Respiratory Syndrome Virus Research Articles

Contribution of critical amino acid residues in the RNA-dependent RNA polymerase to the replication fidelity and viral ribavirin sensitivity of porcine reproductive and respiratory syndrome virus

The porcine reproductive and respiratory syndrome virus (PRRSV) has the highest mutation rate of any known RNA virus. The replication fidelity of RNA viruses can be modulated by subtle amino acid changes in the viral RNA-dependent RNA polymerase (RdRp). In our study, two novel amino acid substitutions (V218I and P386S) in the RdRp of PRRSV were identified under the ribavirin selection. A series of mutant viruses with single or double amino acid replacements were generated from high-fidelity PRRSV NJ-Rb and wild-type NJ-a P80 infectious cDNA clones. Subsequently, we evaluated the genetic stability, ribavirin sensitivity, and biological characteristics of the recombinant viruses. Our findings indicated that the mutation frequencies of the recombinant mutants (vI218V, vS386P, and vVP) based on NJ-Rb were significantly increased and that these recombinant viruses exhibited a loss of ribavirin resistance. The high-fidelity virus NJ-Rb was undetectable using a virus titration assay in porcine alveolar macrophages (PAMs). Our in vivo experiments demonstrated that NJ-Rb was nearly incapable of establishing infection and replicating in the lungs. The recombinant mutants vV218I, vP386S, and vIS, based on NJ-a P80, significantly increased replication fidelity and ribavirin resistance. These results indicated that PRRSV RdRp (NSP9) contained fidelity checkpoints. Furthermore, Val218 and Pro386 were identified as critical sites that determined PRRSV’s genetic stability and ribavirin resistance. These findings contribute to understanding how RdRp affects PRRSV’s genetic stability and ribavirin sensitivity and provide a theoretical basis for designing a genetically stable high-fidelity PRRSV vaccine.

Porcine reproductive and respiratory syndrome virus nsp5 inhibits the activation of the Nrf2/HO-1 pathway by targeting p62 to antagonize its antiviral activity.

Porcine reproductive and respiratory syndrome virus (PRRSV) infections often trigger oxidative stress and cytokine storms, resulting in significant tissue damage that causes fatalities in piglets and reproductive issues in sows. However, it is still unknown how oxidative stress is regulated by viral and host factors in response to PRRSV infection. Here, we found that PRRSV induced cellular oxidative stress by triggering the production of reactive oxygen species and inhibiting the expression of antioxidant enzymes. Although Nrf2 is an important redox regulator that initiates the expression of downstream antioxidant genes, PRRSV can impair the Nrf2/HO-1 pathway. The overexpression of Nrf2 showed a significant anti-PRRSV effect, and inhibiting the expression of Nrf2 promoted the proliferation of PRRSV. Further analysis showed that Nrf2 positively regulated the production of type I interferons and interferon-stimulated genes, which may contribute to its anti-PRRSV effect. By screening the PRRSV-encoded protein, we found that the PRRSV nsp5 protein can degrade Nrf2 at the protein level. Mechanistically, nsp5 promotes Nrf2-Keap1 binding affinity by inhibiting p62-mediated Keap1 sequestration and increasing Keap1 expression. Subsequently, this increased Keap1-mediated degradation of Nrf2 ubiquitination through K48-linked polyubiquitin. Furthermore, we found that the residues Tyr146 and Arg147 of nsp5 are crucial for inhibiting the activation of the p62-mediated Nrf2 antioxidant pathway. Thus, our findings uncover a novel mechanism by which PRRSV disrupts the host antioxidant defense system and highlight the crucial role of the Nrf2/HO-1 antioxidant pathway in host defense against PRRSV.IMPORTANCEOxidative stress-induced redox imbalance is a crucial pathogenic mechanism in viral infections. Nrf2 and its antioxidant genes serve as the main defense pathways against oxidative stress. However, the role of Nrf2 in the context of porcine reproductive and respiratory syndrome virus (PRRSV) infection remains unclear. In this study, we demonstrated that PRRSV infection decreased the expression of antioxidant genes of the Nrf2 signaling pathway and overexpression of Nrf2 triggered a strong anti-PRRSV effect. PRRSV nsp5 enhanced Keap1-dependent degradation of Nrf2 ubiquitination, thereby weakening cellular resistance to oxidative stress and antagonizing the antiviral activity of Nrf2. Our study further revealed a new mechanism by which PRRSV evades host antiviral innate immunity by disturbing cellular redox homeostasis, providing a new target for developing anti-PRRSV drugs.

