PRRSV-1 Strains Research Articles

Sequence analysis of the GP5 protein of porcine reproductive and respiratory syndrome virus in Vietnam from 2007 to 2023.

Porcine reproductive and respiratory syndrome virus (PRRSV) is the causative agent 13 of porcine reproductive and respiratory syndrome (PRRS), which is one of the most economically 14 devastating viruses in the Vietnamese swine industry. With a view toward determining the 15 genetic variation among PRRSV strains in Vietnam, we examined 271 PRRSV GP5 protein 16 sequences obtained from strains isolated in Vietnam from 2007 to 2023, for which we constructed 17 phylogenetic trees. Additionally, a collection of 52 PRRSV-1 strains and 80 PRRSV-2 strains 18 isolated in different years were specifically selected for nucleotide and amino acid homology analysis 19 and amino acid sequence alignment. The results revealed 76.1%-100.0% nucleotide and 20 75.2%-100.0% amino acid homologies for the PRRSV-1 GP5 gene, and 81.8%-100.0% nucleotide 21 and 81.1%-100.0% amino acid homologies for the PRRSV-2 GP5 gene. Amino acid mutation sites 22 in PRRSV-2 were found to be primarily distributed in the signal peptide region, antigenic sites, two 23 T-cell antigen regions, two highly variable regions (HVRs), and in the vicinity of the neutralizing 24 epitope, with a deletion mutation occurring in the neutralizing epitope, whereas amino acid mutations 25 in the PRRSV-1 sequences were found to occur predominantly in two T-cell epitopes. Genetic 26 analysis revealed that PRRSV-1 strains in Vietnam are of subtype 1 (Global), whereas PRRSV-2 27 strains are categorized into sublineages L1A, L5A, and L8E, with L8E being the predominantly 28 prevalent strain at present. Recombination analyses indicated that no significant recombination 29 events have occurred in any of the assessed 271 Vietnamese PRRSV strains. Our 30 analyses of 271 Vietnamese PRRSV strains have yielded valuable insights regarding the 31 epidemiological trends and genetic dynamics of PRRSV in Vietnam, and will provide a theoretical 32 basis for formulating prevention and control measures for PRRS and the development of PRRS 33 vaccines.

Investigation of the distribution and origin of porcine reproductive and respiratory syndrome virus 1 in the swine production chain: A retrospective study of three farms in Thailand.

Porcine reproductive and respiratory syndrome (PRRS), caused by PRRS virus (PRRSV), is a global issue that affects Thai swine as well. In Thailand, PRRSV-2 predominates over PRRSV-1. The origin of PRRSV-1 transmission remains undiscovered. This study traced the source of infected pigs responsible for disease transmission among three pig-fattening farms and analyzed the spread of PRRSV-1. A total of 696 swine samples from breeding and pig-fattening farms in Thailand were screened for PRRSV using open reading frames (ORF7) reverse transcription polymerase chain reaction (RT-PCR). Positive samples were identified as PRRSV-1 using ORF5 RT-PCR. The analysis included the study of nucleotide homology, GP5 amino acid sequences, and N-linked glycosylation patterns to assess the spread of PRRSV-1 across these farms. Genetic examination identified 28 PRRSV-1-positive samples, of which 13 were chosen as representatives. These strains were categorized into three groups based on breeding farm pig houses and showed distinct distribution patterns across pig-fattening farms. Group 1 included piglets transferred from pig house A to Nakhon Pathom, Chonburi, and Sa Kaeo. Groups 2 and 3 showed transfers from pig houses F and H to Chonburi and Sa Kaeo farms. All 13 PRRSV-1 strains were categorized into PRRSV-1 subtype 1/clade H. N-linked glycosylation analysis revealed that nearly all PRRSV-1 strains exhibited a conserved glycosylation pattern at amino acid positions N37, N46, and N53. This pattern is consistent with the glycosylation profile of the previous Thai PRRSV-1 subtype 1/clade H. The present study highlights the persistent presence of PRRSV-1 in Thai swine, which leads to sporadic outbreaks. The molecular genetic analysis identified three primary strain groups dispersed throughout the pig production system, emphasizing the importance of regular monitoring for new PRRSV strains in this herd. Understanding the PRRSV-1 distribution in swine farms is vital for veterinarians. This knowledge supports strategies for eradicating the virus and managing swine health effectively in Thailand.

Characterization and Pathogenicity of the Novel Porcine Reproductive and Respiratory Syndrome Virus 1 Strain SL-01 in China

Currently, PRRSV-1 causes a large number of clinical infections in Chinese swine herds, and the prevalence of new strains has presented great challenges. In this study, the novel PRRSV-1 strain SL-01 was isolated, with a genome length of 14,978 bp, and genetic evolution analysis revealed that it belonged to a new subtype branch. Sequence homology analysis showed that the strain was only 82.2%–86.7% identical to the current classical PRRSV-1 strains. In particular, the novel strain exhibited a unique deletion pattern in Nsp2. In addition, GP3 and GP4 of the SL-01 strain showed four consecutive amino acid deletions in the highly variable regions at amino acids 243–248 and 63–68, respectively. Further challenges in piglet and pregnant sow demonstrated that the SL-01 strain could cause the piglet fever and death but less pathogenic to pregnant sow. Overall, the characterization and pathogenicity of a novel PRRSV-1 strain were first explored and provide a prevention for pig farms.

