and PCV2b. The study showed a high infection rate of PCV2 among wild boars, with PCV2d dominance. Simultaneously, PCV3 also circulates among wild boars. The obtained results can provide a basis for the development of preventive measures to support infection transmission risks between farm and wild animals.

Preventive variables for childhood leukemia incidence (LI) remain unknown. Past assertions that childhood vaccinations, especially BCG, may be potentially protective have remained disputed for over five decades because of the lack of a unifying framework to explain variable outcomes in different studies. An examination of the early childhood LI for 2020 in European Region countries with supposedly similar underlying confounders but differential childhood vaccination coverage displays negative covariation with prevailing Mycobacterium spp. exposure in BCG-vaccinated children. The childhood LI in 0-4-year-old populations with >90% childhood BCG vaccination coverage is found to be strongly but negatively correlated with prevailing tuberculin immunoreactivity [r(24): -0.7868, p-value: < 0.0001]. No such correlation existed for the LI in 0-4-year-old populations without BCG vaccinations, though weak associations are hinted at by the available data for MCV2, PCV3, and DTP3 vaccinations. We hypothesize that early childhood BCG vaccination "priming" and subsequent "trained immunity" augmentation by "natural" boosting from Mycobacterium spp. exposure play a preventive and protective role in childhood LI. The non-consideration of prevailing "trained immunity" could have been a cause behind the conflicting outcomes in past studies. Exploratory studies, preferably performed in high-burden countries and controlling for the trained-immunity correlate and other potential confounders, would be warranted in order to establish a role for BCG vaccination and early-life immune training (or lack thereof) in childhood LI and help put the current controversy to rest. [ABSTRACT FROM AUTHOR]

This study aimed to evaluate the relationship between porcine circovirus type 3 (PCV3) viral load and histopathological findings in perinatal piglet tissues and to develop an immunohistochemical method for detecting the virus in lesions. The quantitative polymerase chain reaction (qPCR) cycle threshold (Ct) when amplifying PCV3 DNA and the area of perivascular inflammatory infiltrates in different organs [central nervous system (CNS), lung, heart, liver, spleen, and lymph nodes] were compared. To develop an immunohistochemistry technique, rabbit sera were produced against PCV3-capsid protein peptides selected using bioinformatic analyses. The assay was initially implemented using a tissue sample previously tested using qPCR and in situ hybridization to optimize the procedure and reagent dilutions. To evaluate immunohistochemistry performance, tissue samples from another 17 cases were analyzed using standardized parameters. The most common microscopic lesion was multisystemic periarteritis, with associated vasculitis, as the mesenteric vascular plexus is one of the most affected organs. Other tissues, such as the heart, lung, CNS, and skeletal muscle, were also affected. Comparison of the Ct values for different tissues showed no significant difference, except in lymphoid organs (spleen and lymph nodes), which had significantly higher viral loads than the CNS tissues. There was no correlation between Ct values and perivascular inflammatory infiltrates. PCV3 immunohistochemistry revealed granular immunolabeling, mainly in the cytoplasm of cells in the vascular mesenteric plexus, heart, lung, kidney, and spleen.

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 viruscarrying 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. [ABSTRACT FROM AUTHOR]

IntroductionPorcine circovirus type 2 (PCV2) is considered one of the viruses with substantial economic impact on swine industry in the word. Recently, porcine circovirus type 3 (PCV3) has been found to be associated with porcine dermatitis and nephropathy syndrome (PDNS)-like disease. And the two viruses were prone to co-infect clinically.MethodsTo further investigate the prevalence and genetic diversity of the two viruses, 257 pig samples from 23 different pig farms in southwest China with suspected PCVAD at different growth stages were analyzed by real-time PCR between 2020 and 2022 to determine the presence of PCV2 and PCV3.ResultsResults showed high prevalence of PCV2 and PCV3: 26.46% samples were PCV2 positive and 33.46% samples were PCV3 positive. The coinfection rate was doubled from 2020 (5.75%) to 2022 (10.45%). Subsequently, the whole genome sequences of 13 PCV2 and 18 PCV3 strains were obtained in this study. Of these, 1 strain was PCV2a, 5 strains were PCV2b and 7 strains were PCV2d, indicating that PCV2d was the predominant PCV2 genotype prevalent in the Southwest of China.DiscussionIn addition, the phylogenetic analysis of PCV3 showed high nucleotide homology (>98%) between the sequences obtained in this study and reference sequences. And 3 mutations (A24V, R27K and E128D) were found in PCV3 antibody recognition domains, which might be related to the mechanism of viral immune escape. Thus, this study will enhance our understanding of the molecular epidemiology and evolution of PCV2 and PCV3, which are conducive to the further study of the genotyping, immunogenicity and immune evasion of PCVs.