Scalable Production of Recombinant Adeno-Associated Virus Vectors Expressing Soluble Viral Receptors for Broad-Spectrum Inhibition of Porcine Reproductive and Respiratory Syndrome Virus Type 2

Porcine reproductive and respiratory syndrome virus (PRRSV) continues to be a major threat to the global swine industry, causing significant economic losses. To address this, we developed a scalable recombinant adeno-associated virus (rAAV)-based strategy for the delivery of soluble viral receptors (SVRs) to treat and potentially eliminate PRRSV infections. This strategy involves fusing the virus-binding domains of two key cellular receptors, sialoadhesin (Sn4D) and CD163 (SRCR5-9), with an Fc fragment. We then used an insect cell–baculovirus expression vector system to produce the rAAV-SRCR59-Fc/Sn4D-Fc vector. Through a series of optimizations, we determined the best conditions for rAAV production, including a baculovirus co-infection ratio of 0.5:1.0, an initial insect cell density of 2.0 × 106 cells/mL, a fetal bovine serum concentration of 2%, and a culture temperature of 30 °C. Under these optimized conditions, we achieved a high titer of rAAV-SRCR59-Fc/Sn4D-Fc in a 2 L bioreactor, reaching 5.4 ± 0.9 × 109 infectious viral particles (IVPs)/mL. Notably, in vitro neutralization assays using a Transwell co-culture system demonstrated a 4.3 log reduction in viral titers across genetically diverse PRRSV-2 strains, including VR2332, JXA1, JS07, and SH1705. Collectively, this study provides a robust platform for large-scale rAAV production and highlights the potential of SVR-based gene therapy to address the antigenic diversity of PRRSV-2.

An intra-family conserved high-order RNA structure within the M ORF is important for arterivirus subgenomic RNA accumulation and infectious virus production

ABSTRACT Synthesis of subgenomic RNAs is a strategy commonly used by polycistronic positive-sense single-stranded RNA viruses to express 3′-proximal genes. Members of the order Nidovirales , including coronaviruses and arteriviruses, use a unique discontinuous transcription strategy to synthesize subgenomic RNAs. In this study, in silico synonymous site conservation analysis and RNA structure folding predicted the existence of intra-family conserved high-order RNA structure within the M ORF of arteriviral genomes, which was further confirmed by RNA secondary structure probing. This RNA structure was determined to be important for the transcription/accumulation of subgenomic RNAs and the production of infectious viral particles. Mutations disrupting the stability of the RNA structures significantly decreased the accumulation of multiple subgenomic RNAs. In contrast, the impact of mutagenesis on full-length genomic RNA accumulation was limited. The degree to which wild-type levels of subgenomic RNA accumulation were maintained was found to correlate with the efficiency of infectious virus production. Moreover, the thermo-stability of stems within the high-order RNA structure is also well correlated with viral replication capacity and the maintenance of subgenomic RNA accumulation. This study is the first to report an intra- Arteriviridae conserved high-order RNA structure that is located in a protein-coding region and functions as an important cis -acting element to control the accumulation/transcription of arteriviral subgenomic RNAs. This work suggests a complex regulation mechanism between genome replication and discontinuous transcription in nidoviruses. IMPORTANCE Arteriviruses are a group of RNA viruses that infect different animal species. They can cause diseases associated with respiratory/reproductive syndromes, abortion, or hemorrhagic fever. Among arteriviruses, porcine reproductive and respiratory syndrome virus (PRRSV) and equine arteritis virus (EAV) are economically important veterinary pathogens. The challenge in control of arterivirus infection reflects our limited knowledge of viral biology. In this study, we conducted a comprehensive analysis of arteriviral genomes and discovered intra-family conserved regions in the M ORF with a high-order RNA structure. The thermo-stability of the RNA structure influences sgRNA transcription/accumulation and correlates with the level of infectious virus production. Our studies provide new insight into arterivirus replication mechanisms, which may have implications for developing disease control and prevention strategies.

