Infectious bursal disease virus (IBDV) is an immunosuppressive pathogen causing enormous economic losses to the poultry industry across the globe. As a double-stranded RNA virus, IBDV undergoes genetic mutation or recombination in replication during circulation among flocks, leading to the generation and spread of variant or recombinant strains. In particular, the recent emergence of variant IBDV causes severe immunosuppression in chickens, affecting the efficacy of other vaccines. It seems that the genetic mutation of IBDV during the battle against host response is an effective strategy to help itself to survive. Therefore, a comprehensive understanding of the viral genome diversity will definitely help to develop effective measures for prevention and control of infectious bursal disease (IBD). In recent years, considerable progress has been made in understanding the relation of genetic mutation and genomic recombination of IBDV to its pathogenesis using the reverse genetic technique. Therefore, this review focuses on our current genetic insight into the IBDV’s genetic typing and viral genomic variation.

/pdf/genetic-insight-into-the-interaction-of-ibdv-with-host-a-3sfgoy0s.pdf

Genetic Insight into the Interaction of IBDV with Host—A Clue to the Development of Novel IBDV Vaccines

Porcine circovirus (PCV), a member of the Circoviridae family within the genus Circovirus, poses a significant economic risk to the global swine industry. PCV2, which has nine identified genotypes (a-i), has emerged as the predominant genotype worldwide, particularly PCV2d. PCV2 has been commonly found in both domestic pigs and wild boars, and sporadically in non-porcine animals. The virus spreads among swine populations through horizontal and vertical transmission routes. Despite the availability of commercial vaccines for controlling porcine circovirus infections and associated diseases, the continuous genotypic shifts from a to b, and subsequently from b to d, have maintained PCV2 as a significant pathogen with substantial economic implications. This review aims to provide an updated understanding of the biology, genetic variation, distribution, and preventive strategies concerning porcine circoviruses and their associated diseases in swine. 

Revisiting Porcine Circovirus Infection: Recent Insights and Its Significance in the Piggery Sector

Simple Summary Porcine circovirus (PCV) is widespread in the world and causes huge economic losses to the pig industry. Recently, the infection rates of PCV2 and PCV3 have been increasing year by year in China, and a new type of virus, PCV4, discovered in 2019, was found to result in complex pathogenic infection. In this study, a TaqMan-probe-based multiplex real-time PCR (qPCR) assay was developed for the differential detection of PCV2, -3, and -4. A total of 535 clinical samples from Shandong, Zhejiang, Jiangsu, and Anhui provinces, East China, were included in the analyses to evaluate the application of multiplex qPCR. The resulting data showed that the individual positive rates of PCV2, PCV3, and PCV4 were 35.33%, 40.37%, and 33.08%, respectively; the mixed infection rates of PCV2 and PCV3, PCV2 and PCV4, and PCV3 and PCV4 were 31.03%, 30.09%, and 30.84%, respectively; the mixed infection rate of the three viruses was 28.22%. The developed assay showed extreme specificity, high sensitivity, and excellent reproducibility for the simultaneous detection and differentiation of three porcine circovirus types. Abstract Porcine circovirus disease (PCVD) caused by porcine circovirus (PCV) is an important swine disease that is characterized by porcine dermatitis, nephropathy syndrome, and reproductive disorders in sows. However, disease caused by PCV2, PCV3, or PCV4 is hard to distinguish, so a rapid and sensitive detection method is urgently needed to differentiate these three types. In this study, four pairs of specific primers and the corresponding probes for PCV 2, -3, and -4, and porcine endogenous gene β-Actin as the positive internal reference index, were designed to establish a TaqMan multiplex real-time PCR (qPCR) assay for the simultaneous differential diagnosis of different types of viruses. The results showed that this assay has good specificity and no cross-reactivity with other important porcine viral pathogens. Furthermore, it has high sensitivity, with a detection limit of 101 copies/μL, and good reproducibility, with intra- and inter-group coefficients of variation below 2%. Subsequently, 535 clinical samples of suspected sow reproductive disorders collected from Shandong, Zhejiang, Anhui, and Jiangsu provinces from 2020 to 2022 were analyzed using the established assay. The results showed that the individual positive rates of PCV2, PCV3, and PCV4 were 31.03%, 30.09%, and 30.84%, respectively; the mixed infection rates of PCV2 and PCV3, PCV2 and PCV4, and PCV3 and PCV4 were 31.03%, 30.09%, and 30.84%, respectively; the mixed infection rate of PCV2, PCV3, and PCV4 was 28.22%. This indicated that this assay provides a convenient tool for the rapid detection and differentiation of PCV2, PCV3, and PCV4 in pig farms in East China. Our findings highlight that there are different types of porcine circovirus infection in pig farms in East China, which makes pig disease prevention and control difficult.

