Porcine Reproductive And Respiratory Syndrome Virus RNA Research Articles

Gene expression in heart, kidney, and liver identifies possible mechanisms underpinning fetal resistance and susceptibility to in utero PRRSV infection

Porcine reproductive and respiratory syndrome (PRRS) is one of the costliest diseases to pork producers worldwide. We tested samples from the pregnant gilt model (PGM) to better understand the fetal response to in-utero PRRS virus (PRRSV) infection. Our goal was to identify critical tissues and genes associated with fetal resilience or susceptibility. Pregnant gilts (N=22) were infected with PRRSV on day 86 of gestation. At 21 days post maternal infection, the gilts and fetuses were euthanized, and fetal tissues collected. Fetuses were characterized for PRRS viral load in fetal serum and thymus, and preservation status (viable or meconium stained: VIA or MEC). Fetuses (N=10 per group) were compared: uninfected (UNIF; <1 log/µL PRRSV RNA), resilient (HV_VIA, >5 log virus/µL but viable), and susceptible (HV_MEC, >5 log virus/µL with MEC). Gene expression in fetal heart, kidney, and liver was investigated using NanoString transcriptomics. Gene categories investigated were hypothesized to be involved in fetal response to PRRSV infection: renin- angiotensin-aldosterone, inflammatory, transporter and metabolic systems. Following PRRSV infection, CCL5 increased expression in heart and kidney, and ACE2 decreased expression in kidney, each associated with fetal PRRS susceptibility. Liver revealed the most significant differential gene expression: CXCL10 decreased and IL10 increased indicative of immune suppression. Increased liver gene expression indicated potential associations with fetal PRRS susceptibility on several systems including blood pressure regulation (AGTR1), energy metabolism (SLC16A1 and SLC16A7), tissue specific responses (KL) and growth modulation (TGFB1). Overall, analyses of non-lymphoid tissues provided clues to mechanisms of fetal compromise following maternal PRRSV infection.

Detection of PRRSV-1 in tongue fluids under experimental and field conditions and comparison of different sampling material for PRRSV sow herd monitoring.

Infection with porcine reproductive and respiratory syndrome virus (PRRSV) leads to significant economic losses worldwide. One of the initial measures following an outbreak is to stabilise the herd and to prevent vertical transmission of PRRSV. The objective of this study was to detect PRRSV in different sampling material, both in an experimental model and on a commercial piglet producing farm, with a focus on evaluating the suitability of tongue fluid samples. In the experimental model, PRRSV negative pregnant gilts were infected with PRRSV-1 AUT15-33 on gestation day 85 and necropsy of gilts and foetuses was performed three weeks later. 38.3% of individual foetal serum and 39.4% of individual foetal thymus samples were considered PRRSV RT-qPCR positive. Tongue fluids from individual foetuses showed a 33.0% positivity rate. PRRSV RNA was detected in all but one sample of litter-wise pooled processing fluids and tongue fluids. In the field study, the investigated farm remained PRRSV positive and unstable for five consecutive farrowing groups after the start of the sampling process. Tongue fluid samples pooled by litter in the first investigated farrowing group had a 54.5% positivity rate, with the overall highest viral load obtained in the field study. In this farrowing group, 33.3% of investigated litter-wise pooled processing fluid samples and all investigated serum samples (pools of 4-6 individuals, two piglets per litter) were considered positive. Across all investigated farrowing groups, tongue fluid samples consistently showed the highest viral load. Moreover, tongue fluid samples contained the virus in moderate amounts for the longest time compared to the other investigated sampling material. It can be concluded that the viral load in individual foetuses is higher in serum or thymus compared to tongue fluid samples. However, litter-wise pooled tongue fluid samples are well-suited for detecting vertical transmission within the herd, even when the suspected prevalence of vertical transmission events is low.