S146 and M148 within the mature chain domain of PSMB4 are crucial for degrading PRRSV nsp1α

Porcine reproductive and respiratory syndrome virus (PRRSV) is a single-stranded positive-sense RNA virus with an envelope. It-encoded non-structural protein 1α (nsp1α) plays a key role in evading host immune responses. Exploring the interaction between host factors and PRRSV nsp1α is crucial for understanding the mechanism of virus immune escape and virus control. Here, we constructed a cDNA library using porcine lung tissues and identified 33 potential host proteins interacting with viral nsp1α using yeast two-hybrid (Y2H) screening. These interactions were further analyzed using Gene Ontology and KEGG pathway analysis. Confocal microscopy revealed that proteasome subunit beta type-4 (PSMB4), carnosine dipeptidase 2 (CNDP2) and poly(rC) binding protein 1 (PCBP1) colocalized with viral nsp1α. The interaction between PSMB4 and nsp1α was further confirmed by Y2H and co-immunoprecipitation. PRRSV infection did not affect PSMB4 expression in both Marc-145 cells and porcine alveolar macrophages (PAMs). Overexpression of PSMB4 reduced nsp1α protein levels in a dose-dependent manner and decreased the accumulation of both viral N and nsp1α proteins in the context of PRRSV infection, while its knockdown promoted PRRSV replication. These data suggest that PSMB4 is a host restriction factor for PRRSV. Structure prediction and truncated mutant assays found that S146 and M148 within the mature chain domain of PSMB4 were crucial for binding and degrading nsp1α. These findings suggest that PRRSV nsp1α interacts with host proteins, with PSMB4 specifically binding to degrade nsp1α, thereby inhibiting PRRSV replication.

Evaluating postmortem tongue fluids as a tool for monitoring PRRSV and IAV in the post-wean phases of swine production

BackgroundPorcine reproductive and respiratory syndrome virus (PRRSV) and influenza A virus (IAV) are swine pathogens that can significantly impact the performance of post-weaning pigs. While oral fluid (OF) samples are widely used for monitoring these viruses, postmortem tongue fluid (TF) samples present a cost-effective alternative with potential advantages in viral detection. This study aimed to compare the performance of TF and OF samples collected from nursery and finishing pig herds in detecting PRRSV and IAV using reverse transcription-quantitative polymerase chain reaction (RT-qPCR). A Bayesian latent class model was used to estimate diagnostic sensitivity and specificity for TF and OF under the assumption of conditional independence. The study also examined the relationship between mortality rates and RT-qPCR outcomes, the success rate of Sanger sequencing for the PRRSV ORF-5 region, and the effect of pooling daily aggregated TF samples on the probability of PRRSV detection.ResultsIAV was detected in 34.9% of OF samples and 30.2% of TF samples, while PRRSV was identified in 67.4% of OF and 53.5% of TF samples. TF samples had a significantly lower mean Ct for PRRSV (29.1) compared to OF samples (32.8) but had a similar Ct (30.9) to OF (29.7) for IAV. The hierarchical latent class Bayesian model estimated the sensitivity and specificity values for OF as 37.3% and 61.7% for IAV, and 64.3% and 35.1% for PRRSV. The estimated sensitivity and specificity values for TF were 33.5% and 66.0% for IAV, and 53.0% and 47.0% for PRRSV. Among 22 matched TF and OF pairs submitted for PRRSV sequencing, 45.5% of OF samples and 63.6% of TF samples were successfully sequenced, with the higher success rate for TF attributed to having lower Ct values. Additionally, mortality rates were notably higher when PRRSV was detected, especially in cases with concurrent IAV detection. Regarding sample pooling, our results indicated that pooling TF samples significantly increased detection probabilities, with a 1/7 dilution achieving a 79% RT-qPCR detection rate, compared to a detection rate of 14.3% when testing a single day's TF sample from a week with only one positive day.ConclusionThe findings support the use of TF samples as a viable complement or alternative to OF samples for PRRSV and IAV surveillance in post-weaning pigs when mortalities are available. The cost-efficiency of TF sampling can enhance monitoring compliance, improve early pathogen detection, and facilitate timely responses to emerging threats in swine production. This study advocates for the adoption of TF as a risk-based sampling strategy in nursery and grow-finish settings, complementing live animal samples such as OF, ultimately contributing to better herd health management.