https://www.mdpi.com/2306-7381/10/1/29/pdf?version=1673493647

Development of a TaqMan-Probe-Based Multiplex Real-Time PCR for the Simultaneous Detection of Porcine Circovirus 2, 3, and 4 in East China from 2020 to 2022

Protein arginine methyltransferase (PRMT)-mediated arginine methylation is an important post-transcriptional modification that regulates various cellular processes including epigenetic gene regulation, genome stability maintenance, RNA metabolism, and stress-responsive signal transduction. The varying substrates and biological functions of arginine methylation in cancer and neurological diseases have been extensively discussed, providing a rationale for targeting PRMTs in clinical applications. An increasing number of studies have demonstrated an interplay between arginine methylation and viral infections. PRMTs have been found to methylate and regulate several host cell proteins and different functional types of viral proteins, such as viral capsids, mRNA exporters, transcription factors, and latency regulators. This modulation affects their activity, subcellular localization, protein-nucleic acid and protein-protein interactions, ultimately impacting their roles in various virus-associated processes. In this review, we discuss the classification, structure, and regulation of PRMTs and their pleiotropic biological functions through the methylation of histones and non-histones. Additionally, we summarize the broad spectrum of PRMT substrates and explore their intricate effects on various viral infection processes and antiviral innate immunity. Thus, comprehending the regulation of arginine methylation provides a critical foundation for understanding the pathogenesis of viral diseases and uncovering opportunities for antiviral therapy. 

Protein arginine methylation in viral infection and antiviral immunity

Infectious bursal disease virus still poses a great economic threat to the global poultry farming industry. Detailed information on the steps of viral genome replication is essential for the development of antiviral therapeutics. ABSTRACT Infectious bursal disease virus (IBDV) is a double-stranded RNA (dsRNA) virus belonging to the genus Avibirnavirus in the family Birnaviridae. It can cause serious failure of vaccination in young poultry birds with impaired immune systems. Post-translational modifications of the VP1 protein are essential for viral RNA transcription, genome replication, and viral multiplication. Little information is available so far regarding the exact mechanism of phosphorylation of IBDV VP1 and its significance in the viral life cycle. Here, we provide several lines of evidence that the cyclin-dependent kinase 1 (CDK1)-cyclin B1 complex phosphorylates VP1, which facilitates viral replication. We show that the CDK1-cyclin B1 specifically interacts with VP1 and phosphorylates VP1 on the serine 7 residue, located in the N-terminal 7SPAQ10 region, which follows the optimal phosphorylation motif of CDK1, p-S/T-P. Additionally, IBDV infection drives the cytoplasmic accumulation of CDK1-cyclin B1, which co-localizes with VP1, supporting the kinase activity of CDK1-cyclin B1. Treatment with CDK1 inhibitor RO3306 and knockdown of CDK1-cyclin B1 severely disrupts the polymerase activity of VP1, resulting in diminished viral replication. Moreover, the replication of S7A mutant recombinant IBDV was significantly decreased compared to that of wild-type (WT) IBDV. Thus, CDK1-cyclin B1 is a crucial enzyme which phosphorylates IBDV VP1 on serine 7, which is necessary both for the polymerase activity of VP1 and for viral replication. IMPORTANCE Infectious bursal disease virus still poses a great economic threat to the global poultry farming industry. Detailed information on the steps of viral genome replication is essential for the development of antiviral therapeutics. Phosphorylation is a common post-translational modification in several viral proteins. There is a lack of information regarding the significance of VP1 phosphorylation and its role in modulating the viral life cycle. In this study, we found that CDK1-cyclin B1 accumulates in the cytoplasm and phosphorylates VP1 on serine 7. The presence of a CDK1 inhibitor and the silencing of CDK1-cyclin B1 decrease IBDV replication. The mutation of VP1 serine 7 to alanine reduces VP1 polymerase activity, disrupting the viral life cycle, which suggests that this residue serves an essential function. Our study offers novel insights into the regulatory mechanism of VP1 phosphorylation.