67 Decontamination of a feed manufacturing environment following inoculation of porcine epidemic diarrhea virus, porcine reproductive and respiratory virus, and Seneca Valley virus 1

Abstract Feed mill decontamination is difficult because equipment is not designed to be cleaned with water. Alternate strategies may improve the ability of a mill to decontaminate in the event of viral contamination. The objective was to evaluate different decontamination strategies within a mill following the inoculation of porcine epidemic diarrhea virus (PEDV), porcine reproductive and respiratory syndrome virus (PRRSV), and Seneca Valley virus 1 (SVV1) contaminated feed. A batch of feed was inoculated with PEDV, PRRSV, and SVV1 and ran through a mixer, bucket elevator, corn cleaner, screw conveyor, distributor head, and spout. Following mill inoculation, decontamination strategies were implemented with environmental samples collected after each decontamination step. Decontamination strategies included: 1) complete facility decontamination including organic matter removal with hot water power washing, disinfection with 1% peroxygen (Virkon S, Lanxess, Germany) followed by a rinse, and a second disinfection with 5% sodium hypochlorite (Clorox, Oakland, CA) followed by a heating period for 48 h once the facility reached 60°C; 2) chlorine dioxide application (ProOxine AH, Bio-Cide International, Inc., Norman, OK); 3) organic matter removal using vacuums (Ridge Tool Company, Elyria, OH) and chlorine dioxide application; 4) heat up with portable electric heaters for exactly 48 h; and 5) organic matter removal and heat up with portable heaters for exactly 48 h. Samples were analyzed via triplex PCR. Cycle threshold and proportion PCR positive were analyzed using SAS GLIMMIX v 9.4 (SAS, Inc., Cary, NC). A swine bioassay was completed to determine the infectivity of samples following the final decontamination step of each treatment. A treatment × decontamination step × location interaction was observed (P &amp;lt; 0.05) for SVV1, where less RNA was detected post-treatment compared with pre-treatment following complete facility decontamination on surfaces including the mixer, corn cleaner, screw conveyor, and floor around the feed discharge (P &amp;lt; 0.05). There was no statistical difference between pre- and post-treatment samples for other combinations of decontamination treatment and location (P &amp;gt; 0.05), although numerical increases in Ct were generally observed following treatment. Across all treatments, the act of decontamination reduced detectable PEDV (P &amp;lt; 0.05) and PRRSV (P &amp;lt; 0.05) RNA when compared with samples immediately following inoculation, but complete facility decontamination was the only treatment where PEDV and PRRSV was non-detectable in all locations. Pigs given samples post-treatment showed no evidence of SVV1 or PEDV infection. Although PRRSV RNA was rarely found via PCR from samples collected within the mill, PRRSV infection was observed in pigs given the chlorine dioxide with and without organic matter removal treatments and the organic matter removal plus heat treatment. Overall, all treatments reduced detectable RNA for all viruses between the inoculation step and the final decontamination step; however, PRRSV particles remained infectious following decontamination regardless of PCR results in a swine bioassay.

Evaluation of Truck Cab Decontamination Procedures following Inoculation with Porcine Epidemic Diarrhea Virus and Porcine Reproductive and Respiratory Syndrome Virus.

This experiment aimed to evaluate commercially available disinfectants and their application methods against porcine epidemic diarrhea virus (PEDV) and porcine reproductive and respiratory syndrome virus (PRRSV) on truck cab surfaces. Plastic, fabric, and rubber surfaces inoculated with PEDV or PRRSV were placed in a full-scale truck cab and then treated with one of eight randomly assigned disinfectant treatments. After application, surfaces were environmentally sampled with cotton gauze and tested for PEDV and PRRSV using qPCR duplex analysis. There was a disinfectant × surface interaction (p < 0.0001), indicating a detectable amount of PEDV or PRRSV RNA was impacted by disinfectant treatment and surface material. For rubber surfaces, 10% bleach application had lower detectable amounts of RNA compared to all other treatments (p < 0.05) except Intervention via misting fumigation, which was intermediate. In both fabric and plastic surfaces, there was no evidence (p > 0.05) of a difference in detectable RNA between disinfectant treatments. For disinfectant treatments, fabric surfaces with no chemical treatment had less detectable viral RNA compared to the corresponding plastic and rubber (p < 0.05). Intervention applied via pump sprayer to fabric surfaces had less detectable viral RNA than plastic (p < 0.05). Furthermore, 10% bleach applied via pump sprayer to fabric and rubber surfaces had less detectable viral RNA than plastic (p < 0.05). Also, a 10 h downtime, with no chemical application or gaseous fumigation for 10 h, applied to fabric surfaces had less detectable viral RNA than other surfaces (p < 0.05). Sixteen treatments were evaluated via swine bioassay, but all samples failed to produce infectivity. In summary, commercially available disinfectants successfully reduced detectable viral RNA on surfaces but did not eliminate viral genetic material, highlighting the importance of bioexclusion of pathogens of interest.