Evaluation of an electrostatic precipitator in mitigating the transmission of airborne viruses in experimentally infected pigs

Airborne viruses spread rapidly in animal premises, which makes them difficult to contain. Electrostatic precipitators (ESP) are air cleaning devices that charge airborne particles and electrophoretically deposit them on collection surfaces, thereby removing them from an airstream. We evaluated the effect of a single-stage wire-plate ESP on mitigating airborne transmission of influenza A virus (IAV) and porcine reproductive and respiratory syndrome virus (PRRSV), using experimentally infected pigs. Inoculated pigs were placed in isolators upstream of the ESP, and sentinel pigs were placed downstream in an isolator. The airflow moved unidirectionally from inoculated pigs to sentinel pigs. Nasal swabs of pigs, air samples and surface wipes from all the isolators were collected daily and tested by RT-qPCR. Without the ESP powered, sentinel pigs tested positive within 1 day of exposure to IAV aerosols and 2 days to PRRSV aerosols. Airborne IAV RNA was detected upstream and downstream of the ESP in particles ranging from 0.22 μm to > 8 μm. In contrast, with the ESP powered, sentinel pigs tested positive after 5–6 days of exposure to IAV aerosols, and 7–8 days to PRRSV aerosols. Limited levels of IAV RNA were detected in air samples in the downstream isolator before sentinel pigs tested positive. The RNA-based virus removal efficiency of the ESP ranged from 96.91 to 99.97%, with higher removal observed in particles > 6.5 μm. Under the conditions of this study, the ESP efficiently removed IAV aerosol particles and delayed the onset of IAV and PRRSV infections in the sentinel pigs. Our study shows the potential of the ESPs to help prevent the spread of airborne viruses in agricultural animal farming facilities.

Genetic evolution analysis of PRRSV ORF5 gene in five provinces of Northern China in 2024

BackgroundPorcine reproductive and respiratory syndrome (PRRS) was first discovered in North America in 1987, and since then it has been spread widely all over the world. The prevalence of PRRS has caused significantly economic losses to pig industry in many countries.ObjectivesInvestigate the prevalence and genetic evolution of porcine reproductive and respiratory syndrome virus (PRRSV) in five provinces of northern China.Methods190 samples suspected of PRRS were collected from 28 pig farms in five provinces of northern China. The PRRSV ORF7 and ORF5 gene were detected by RT-PCR, and the ORF5 gene were sequenced for the homology and genetic evolution analysis.ResultsThe positive samples of ORF7 gene were 50, and its positive rate was 26.32%. The positive samples of ORF5 gene were 48, and its positive rate was 25.26%. The sequenced results of the ORF5 gene showed that 48 positive samples all belonged to PRRSV-2. Among them, 26 samples were NADC34-like strains, 17 samples were NADC30-like strains, and 5 samples were classical strains. The amino acid sequence analysis of PRRSV GP5 indicated that there was a deletion at the 37th amino acid in 4 NADC30-like strains. The amino acids of the transmembrane region 1 in all positive strains are relatively conserved, and multiple amino acid mutations were observed in the signal peptide, transmembrane region 2, and B cell epitope. The amino acid mutations were different in different strains and regions. The above results demonstrated that the complexity and diversity of PRRSV genetics.ConclusionThe strains from lineage 1 became the dominant strains in five provinces of northern China in 2024. The positive rate of NADC34-like strains was the highest in Heilongjiang Province and the NADC30-like strains were the most prevalent in these regions. The genetic evolution of PRRSV presented a complex trend. This study provided the data support for understanding PRRSV variation and for PRRS prevention and control in five provinces of northern China.