Porcine circovirus type 3 (PCV3), a novel circovirus, imposes great burdens on the global pig industry. The penside tests for detecting PCV3 are critical for assessing the epidemiological status and working out disease prevention and control programs due to the unavailability of a commercial vaccine. A one-step molecular assay based on visual loop-mediated isothermal amplification (vLAMP) was developed for simple and rapid detection of PCV3. We compared its sensitivity and specificity with TaqMan quantitative real-time polymerase chain reaction (qPCR) and applied the developed assay in the epidemiological study of (n = 407) pooled swine sera collected from almost the entire mainland China during the years 2017–2018. We also explored the feasibility of the vLAMP assay for detecting raw samples without a prior DNA isolation step to expand its application capability. Results showed that the vLAMP assay could reliably detect the PCV3 cap gene with a detection limit of 10 DNA copies equal to that of the Taqman qPCR assay. In the epidemiological study, the PCV3 positive detection rate for 407 swine pooled sera detected by the vLAMP assay was 37.35% (152/407), whereas it was 39.01% (159/407) for Taqman qPCR. For the detection method without genome extraction, the results kept satisfactory specificity (100%) but displayed lower sensitivity (100% for CT < 32), indicating the direct detection is not sensitive enough to discriminate the samples with low viral loads. The one-step vLAMP is a convenient, rapid, and cost-effective diagnostic for penside detection and will enable the epidemiological surveillance of PCV3, which has widely spread in mainland China. [ABSTRACT FROM AUTHOR]

Porcine circovirus type 3 (PCV3), a novel circovirus, imposes great burdens on the global pig industry. The penside tests for detecting PCV3 are critical for assessing the epidemiological status and working out disease prevention and control programs due to the unavailability of a commercial vaccine. A one-step molecular assay based on visual loop-mediated isothermal amplification (vLAMP) was developed for simple and rapid detection of PCV3. We compared its sensitivity and specificity with TaqMan quantitative real-time polymerase chain reaction (qPCR) and applied the developed assay in the epidemiological study of (n = 407) pooled swine sera collected from almost the entire mainland China during the years 2017–2018. We also explored the feasibility of the vLAMP assay for detecting raw samples without a prior DNA isolation step to expand its application capability. Results showed that the vLAMP assay could reliably detect the PCV3 cap gene with a detection limit of 10 DNA copies equal to that of the Taqman qPCR assay. In the epidemiological study, the PCV3 positive detection rate for 407 swine pooled sera detected by the vLAMP assay was 37.35% (152/407), whereas it was 39.01% (159/407) for Taqman qPCR. For the detection method without genome extraction, the results kept satisfactory specificity (100%) but displayed lower sensitivity (100% for CT < 32), indicating the direct detection is not sensitive enough to discriminate the samples with low viral loads. The one-step vLAMP is a convenient, rapid, and cost-effective diagnostic for penside detection and will enable the epidemiological surveillance of PCV3, which has widely spread in mainland China.

Porcine circovirus type 3 (PCV3), a virus belonging to the Circoviridae family, is considered to be associated with respiratory and neurological signs, cardiac and multisystemic inflammation, reproductive failure, and porcine dermatitis and nephropathy syndrome-like disease in pigs (Sus scrofa). In this study, epidemiological and serological investigations of PCV3 in clinically healthy pigs from different regions of China were performed. Overall, 42.87% (1,101/2,568) of pigs were positive for PCV3 Cap antibody via indirect enzyme-linked immunosorbent assay, with a higher prevalence of PCV3 in multiparous sows (62.22%, 881/1,416) and fattening pigs (28.96%, 159/549) than in suckling piglets (8.96%, 32/357) and nursery pigs (11.79%, 29/246). Of the 2,568 samples, 255 were further tested for PCV3 DNA using real-time polymerase chain reaction, and 63.14% of these were positive, with nearly half having

This document is an editorial published in the journal Frontiers in Veterinary Science. It discusses the emergence of swine infectious diseases and their impact on the global swine industry. The editorial highlights the economic losses and potential public health issues caused by diseases such as porcine reproductive and respiratory syndrome virus (PRRSV), porcine epidemic diarrhea virus (PEDV), and pseudorabies virus (PRV). It also mentions the identification of novel pathogens like Senecavirus A, atypical porcine pestivirus (APPV), porcine circovirus 3 (PCV3), porcine circovirus 4 (PCV4), porcine deltacoronavirus (PDCoV), swine acute diarrhea syndrome coronavirus (SADS-CoV), and influenza D virus (IDV). The editorial emphasizes the need for further research and attention to emerging swine infectious diseases. [Extracted from the article]