ZNF283, a Krüppel-associated box zinc finger protein, inhibits RNA synthesis of porcine reproductive and respiratory syndrome virus by interacting with Nsp9 and Nsp10

Porcine reproductive and respiratory syndrome virus (PRRSV) is a viral pathogen with substantial economic implications for the global swine industry. The existing vaccination strategies and antiviral drugs offer limited protection. Replication of the viral RNA genome encompasses a complex series of steps, wherein a replication complex is assembled from various components derived from both viral and cellular sources, as well as from the viral genomic RNA template. In this study, we found that ZNF283, a Krüppel-associated box (KRAB) containing zinc finger protein, was upregulated in PRRSV-infected Marc-145 cells and porcine alveolar macrophages and that ZNF283 inhibited PRRSV replication and RNA synthesis. We also found that ZNF283 interacts with the viral proteins Nsp9, an RNA-dependent RNA polymerase, and Nsp10, a helicase. The main regions involved in the interaction between ZNF283 and Nsp9 were determined to be the KRAB domain of ZNF283 and amino acids 178–449 of Nsp9. The KRAB domain of ZNF283 plays a role in facilitating Nsp10 binding. In addition, ZNF283 may have an affinity for the 3' untranslated region of PRRSV. These findings suggest that ZNF283 is an antiviral factor that inhibits PRRSV infection and extend our understanding of the interactions between KRAB-containing zinc finger proteins and viruses.

Transmission of porcine reproductive and respiratory syndrome virus in domestic pigs via oral ingestion of feed material.

The purpose of this case study was to describe the transmission of porcine reproductive and respiratory syndrome virus (PRRSV) under field and experimental conditions via the consumption of PRRSV-positive swine feed. 1 domestic swine breeding herd and 20 laboratory-maintained experimental domestic pigs. A 2,500-sow PRRSV-naïve biosecure breeding herd became infected during the autumn months. It experienced a feed outage involving a specific bin on October 23 (day 0), with the bin refilled on October 24 (day 1). From October 28 to 30 (days 5 to 7), signs of anorexia and hyperemia were observed in 30 gestating sows after consuming feed from this bin. On November 1 (day 9), blood samples from 10 affected sows were PRRSV positive by reverse transcriptase PCR. In contrast, sows in the same room that had consumed feed from other bins were clinically normal and PRRSV negative. To investigate whether the feed delivery introduced PRRSV to the herd, on November 2 (day 10) 4 samples of feed material from the interior walls of the index bin were collected and tested by reverse transcriptase PCR. All 4 samples were positive for PRRSV RNA with cycle threshold values ranging from 26 to 29. Nucleic acid sequencing indicated that the open reading frame 5 region of the PRRSV in feed samples was 100% homologous to PRRSV from index cases. To assess viability of the virus, PRRSV-naïve pigs were allowed to consume the index feed bin samples and became infected with PRRSV based on viral RNA in oral fluid samples, clinical signs, and postmortem lesions. These results suggest that feed was a likely source of PRRSV introduction to the herd. This is the first report of PRRSV transmission through feed.

A cross-sectional assessment of PRRSV nucleic acid detection by RT-qPCR in serum, ear-vein blood swabs, nasal swabs, and oral swabs from weaning-age pigs under field conditions.