A qualitative risk assessment for the importation of fresh porcine semen into Australia.

The importation of fresh boar semen was qualitatively assessed for the biosecurity risk of introducing specific diseases into the Australian pig herd. The methodology used was as described by the World Organisation for Animal Health (WOAH) and applied for official import risk assessments undertaken by Australia. It was found that the risk of introducing porcine reproductive and respiratory syndrome virus, Senecavirus A, porcine epidemic diarrhoea virus and African swine fever virus exceeded Australia's appropriate level of protection even when WOAH mitigations were applied. Findings of the assessment also support predictions of new diseases transmitted via semen emerging that may cause serious and/or irreversible harm to the Australian pig industry.

Emergence, prevalence and evolution of porcine reproductive and respiratory syndrome virus 1 in China from 1994 to 2024.

Emergence, prevalence and evolution of porcine reproductive and respiratory syndrome virus 1 in China from 1994 to 2024.

Machine learning models provide modest accuracy in predicting clinical impact of porcine reproductive and respiratory syndrome type 2 in Canadian sow herds.

To determine the predictive potential of the open reading frame 5 nucleotide sequence of porcine reproductive and respiratory syndrome (PRRS) virus and the basic demographic data on the severity of the impact on selected production parameters during clinical PRRS outbreaks in Ontario sow herds. A retrospective longitudinal study of clinical outbreaks in Ontario sow herds at various points between September 5, 2009, and February 5, 2019, was conducted using herds as units of analysis. Data were gathered from study sow farms in Ontario at the start of each clinical outbreak. Six machine learning models and 2 different genetic input structures of open reading frame 5 sequences were utilized to predict the impact on abortion and preweaning mortality. Extreme boosting machine learning models with genetic data represented through 2-dimensional multiple correspondence analysis had the highest accuracy when predicting clinical outcomes (60.8% [SD = 12.4%] and 74.4% [SD = 13.2%]) for abortion and preweaning mortality outcomes, respectively. The mean sensitivity of classifying outbreaks with a high impact on abortion was 50%, with a specificity of 89.2%. The mean sensitivity of classifying outbreaks with high preweaning mortality was 56.2%, with a specificity of 85.2%. The data and methods utilized herein exhibited improvement in accuracy over the baseline; however, this increase was not sufficient to warrant field implementation. Predictive models based on observed data could assist practitioners in linking the genetics of the PRRS virus with clinical impact in clinical settings. Models trained in this study show promise for PRRS clinical impact prediction.

Development of a triplex crystal digital PCR for the detection of PRCoV, PRRSV, and SIV.

Porcine respiratory coronavirus (PRCoV), porcine reproductive and respiratory syndrome virus (PRRSV), and swine influenza virus (SIV) are important pathogens of significant infectious diseases. They cause similar clinical respiratory symptoms, including fever, cough, runny nose, and respiratory distress, which makes these diseases difficult to distinguish from each other. In this study, three pairs of specific primers and TaqMan probes were designed for the conserved regions of the PRCoV S gene, PRRSV N gene, and SIV M gene, respectively. The annealing temperature, primer and probe concentrations, and reaction cycle were optimized, and a triplex crystal digital PCR (cdPCR) assay was established for the detection of PRCoV, PRRSV, and SIV. According to the test results, the assay was capable of specifically detecting PRCoV, PRRSV, and SIV, and there was no cross-reaction with other control swine viruses. Based on the Poisson distribution analysis, the limits of detection (LODs) for PRCoV, PRRSV, and SIV were 6.00, 5.75 and 6.00 copies/reaction, respectively, and the sensitivity was 26 times higher than those of the corresponding multiplex RT-qPCR. The coefficients of variation (CVs) of the intra-assay and inter-assay ranged from 0.19 to 1.84%. The assay was used to test 1,657 clinical samples, and the positivity rates of PRCoV, PRRSV, and SIV were 1.15, 12.79, and 2.05%, respectively. It showed diagnostic sensitivity and specificity of 100 and 99.82% for PRCoV, 100 and 99.24% for PRRSV, and 100 and 99.69% for SIV, respectively. These results indicated that the triplex cdPCR assay has strong specificity, high sensitivity, and excellent repeatability, which provides a valuable tool for the detection and differentiation of PRCoV, PRRSV, and SIV.