Since 2016, a novel porcine circovirus, PCV3, has been infecting pigs, causing significant economic losses to the pig industry. In recent years, the infection rate of PCV3 has been increasing, and thus rapid and accurate detection methods for PCV3 are essential. In this study, we established a novel probe-based single multiple cross displacement amplification (P-S-MCDA) method for PCV3. The method was termed as P-S-MCDA. The P-S-MCDA uses seven primers to amplify the capsid gene, and the assay can be performed at 60°C for 30 min, greatly shortening the reaction time. The results of P-S-MCDA can not only be monitored in real time through fluorescence signals but also be determined by observing the fluorescence of the reaction tubes using a smartphone-based cassette. This method demonstrated good specificity and the same sensitivity as qPCR, with a minimum detection limit of 10 copies. In 139 clinical samples, the coincidence rate with qPCR was 100%. The P-S-MCDA can be widely applied in PCV3 detection in laboratories or in rural areas.

Porcine circovirus type 3 (PCV3) invades multiple tissues and organs of pigs of different ages and are widely spread throughout pig farms, emerging as an important viral pathogen that can potentially damage the pig industry worldwide. Since PCV3 is a newly discovered virus, many aspects of its life cycle remain unknown. Porcine kidney epithelial cells are important host targets for PCV3. Here, we used systematic approaches to dissect the molecular mechanisms underlying the cell entry and intracellular trafficking of PCV3 in PK15 cells, a cell line of porcine kidney epithelial origin. A large number of PCV3 viral particles were found to colocalize with clathrin but not caveolin-1 after entry, and PCV3 infection was significantly decreased when treated with chlorpromazine, dynasore, knockdown of clathrin heavy chain expression via RNA interference, or overexpression of a dominant-negative mutant of EPS15 in PCV3-infected cells. After internalization, the viral particles were further observed to colocalize with Rab5 and Rab7, and knockdown of both expression by RNA interference significantly inhibited PCV3 replication. We also found that PCV3 infection was impeded by ammonium chloride treatment, which indicated the requirement of an acidic environment for viral entry. Taken together, our findings demonstrate that PCV3 enters PK15 cells through a clathrin- and dynamin-2-mediated endocytic pathway, which requires early and late endosomal trafficking, as well as an acidic environment, providing an insightful theoretical basis for further understanding the PCV3 life cycle and its pathogenesis.

Porcine circovirus 3 (PCV3) infections cause clinical diseases similar to those seen in porcine circovirus 2 (PCV2) infections. It is unclear whether PCV3 infections can also cause immunosuppression like that seen with PCV2. Here, we report that Cap inhibits DNA-induced IFN-β mRNA transcription and IFN promoter activation. Cap was also found to inhibit cyclic GMP-AMP (cGAMP) synthase (cGAS) binding to interferon-stimulating DNA (ISD). Immunoprecipitation and mass spectrometry were used to identify cellular interaction partners of Cap. Cap interacted with G3BP1 and inhibited the interaction between GTPase-activating protein-(SH3 domain)-binding protein 1 (G3BP1) and cGAS. Furthermore, the destruction of endogenously expressed G3BP1 by siRNA significantly reduced IFN promoter activation, and phosphorylation of tank-binding kinase 1 (TBK1) was induced by ISD. Overexpression of G3BP1 attenuated the inhibition of ISD binding of cGAS by Cap and promoted phosphorylation of TBK1 and IRF3 induced by ISD. Collectively, our results show that the interaction between Cap and G3BP1 prevents cGAS from recognizing DNA, thereby inhibiting the IFN production.