The porcine reproductive and respiratory syndrome virus (PRRSV) continues to challenge swine production in the US and most parts of the world. Effective PRRSV surveillance in swine herds can be challenging, especially because the virus can persist and sustain a very low prevalence. Although weaning-age pigs are a strategic subpopulation in the surveillance of PRRSV in breeding herds, very few sample types have been validated and characterized for surveillance of this subpopulation. The objectives of this study, therefore, were to compare PRRSV RNA detection rates in serum, oral swabs (OS), nasal swabs (NS), ear-vein blood swabs (ES), and family oral fluids (FOF) obtained from weaning-age pigs and to assess the effect of litter-level pooling on the reverse transcription-quantitative polymerase chain reaction (RT-qPCR) detection of PRRSV RNA. Three eligible PRRSV-positive herds in the Midwestern USA were selected for this study. 666 pigs across 55 litters were sampled for serum, NS, ES, OS, and FOF. RT-qPCR tests were done on these samples individually and on the litter-level pools of the swabs. Litter-level pools of each swab sample type were made by combining equal volumes of each swab taken from the pigs within a litter. Ninety-six piglets distributed across 22 litters were positive by PRRSV RT-qPCR on serum, 80 piglets distributed across 15 litters were positive on ES, 80 piglets distributed across 17 litters were positive on OS, and 72 piglets distributed across 14 litters were positive on NS. Cohen's kappa analyses showed near-perfect agreement between all paired ES, OS, NS, and serum comparisons (). The serum RT-qPCR cycle threshold values (Ct) strongly predicted PRRSV detection in swab samples. There was a ≥ 95% probability of PRRSV detection in ES-, OS-, and NS pools when the proportion of positive swab samples was ≥ 23%, ≥ 27%, and ≥ 26%, respectively. ES, NS, and OS can be used as surveillance samples for detecting PRRSV RNA by RT-qPCR in weaning-age pigs. The minimum number of piglets to be sampled by serum, ES, OS, and NS to be 95% confident of detecting ≥ 1 infected piglet when PRRSV prevalence is ≥ 10% is 30, 36, 36, and 40, respectively.

Effect of extrinsic factors on the detection of PRRSV and a porcine-specific internal sample control in serum, oral fluid, and fecal specimens tested by RT-rtPCR.

We characterized the effect of 1) temperature × time, 2) freeze-thaw cycles, and 3) high porcine reproductive and respiratory syndrome virus (PRRSV) RNA concentrations on the detection of PRRSV and a porcine-specific internal sample control (ISC) in serum, oral fluid, and fecal samples using a commercial PRRSV RT-rtPCR assay (Idexx). In study 1, the effect of temperature × time on PRRSV and ISC detection was shown to be specimen dependent. In serum stored at 4, 10, or 20°C, PRRSV detection was consistent for up to 168 h, but storage at 30°C reduced detectable PRRSV RNA. ISC RNA was stable in serum held at 4 and 10°C, but not at 20 and 30°C. In contrast, PRRSV and ISC RNAs in oral fluid and fecal samples continuously decreased at all temperature × time treatments. Based on these data, serum samples should be stored at ≤ 20°C to optimize PRRSV RNA detection. Oral fluid and fecal samples should be frozen in a non-self-defrosting freezer until tested. In study 2, freeze-thaw cycles had little impact on PRRSV and ISC detection, but more so in oral fluids than serum or fecal samples. Thus, freeze-thaw cycles in oral fluids should be minimized before RT-rtPCR testing. In study 3, the ISC was not affected by high concentrations of PRRSV RNA in serum, oral fluid, or fecal samples. It should not be assumed that data from our PRRSV study are applicable to other pathogens; additional pathogen-specific studies are required.

PCNA promotes PRRSV replication by increasing the synthesis of viral genome

Porcine reproductive and respiratory syndrome virus (PRRSV) is an RNA virus belonging to the Arteriviridae family. Currently, the strain has undergone numerous mutations, bringing massive losses to the swine industry worldwide. Despite several studies had been conducted on PRRSV, the molecular mechanisms by which it causes infection remain unclear. Proliferating cell nuclear antigen (PCNA) is a sign of DNA damage and it participates in DNA replication and repair. Therefore, in this study, we investigated the potential role of PCNA in PRRSV infection. We observed that PCNA expression was stable after PRRSV infection in vitro; however, PCNA was translocated from the nucleus to the cytoplasm. Notably, we found the redistribution of PCNA from the nucleus to the cytoplasm in cells transfected with the N protein. PCNA silencing inhibited PRRSV replication and the synthesis of PRRSV shorter subgenomic RNA (sgmRNA) and genomic RNA (gRNA), while PCNA overexpression promoted virus replication and PRRSV shorter sgmRNA and gRNA synthesis. By performing immunoprecipitation and immunofluorescence colocalization, we confirmed that PCNA interacted with replication-related proteins, namely NSP9, NSP12, and N, but not with NSP10 and NSP11. Domain III of the N protein (41–72 aa) interacted with the IDCL domain of PCNA (118–135 aa). Therefore, we propose cytoplasmic transport of PCNA and its subsequent influence on PRRSV RNA synthesis could be a viral strategy for manipulating cell function, thus PCNA is a potential target to prevent and control PRRSV infection.