Cinnamaldehyde Inhibits the Replication of Porcine Reproductive and Respiratory Syndrome Virus Type 2 In Vitro

Globally, the swine industry suffers significant economic losses due to the presence of porcine reproductive and respiratory syndrome virus (PRRSV). Unfortunately, existing vaccines fail to offer adequate protection against the various strains of PRRSV, and there are currently no specific treatments available for this virus. In this study, we screened four natural products and identified cinnamaldehyde (CA) as an effective inhibitor of PRRSV infection in Marc-145 cells. CA could achieve an inhibition rate of up to 93% on PRRSV N protein at 160 μM. Mechanistically, CA exerted anti-PRRSV ability in different treatment modes. CA could directly interact with PRRSV particles. Cinnamaldehyde blocks the binding, entry, replication, and release of PRRSV. Furthermore, a significant reduction in dsRNA levels was observed in the CA-treated groups compared to the control groups. In conclusion, our research demonstrated that CA could inhibit essential stages of the PRRSV lifecycle: binding, entry, replication, and release. CA could directly interact with PRRSV. Additionally, CA disrupted the expression of dsRNA during viral replication, thereby suppressing in vitro PRRSV replication in Marc-145 cells. This study provides crucial perspectives on the potential application of CA for the prevention and treatment of PRRS.

Porcine Reproductive and Respiratory Syndrome Virus Prevalence and Pathogenicity of One NADC34-like Virus Isolate Circulating in China

Porcine Reproductive and Respiratory Syndrome Virus (PRRSV) is one of the most significant infectious agents threatening the global pig industry. Due to its high mutation and recombination rates, the prevalence of PRRSV in domestic pig populations is complex. To better understand the epidemiology of PRRSV, we conducted a large-scale investigation in eastern China, focusing on pig farms with a history of high abortion rates. A total of 14,934 pig samples were collected from 11 sow farms and 53 fattening farms across three provinces. Among these, 13.0% of the collected samples tested positive for PRRSV, with specific prevalence rates of 19.7% in sows and 12.4% in piglets. Genetic evolution analysis of the GP5 gene from 43 PRRSV strains identified in this study revealed that NADC30-like, NADC34-like, and HP-PRRSV were the predominant lineages in domestic pig farms. The NADC30-like genotype was the most dominant and had evolved into three subgenotypes, while the NADC34-like strains had diverged into two subgenotypes. Further analysis of the Nsp2 gene from 18 strains indicated that the NSP2 gene of multiple NADC34-like strains was closely related to that of the NADC30-like, suggesting that the NADC34-like strains are primarily recombinant viruses. Sequence comparison of the Nsp2 gene showed that both NADC30-like and NADC34-like viruses share 111 amino acid deletions at positions 322–433 and 21 amino acid deletions at positions 539–558 in the Nsp2 gene coding region. For the first time, the pathogenicity of a representative NADC34-like virus isolated in China was evaluated in pregnant sow. The results showed that infected sows exhibited an increased body temperature, ear cyanosis, and typical edema and cyanosis of the external genitalia. Moreover, all infected sows experienced miscarriage, with 100% of the aborted piglets being stillbirths exhibiting a high virus load. These findings indicate that this NADC34-like virus is highly virulent to sows.

Development and application of a blocking ELISA method based on Cap protein for detecting antibodies against porcine circovirus 2