Porcine circovirus type 3 (PCV3) invades multiple tissues and organs of pigs of different ages and are widely spread throughout pig farms, emerging as an important viral pathogen that can potentially damage the pig industry worldwide. Since PCV3 is a newly discovered virus, many aspects of its life cycle remain unknown. Porcine kidney epithelial cells are important host targets for PCV3. Here, we used systematic approaches to dissect the molecular mechanisms underlying the cell entry and intracellular trafficking of PCV3 in PK15 cells, a cell line of porcine kidney epithelial origin. A large number of PCV3 viral particles were found to colocalize with clathrin but not caveolin-1 after entry, and PCV3 infection was significantly decreased when treated with chlorpromazine, dynasore, knockdown of clathrin heavy chain expression via RNA interference, or overexpression of a dominant-negative mutant of EPS15 in PCV3-infected cells. After internalization, the viral particles were further observed to colocalize with Rab5 and Rab7, and knockdown of both expression by RNA interference significantly inhibited PCV3 replication. We also found that PCV3 infection was impeded by ammonium chloride treatment, which indicated the requirement of an acidic environment for viral entry. Taken together, our findings demonstrate that PCV3 enters PK15 cells through a clathrin- and dynamin-2-mediated endocytic pathway, which requires early and late endosomal trafficking, as well as an acidic environment, providing an insightful theoretical basis for further understanding the PCV3 life cycle and its pathogenesis. [ABSTRACT FROM AUTHOR]

In 2016, a novel porcine circovirus (PCV), PCV3, was identified in USA. Subsequently, it was proved to be also epidemic in China, Poland, and Korea. To analyze and control the epidemic situation of PCV3, it is necessary to establish accurate and high-throughput detection methods. In this study, the colorimetric isothermal multiple-self-matching-initiated amplification (IMSA) using cresol red was developed to detect PCV3 for the first time. The reaction can be easily performed by incubating the tube at 63°C for 60 min. By the addition of pH-sensitive indicator dye cresol red, the initial color of the reaction mixture is red. When PCV3 capsid gene DNA was positive in the sample, the color of the reaction mixture changed from red to yellow after the isothermal incubation at 63°C, while the negative control maintained the red color. The colorimetric IMSA displayed good specificity in detecting PCV3, PCV2, and PCV1 and 4 porcine DNA pathogens. Moreover, it has a low and repeatable detection limit of 10 copies, which is consistent with TaqMan-based qPCR, but 10 times more sensitive than PCR. In diagnosing 128 clinical specimens, it not only showed 100% agreement with qPCR but also detected 15 positive results more than PCR. The colorimetric IMSA we offered might be a good choice for PCV3 epidemiological investigation and point-of-care testing.

Next generation sequencing (NGS) can be applied to identify and characterize the entire set of microbes within a sample. However, this platform does not provide a morphological context or specific association between the viral or bacterial sequences detected and the histological lesions. This limitation has generated uncertainty whether the sequences identified by NGS are actually contributing or not for the clinical outcome. Although in situ hybridization (ISH) and immunohistochemistry (IHC) can be used to detect pathogens in tissue samples, only ISH has the advantage of being rapidly developed in a context of an emerging disease, especially because it does not require development of specific primary antibodies against the target pathogen. Based on the sequence information provided by NGS, ISH is able to check the presence of a certain pathogen within histological lesions, by targeting its specific messenger RNA, helping to build the relationship between the pathogen and the clinical outcome. In this mini review we have compiled results of the application of NGS-ISH to the investigation of challenging diagnostic cases or emerging pathogens in pigs, that resulted in the detection of porcine circovirus type 3, porcine parvovirus type 2, Senecavirus A, and Mycoplasma hyorhinis.

Porcine circovirus (PCV) is one of the smallest known DNA viruses in mammals. At present, PCVs are divided into three species, PCV1, PCV2, and PCV3. PCV1 and PCV2 were found in the 1970s and the 1990s, respectively, whereas PCV3 was discovered recently in 2016. PCV1 does not cause diseases in pigs. However, PCV3, similar to PCV2, is reported to be associated with several swine diseases, including porcine dermatitis and nephropathy syndrome (PDNS) and reproductive failure. PCVs are very common in domestic pigs as well as wild boars. However, PCVs have been occasionally isolated from non-porcine animals, including ruminants (such as cattle, goats, wild chamois, and roe deers), rodents (such as NMRI mice, BALB/c mice, Black C57 mice, ICR mice, Mus musculus, and Rattus rattus), canines (such as dogs, minks, foxes, and raccoon dogs), insects (such as flies, mosquitoes, and ticks), and shellfish. Moreover, PCVs are frequently reported in biological products, including human vaccines, animal vaccines, porcine-derived commercial pepsin products, and many cell lines. PCVs are also abundant in the environment, including water samples and air samples. Interestingly, PCV1 and/or PCV2 antibody or antigen has also been detected in sera, stool samples and respiratory swab samples of human, revealing zoonotic potential of PCVs. Thus, PCVs inhabit many types of reservoirs. In this review, we summarize the reservoirs of PCVs, and this information would be helpful in understanding the natural circulating status and possible cross-species transmission of PCVs.