Evaluating oral swab samples for PRRSV surveillance in weaning-age pigs under field conditions.

The use of serum and family oral fluids for porcine reproductive and respiratory syndrome virus (PRRSV) surveillance in weaning-age pigs has been previously characterized. Characterizing more sample types similarly offers veterinarians and producers additional validated sample options for PRRSV surveillance in this subpopulation of pigs. Oral swab sampling is relatively easy and convenient; however, there is sparse information on how it compares to the reference sample type for PRRSV surveillance under field conditions. Therefore, this study's objective was to compare the PRRSV reverse-transcription real-time polymerase chain reaction (RT-rtPCR) test outcomes of oral swabs (OS) and sera samples obtained from weaning-age pig litters. At an eligible breeding herd, six hundred twenty-three weaning-age piglets from 51 litters were each sampled for serum and OS and tested for PRRSV RNA by RT-rtPCR. PRRSV RT-rtPCR positivity rate was higher in serum samples (24 of 51 litters, 83 of 623 pigs, with a mean cycle threshold (Ct) value of RT-rtPCR-positive samples per litter ranging from 18.9 to 32.0) compared to OS samples (15 of 51 litters, 33 of 623 pigs, with a mean Ct of RT-rtPCR positive samples per litter ranging from 28.2 to 36.9); this highlights the importance of interpreting negative RT-rtPCR results from OS samples with caution. Every litter with a positive PRRSV RT-rtPCR OS had at least one viremic piglet, highlighting the authenticity of positive PRRSV RT-rtPCR tests using OS; in other words, there was no evidence of environmental PRRSV RNA being detected in OS. Cohen's kappa analysis (Ck = 0.638) indicated a substantial agreement between both sample types for identifying the true PRRSV status of weaning-age pigs.

Brazilin from Caesalpinia sappan inhibits viral infection against PRRSV via CD163ΔSRCR5 MARC-145 cells: an in silico and in vitro studies

This research aimed to identify bioactive compounds from Caesalpinia sappan extract that function as novel porcine reproductive and respiratory syndrome virus (PRRSV) infection inhibitors by computational molecular screening. We obtained a set of small-molecule compounds predicted to target the scavenger receptor cysteine-rich domain 5 (SRCR5) of CD163. In addition, the functions of positive hits were assessed and verified utilizing an in vitro antiviral activity assay with PRRSV-infected MARC-145 cells. Combining molecular docking with the results of binding affinity and ligand conformation, it was found that brazilin had the highest binding energy with the SRCR5 receptor compared to catechin and epicatechin (− 5.8, − 5.5, and − 5.1 kcal/mol, respectively). In terms of molecular mechanics, the binding free energy between the SRCR5 receptor was − 15.71 kcal/mol based on the Poisson-Boltzmann surface area of brazilin. In addition, PRRSV infection in MARC-145 cells was significantly inhibited by brazilin compared to the control (virus titer, 4.10 vs. 9.25 TCID50/mL, respectively). Moreover, brazilin successfully limited the number of PRRSV RNA copies in MARC-145 cells as determined by RT-qPCR. By inhibiting the PRRSV-CD163 interaction with brazilin from Caesalpinia sappan, it may be possible to prevent PRRSV infection in pigs, as suggested by this research.

Porcine reproductive and respiratory syndrome virus RNA detection in tongue tips from dead animals.

The control of porcine reproductive and respiratory syndrome virus (PRRSV) hinges on monitoring and surveillance. The objective of this study was to assess PRRSV RNA detection by RT-PCR in tongue tips from dead suckling piglets compared to serum samples, processing fluids, and family oral fluids. Tongue tips and serum samples were collected from three PRRSV-positive breeding herd farms (farms A, B, and C) of three different age groups: newborns (<24 h), processing (2 to 7 days of age), and weaning (18 to 22 days of age). Additionally, processing fluids and family oral fluids were collected from 2–7 days of age and weaning age, respectively. In farms A and B, PRRSV RNA was detected in tongue tips from all age groups (100 and 95%, respectively). In addition, PRRSV RNA was detected in pooled serum samples (42 and 27%), processing fluids (100 and 50%), and family oral fluids (11 and 22%). Interestingly, the average Ct value from tongue tips was numerically lower than the average Ct value from serum samples in the newborn age. In farm C, PRRSV RNA was only detected in serum samples (60%) and family oral fluids (43%), both from the weaning age. Further, no PRRSV RNA was detected in tongue tips when pooled serum samples from the same age group tested PRRSV RNA-negative. Taken together, these results demonstrate the potential value of tongue tips for PRRSV monitoring and surveillance.