ABSTRACT Porcine circovirus type 2 (PCV2) serves as the key pathogen linked to porcine circovirus-associated disease, representing a considerable risk to the worldwide swine industry. A blocking enzyme-linked immunosorbent assay (ELISA) was established to identify antibodies specifically targeting PCV2, employing recombinant Cap protein as the antigen and a monoclonal antibody against PCV2 Cap protein as the detector antibody. Utilizing receiver operating characteristic curve analysis, a cutoff value of 33.8% was determined to distinguish between positive and negative serum samples. The sensitivity and specificity of this blocking ELISA method were reported at 94.7% and 96.1%, respectively. Notably, this approach exclusively identified antibodies for PCV2, showing no cross-reactivity with antibodies related to African swine fever virus (ASFV), Porcine epidemic diarrhea virus (PEDV), Porcine pseudorabies virus (PRV), and porcine reproductive and respiratory syndrome virus. Both intra-assay and inter-assay coefficients of variation were less than 10%. In a comparison involving 402 porcine serum samples, the agreement rate with a commercial indirect ELISA kit reached 98.76%, with a kappa value of 0.888, reflecting high concordance between the two testing methods. This study showcases the blocking ELISA method as an efficient and standardized approach for serological monitoring of PCV2 in swine populations and for assessing seroconversion in vaccinated pigs. IMPORTANCE Porcine circovirus type 2 (PCV2) has become recognized as a pathogen of significant economic concern within the swine industry. PCV2 mainly affects the immune systems of pigs, leading to a reduction in lymphocytes and resulting in immune suppression in the affected animals. Co-infection with other porcine pathogens can enhance PCV2 infection and exacerbate porcine circovirus disease. Currently, the kits available for detecting PCV2 antibodies primarily employ indirect enzyme-linked immunosorbent assay (ELISA); however, this method is prone to false positives. In contrast, the blocking ELISA method offers enhanced specificity and provides a more straightforward interpretation of results. Previous studies utilizing blocking ELISA for PCV2 antibody detection have depended on plates coated with purified PCV2 virus, a process that is both technically challenging and time-consuming. Consequently, there is a pressing need to develop a new blocking ELISA method that is more efficient to detect antibodies against PCV2.

Impact of weaning procedures on Porcine Reproductive and Respiratory Syndrome Virus (PRRSV) circulation in the nursery section

The impact of different weaning strategies on the downstream circulation of PRRSV has not been widely described. It is, however, believed that mixing pigs of different age groups is increasing the risk of PRRSV circulation in the nursery section. In this study, pigs were sampled in three herds that performed “mixed at weaning (MIX)” and three herds that performed “all in/all out at weaning (AIAO)”. MIX included holding underweighted piglets back in containers for two weeks and then move them to nursery facilities with newly weaned piglets from subsequent batches. Oral fluid samples were collected from four batches of pigs in each herd, three times from weaning until 30 kg for each batch, and tested for PRRSV and PRRSV antibodies. Herds that performed MIX at weaning had an eightfold increase in risk of detecting PRRSV in oral fluids compared to herds with AIAO. In total, 41 oral fluid samples from eight batches in MIX herds and five oral fluid samples from two batches in AIAO herds tested positive for PRRSV. The PRRSV ELISA S/P ratio in oral fluid samples from weaners seem to decrease in most of the batches in the AIAO herds and to increase in most MIX herds. In addition to oral fluids, tongue tip samples were collected from dead pigs and tested for PRRSV. In 17 of 23 batches the results of the tongue tip samples correlated with the results of the oral fluid samples (κ = 0.44) indicating a good agreement between the two materials for sampling. Overall, the results of the study confirmed that the weaning strategy had a significant impact on the circulation of PRRSV post weaning.

Formononetin and mizoribine inhibit Porcine Reproductive and Respiratory Syndrome Virus replication in vitro.

This study delves into the antiviral efficacy of Formononetin (FMN) and Mizoribine (MZR) against the Porcine Reproductive and Respiratory Syndrome Virus (PRRSV), a virus with a considerable economic impact and a current void in effective treatments. FMN and MZR were found to inhibit various PRRSV strains in vitro, predominantly in the early stages of viral infection. Noteworthy was the observation of their synergistic effects when combined with Ribavirin. The study underscores the antiviral potential of FMN and MZR, particularly emphasizing their low cytotoxicity at specific concentrations. These results position FMN and MZR as promising antiviral agents against PRRSV, underscoring their low cytotoxicity and efficacy in early-stage viral inhibition. Such findings pave the way for their potential inclusion in future PRRSV management strategies.