Phosphorylation of VP1 Mediated by CDK1-Cyclin B1 Facilitates Infectious Bursal Disease Virus Replication

Simple Summary Since 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. Abstract 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.

/pdf/detection-of-porcine-circovirus-1-2-3-and-genetic-analysis-2enj0mvg.pdf

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

The infectious bursal disease virus (IBDV), one member of the Birnaviridae family, causes immunosuppression in young chickens by damaging the mature B cells of the bursa of Fabricius (BF), the central immune system of young chickens. The genome of IBDV is a bisegmented, double-strand RNA (dsRNA). Reverse genetics systems for IBDV allow the generation of genetically manipulated infectious virus via transfected plasmid DNA, encoding the two genomic viral RNA segments as well as major viral proteins. For this purpose, the minus-sense of both segment A and segment B are inserted into vectors between the polymerase I promoter and the corresponding terminator I. These plasmids facilitate the transcription of the viral minus-sense genome but copy the plus-sense genome as well viral protein translation depends on the activity of VP1 and VP3, when transfected into 293T cells. To further improve rescue efficiency, dual-direction promoters were generated based on the polymerase II promoter in the reverse direction in the backbone of the pCDNA3.0 vector. Therefore, the polymerase I promoter transcribes the viral minus-sense genome in the forward direction and the polymerase II promoter transcribes viral mRNA, translated into viral proteins that produce infectious IBDV. We also found that the rescue efficiency of transfecting two plasmids is significantly higher than that of transfecting four plasmids. In addition, this dual-direction promoter rescue system was used to generate R186A mutant IBDV since Arg186 is the arginine monomer-methylation site identified by LC–MS. Our data furtherly showed that the Arg186 monomer methylation mutant was due to a reduction in VP1 polymerase activity as well as virus replication, suggesting that the Arg186 methylation site is essential for IBDV replication.

/pdf/an-improved-dual-direction-promoter-driven-reverse-genetics-1tnn33t2.pdf

An Improved, Dual-Direction, Promoter-Driven, Reverse Genetics System for the Infectious Bursal Disease Virus (IBDV)

In mammals, heterogeneous ribonucleoprotein U (hnRNPU), also named as nuclear matrix protein-nuclear scaffold attachment factor (SAFA), was originally identified as a DNA/RNA interactor protein. It has been reported that human hnRNPU facilitates IFN-β generation after vesicular stomatitis virus (VSV) infection. Nevertheless, the role of chicken hnRNPU (chhnRNPU) in IFN-β regulation as well as in infectious bursal diseases virus (IBDV) replication is still unclear. Here, we found that chhnRNPU inhibits IFN-β production via interacting with MDA5 and MAVS, and facilitates IBDV replication via associating with genomic dsRNA of IBDV. Firstly, chicken hnRNPU (chhnRNPU) was widely expressed in different tissues of chickens and was distributed in the nucleus of DF-1 cells. Overexpression of chhnRNPU significantly suppresses IFN-β promoter activities induced by MDA5 and MAVS. Additionally, immunoprecipitated by dsRNA antibodies, which followed LC-MS analysis demonstrate that chhnRNPU is a partner of viral genomic dsRNA. chhnRNPU is translocated from nucleus to cytosol to co-localize with replication complex of IBDV after IBDV infection. Over-expression of chhnRNPU significantly promotes IBDV replication, which was determined by western blotting, qRT-PCR and TCID50 assay. Furthermore, knock down chhnRNPU by siRNA remarkably facilitates IFN-β production, and inhibits IBDV proliferation. These data collectively reveal that chhnRNPU positively regulates IBDV replication via negatively regulating IFN-β response. 

Characterization and functional analysis of chicken dsRNA binding protein hnRNPU.

Frequency detection of porcine circovirus-like viruses in pigs with porcine respiratory disease.

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