Considerations in the use of processing fluids for the detection of PRRSV RNA and antibody.

Surveillance is mandatory for tracking the progress of porcine reproductive and respiratory syndrome virus (PRRSV) control and elimination efforts in breeding herds. Processing fluids, the fluid recovered from tissues collected at castration and/or tail docking, are used for breeding herd surveillance by large segments of the industry, but the basic diagnostic characteristics of processing fluids are largely undescribed. We undertook 3 studies to address this information gap. In study 1, we found no differences among the PRRSV RT-rtPCR results obtained with 4 commercial RNA extraction kits. In study 2, we found that PRRSV RNA was highly stable in processing fluid samples at -20°C or 4°C, but detrimental effects were observed at ≥22°C within 24 h. In study 3, using a modified PRRSV ELISA at a sample:positive cutoff of ≥0.5, we found excellent discrimination in the detection of PRRSV antibody (IgM, IgA, IgG) in processing fluids from herds of known PRRSV status. Judicious handling of processing fluid samples from sow herds, and the use of methods available in veterinary diagnostic laboratories, can provide a foundation for reliable PRRSV surveillance.

Effect of pooling family oral fluids on the probability of PRRSV RNA detection by RT-rtPCR

Family oral fluids (FOFs) are an aggregate sample type shown to be a cost-efficient and convenient option for determining the porcine reproductive and respiratory syndrome virus (PRRSV) status of weaning age pigs. This study investigates the effect of pooling PRRSV-positive FOF samples with PRRSV-negative FOF samples at different levels (1/3, 1/5, 1/10, 1/20) on the probability of PRRSV RNA detection by reverse-transcription real-time polymerase chain reaction (RT-rtPCR). Mathematical models were built to assess how much the probability of RT-rtPCR PRRSV detection changed with increasing proportion of PRRSV-positive samples present within pools and how partially sampling a farrowing room influenced the probability of RT-rtPCR detection of PRRSV RNA in pooled samples at different prevalence scenarios. A general example of a guideline for FOF-based sampling under different prevalence scenarios to detect PRRSV RNA by RT-rtPCR with at least 95 % certainty is presented. At the sample level, the probability of detecting PRRSV RNA by RT-rtPCR decreased from 100 % to 87 %, 68 %, and 26 % when diluting up to 1/20 for PRRSV positive FOF having an initial Cycle threshold (Ct) below 34, between 34 and 36, or above 36, respectively. When PRRSV prevalence is near-zero (1 or 2 litters positive out of 56), the most cost-efficient farrowing room sampling strategy to detect PRRSV RNA with at least 95 % certainty was pooling FOF samples up to 1/10; at higher prevalence (≥ 3 of 56 litters positive), the most cost-efficient strategy was submitting samples in pools of 20. Subsampling a farrowing room for FOF pools was also demonstrated to be a valuable cost-saving strategy. Overall, based on the conditions of this study, pooling FOFs up to 1/20 is a valid option in situations of cost constraint and regardless of pooling level chosen, capturing as many litters as possible improves the probability of PRRSV detection.

The efficacy of a modified live virus vaccine Fostera® PRRS against heterologous strains of porcine reproductive and respiratory syndrome virus: A meta-analysis

Porcine reproductive and respiratory syndrome virus (PRRSV) is currently one of the most economically important health challenges in the global swine industry. The primary aim of this study was to evaluate the overall efficacy of a modified live virus vaccine Fostera® PRRS (F-PRRS) compared to no vaccination as reported in published studies, using meta-analytic techniques. Additionally, we aimed to evaluate the potential impact of age at vaccination and F-PRRS cross-protection against different genetically distanced PRRS strains. In total, 20 papers fulfilled the predefined inclusion criteria. Vaccinated pigs had on average 38.52 g/d higher daily weight gain and a 65% lower mortality (relative risk = 0.35) compared to non-vaccinates. F-PRRS reduced the maximum macroscopic lung lesion score on average by 16.82% points and the maximum viral load in serum by 1.36 log10 PRRSV RNA copies. Vaccination at 1 day and 21 days of age was similarly effective, and the pathogenic PRRS strain(s) used for challenge or being endemic in field studies (PRRSV-1, PRRSV-2, or PRRSV-1 & -2) did not significantly influence the outcomes. Our findings confirm the effectiveness of F-PRRS against heterologous PRRSV infection.