Dynamic interplay between RNA N6-methyladenosine modification and porcine reproductive and respiratory syndrome virus infection

N6-methyladenosine (m6A) has attracted significant attention for its role in regulating the complex interaction between viruses and host cells. Porcine reproductive and respiratory syndrome virus (PRRSV) is a significant pathogen affecting swine health worldwide. Here, we first identified seven m6A-enriched peaks in PRRSV genomic RNA by m6A RNA immunoprecipitation sequencing (m6A-seq). Moreover, functional analyses revealed a positive correlation between the m6A modification level and PRRSV replication. Treatment with the universal methylation inhibitor 3-deazaadenosine (3-DAA) effectively suppressed PRRSV replication in a dose-dependent manner. Furthermore, m6A-seq was also used to determine the m6A landscape of the transcriptome in PAMs infected with pandemic or highly pathogenic PRRSV strains. Among the 4677 transcripts exhibiting altered m6A modification levels, the MAPK14 gene and the p38/MAPK signalling pathway emerged as preliminary targets of m6A-mediated epigenetic regulation during PRRSV infection. These findings provide new insights into the epigenetic mechanisms underlying PRRSV infection and may facilitate the development of anti-PRRSV therapeutics.

Validation of Polymorphisms Associated with the Immune Response After Vaccination Against Porcine Reproductive and Respiratory Syndrome Virus in Yorkshire Gilts

Validation of Polymorphisms Associated with the Immune Response After Vaccination Against Porcine Reproductive and Respiratory Syndrome Virus in Yorkshire Gilts

Altered structural and transporter-related gene expression patterns in the placenta play a role in fetal demise during Porcine reproductive and respiratory syndrome virus infection

BackgroundPorcine reproductive and respiratory syndrome virus (PRRSV) can be transmitted across the maternal-fetal-interface from an infected gilt to her fetuses. Although fetal infection status and disease outcomes vary, the mechanisms are not completely understood. The objective was to assess targeted placental structural and transporter-related gene expression patterns. At day 85 of gestation pregnant pigs were challenged with PRRSV, and at 12 days post maternal infection sows and fetuses were sacrificed, and the placental tissue was collected. Grouping of fetuses was by preservation status and PRRS viral load (VL): control (CTRL, n = 14), viable and low VL fetus (VIA_LVF, n = 15), viable and high VL fetus (VIA_HVF, n = 21), meconium mild and low VL fetus (MECm_LVF, n = 14), meconium mild and high VL fetus (MECm_HVF, n = 14), and meconium severe and high VL fetus (MECs_HVF, n = 13). NanoString was used to evaluate the expression of 86 genes: actin cytoskeleton signaling, arachidonic acid pathway, integrin signaling, intercellular junctions, transporters, and VEGF signaling. Statistical analyses were performed using Limma with P ≤ 0.05 considered significant.ResultsWe identified 1, 7, 0, 29, and 39 differentially expressed genes in VIA_LVF, VIA_HVF, MECm_LVF, MECm_HVF, and MECs_HVF, respectively, contrasted to CTRL. Placental transporter genes were significantly impacted (i.e., downregulation of SLC1A3, SLC1A5, SLC2A1, SLC2A3, SLC2A5, SLC2A10, SLC2A12, SLC7A4, SLC16A5, SLC16A10, and SLC27A6; and upregulation of SLC2A2, SLC16A3, and SLC27A4), compared to CTRL. Actin cytoskeleton signaling (ARHGEF6 and ARHGEF7), arachidonic acid (PTGES3 and PTGIS), integrin signaling (FN1 and ITGB6), intercellular junctions (CDH3 and CDH11), and VEGF signaling (MAPK3 and HPSE) gene groupings were significantly impacted, compared to CTRL.ConclusionData reported here indicate that fetal PRRSV infection levels rather than fetal demise is necessary for transcriptional dysregulation of the fetal placenta, with a tendency towards more downregulation in the target gene sets among susceptible fetuses. These results generally support that in susceptible fetuses there is altered solute transportation, placental structural integrity, and reduced angiogenesis. The data described here is associated with fetal PRRS resistance/resilience and susceptibility.