Virological and Histopathological Findings in Boars Naturally Infected With Porcine Reproductive and Respiratory Syndrome Virus Type 1.

Major geographical transmission of porcine reproductive and respiratory syndrome virus (PRRSV) occurs via semen when a boar stud is infected. This happened in Denmark in 2019, providing an opportunity to compare previous experimental PRRSV boar studies with natural PRRSV-1 infection in boars. The aim of this study was to investigate the association between the presence of PRRSV RNA in serum, semen, testicles, and epididymis of boars naturally infected with PRRSV and to describe the histological lesions in the testes and epididymis combined with direct visualisation of PRRSV-infected cells by immunohistochemical staining (IHC). The exact timing of infection of each boar was not determined, but based on serology the boars were divided into two groups: acute and late infections. All boars included were sampled the same day. In this study, 35 boars and 10 healthy boars from another PRRSV-negative boar stud were included as histological controls. PRRSV RNA was found most often in serum (51%) and least frequently in semen (22%) and was more often detected in the reproductive tract in the acute phase of infection (p < 0.0001; RR: 2.58). Mononuclear cells and multinuclear giant cells were present in the adluminal compartment of the testis and epididymis in PRRSV-infected boars, but not in control boars (p < 0.05), which supports the hypothesis that macrophages are involved in the venereal spread of the virus.

Phenotypic Characterization of a Virulent PRRSV-1 Isolate in a Reproductive Model With and Without Prior Heterologous Modified Live PRRSV-1 Vaccination.

Reproductive disorders induced by porcine reproductive and respiratory syndrome virus (PRRSV) cause high economic losses in the pig industry worldwide. In this study, we aimed to phenotypically characterize a virulent PRRSV-1 subtype 1 isolate (AUT15-33) in a reproductive model. Furthermore, the protective effect of a heterologous modified live virus vaccine (ReproCyc® PRRS EU) was evaluated. In addition, PRRSV AUT15-33 was genotypically compared to other well-characterized isolates. Sixteen gilts were equally divided into four groups: a vaccinated and infected group (V–I), a vaccinated and non-infected group (V–NI), a non-vaccinated and infected group (NV–I), and a non-vaccinated and non-infected (NV–NI) group. After PRRSV infection on gestation day 84, all gilts were clinically examined on a daily basis, and blood samples were taken at five timepoints. Necropsy was performed 3 weeks after infection. The fetal preservation status was assessed, and PRRSV RNA concentrations were measured in the blood and tissue samples from all gilts and fetuses. After infection, all four gilts in the NV–I group were viremic throughout 17 days post-infection (dpi), whereas two gilts in the V–I group were viremic at only one timepoint at 6 dpi. The viral load was significantly higher in gilt serum, tracheobronchial lymph nodes, uterine lymph nodes, maternal endometrium, and fetal placenta of NV–I gilts compared to the V–I ones (p < 0.05). Moreover, the preservation status of the fetuses derived from NV–I gilts was significantly impaired (55.9% of viable fetuses) compared to the other groups (p < 0.001). Upon comparison with other known isolates, the phylogenetic analyses revealed the closest relation to a well-characterized PRRSV-1 subtype 1 field isolate from Belgium. In conclusion, the high virulence of AUT15-33 was phenotypically confirmed in an experimental reproductive model. The vaccination of the gilts showed promising results in reducing viremia, fetal damage, and transplacental transmission of the PRRSV-1 strain characterized in this study.

Intranasal delivery of inactivated PRRSV loaded cationic nanoparticles coupled with enterotoxin subunit B induces PRRSV-specific immune responses in pigs

This study was conducted to evaluate the induction of systemic and mucosal immune responses and protective efficacy following the intranasal administration of inactivated porcine reproductive and respiratory syndrome virus (PRRSV) loaded in polylactic acid (PLA) nanoparticles coupled with heat-labile enterotoxin subunit B (LTB) and dimethyldioctadecylammonium bromide (DDA). Here, 42- to 3-week-old PRRSV-free pigs were randomly allocated into 7 groups of 6 pigs each. Two groups represented the negative (nonvaccinated pigs/nonchallenged pigs, NoVacNoChal) and challenge (nonvaccinated/challenged, NoVacChal) controls. The pigs in the other 5 groups, namely, PLA nanoparticles/challenged (blank NPs), LTB-DDA coupled with PLA nanoparticles/challenged (adjuvant-blank NPs), PLA nanoparticles-encapsulating inactivated PRRSV/challenged (KNPs), LTB-DDA coupled with PLA nanoparticles loaded with inactivated PRRSV/challenged pigs (adjuvant-KNPs) and inactivated PRRSV/challenged pigs (inactivated PRRSV), were intranasally vaccinated with previously described vaccines at 0, 7 and 14 days post-vaccination (DPV). Serum and nasal swab samples were collected weekly and assayed by ELISA to detect the presence of IgG and IgA, respectively. Viral neutralizing titer (VNT) in sera, IFN-γ-producing cells and IL-10 secretion in stimulated peripheral blood mononuclear cells (PBMCs) were also measured. The pigs were intranasally challenged with PRRSV-2 at 28 DPV and necropsied at 35 DPV, and then macro- and microscopic lung lesions were evaluated. The results demonstrated that following vaccination, adjuvant-KNP-vaccinated pigs had significantly higher levels of IFN-γ-producing cells, VNT and IgG in sera, and IgA in nasal swab samples and significantly lower IL-10 levels than the other vaccinated groups. Following challenge, the adjuvant-KNP-vaccinated pigs had significantly lower PRRSV RNA and macro- and microscopic lung lesions than the other vaccinated groups. In conclusion, the results of the study demonstrated that adjuvant-KNPs are effective in eliciting immune responses against PRRSV and protecting against PRRSV infections over KNPs and inactivated PRRSV and can be used as an adjuvant for intranasal PRRSV vaccines.

A serological survey of porcine reproductive and respiratory syndrome virus in wild boar in Gifu Prefecture, Japan.

Porcine reproductive and respiratory syndrome (PRRS) is an infectious swine disease caused by the PRRS virus (PRRSV) that results in economic loss to the pig-rearing industry. To study PRRSV infection in wild boars and pigs, we conducted a serological survey in Gifu Prefecture, Japan, from 2020 to 2021. Three out of 453 (0.7%) wild boar sera were positive for PRRSV antibodies in a commercial ELISA. However, given that PRRSV RNA was not detected in these three wild boars and the specificity and sensitivity of the test kit, these are considered as false positives. Although seropositive pigs were found in multiple pig farms in the study area, the role of wild boars as a source of PRRS to pig farms appeared to be minimal.

Probability of PRRS virus detection in pooled processing fluid samples

There has been a tremendous increase in recent years of population-based diagnostic monitoring and surveillance strategies in swine populations. One example is the use of processing fluids (PF) to screen breeding herds for porcine reproductive and respiratory syndrome virus (PRRSV) activity. An important question from practitioners using such methods is on how intensively can the sample be pooled. More specifically, processing fluids of how many litters can be pooled into a single sample for diagnostic testing to preserve a high probability of PRRSV RNA detection at low prevalence situations? The objective of this study was to model the effect of pooling PF samples on the probability of PRRSV RNA detection. For this study, a PRRSV-positive PF field sample with a RT-rtPCR quantification cycle (Cq) value of 28 was selected to represent a litter of 11 pigs with a single viremic piglet. PF samples from a PRRSV-naïve herd were used to perform 6 replications of 8 two-fold serial dilutions of the PRRSV-positive sample, thus modeling the pooling effect (dilution). Each two-fold dilution represented an increase in the number of PRRS-negative pigs in the sample by a factor of 2. Samples were tested for PRRSV RNA by RT-rtPCR and the data was analyzed using linear and probit regression models. There was an average increment of 1.37 points in Ct for each two-fold dilution. The estimated probability of testing positive on RT-rtPCR was 43 %, 80 %, and 95 % when there was a single PRRSv-positive piglet among 784, 492, and 323 PRRSv-negative piglets contributing to the sample respectively. Results from this study support the practice of collecting and aggregating PF samples from multiple litters for PRRSV RNA testing.