Mass Spectrometry Imaging (MALDI-MSI) as a molecular histology application for characterizing pulmonary lesions caused by PRRSV-1 strains of different virulence

Mass Spectrometry Imaging (MALDI-MSI) as a molecular histology application for characterizing pulmonary lesions caused by PRRSV-1 strains of different virulence

The scene of lung pathology during PRRSV-1 infection.

Porcine reproductive and respiratory syndrome (PRRS) is one of the most economically important infectious diseases for the pig industry worldwide. The disease was firstly reported in 1987 and became endemic in many countries. Since then, outbreaks caused by strains of high virulence have been reported several times in Asia, America and Europe. Interstitial pneumonia, microscopically characterised by thickened alveolar septa, is the hallmark lesion of PRRS. However, suppurative bronchopneumonia and proliferative and necrotising pneumonia are also observed, particularly when a virulent strain is involved. This raises the question of whether the infection by certain strains results in an overstimulation of the proinflammatory response and whether there is some degree of correlation between the strain involved and a particular pattern of lung injury. Thus, it is of interest to know how the inflammatory response is modulated in these cases due to the interplay between virus and host factors. This review provides an overview of the macroscopic, microscopic, and molecular pathology of PRRSV-1 strains in the lung, emphasising the differences between strains of different virulence.

Diltiazem HCl suppresses porcine reproductive and respiratory syndrome virus infection in susceptible cells and in swine

Porcine reproductive and respiratory syndrome virus (PRRSV) is a pathogen for swine, resulting in substantial economic losses to the swine industry. However, there has been little success in developing effective vaccines or drugs for PRRSV control. In the present study, we discovered that Diltiazem HCl, an inhibitor of L-type Ca2+ channel, effectively suppresses PRRSV replication in MARC-145, PK-15CD163 and PAM cells in dose-dependent manner. Furthermore, it demonstrates a broad-spectrum activity against both PRRSV-1 and PRRSV-2 strains. Additionally, we explored the underlying mechanisms and found that Diltiazem HCl -induced inhibition of PRRSV associated with regulation of calcium ion homeostasis in susceptible cells. Moreover, we evaluated the antiviral effects of Diltiazem HCl in PRRSV-challenged piglets, assessing rectal temperature, viremia, and gross and microscopic lung lesions. Our results indicate that Diltiazem HCl treatment alleviates PRRSV-induced rectal temperature spikes, pulmonary pathological changes, and serum viral load. In conclusion, our data suggest that Diltiazem HCl could serve as a novel therapeutic drug against PRRSV infection.

Genetic variation and recombination analysis of the GP5 gene of the porcine reproductive and respiratory syndrome virus in Thailand.

Porcine reproductive and respiratory syndrome (PRRS) is a significant threat to the global swine industry, and its prevalence in Thailand spans over two decades. To understand the genetic variation and recombination of the PRRS virus (PRRSV) GP5 gene in Thailand, we retrieved 726 GP5 gene sequences from the NCBI database. Phylogenetic trees were constructed using the neighbor-joining (NJ) and maximum likelihood (ML) methods, and recombination analysis was performed. Homology analysis was conducted on 83 PRRSV-1 and 83 PRRSV-2 strains. Phylogenetic analysis revealed the prevalence of both PRRSV-1 and PRRSV-2 strains in Thailand, with the latter exhibiting wider distribution. PRRSV-1 strains clustered into clades A, D, and H, while PRRSV-2 strains grouped into lineages 1, 5, and sublineage 8.7, further divided into 8.7/HP and 8.7/NA sublineages. Sublineage 8.7/NA strains accounted for a significant proportion of circulating PRRSV-2 strains. Homology analysis showed nucleotide and amino acid similarities ranging from 75.4 to 100.0% and 41.3 to 100.0% for PRRSV-1, and 78.6 to 100.0% and 70.8 to 100.0% for PRRSV-2 strains. Amino acid sequence alignments revealed mutations, insertions, and deletions in PRRSV-1 GP5, and key residue mutations in PRRSV-2 GP5 associated with biological functions. Recombination analysis identified two recombination events within PRRSV-2 sublineage 8.7 strains. These findings confirm the variability of the GP5 protein. This study enhances our understanding of PRRSV prevalence and genetic variation in Thailand, contributing valuable insights for PRRS prevention and control.

Levels of neutralizing antibodies against resident farm strain or vaccine strain are not indicators of protection against PRRSV-1 vertical transmission under farm conditions

BackgroundVertical transmission is key for the maintenance of porcine reproductive and respiratory syndrome virus (PRRSV) infection. In vaccinated farms, vertical transmission can still occur despite sows having some level of immunity because of repeated vaccination or contact with the wild-type virus. The present study aimed to correlate the age of sows and the amplitude of neutralizing antibodies (Nab) (heterologous neutralization) with PRRSV-1 vertical transmission (VT). For this purpose, umbilical cords of 1,554 newborns (corresponding to 250 litters) were tested for PRRSV by RT-PCR in two PRRSV-unstable vaccinated farms. In parallel, the sows were bled after farrowing and the levels of antibodies were determined by ELISA and by the viral neutralization test against the vaccine virus, the virus circulating in the farm, and other unrelated contemporary PRRSV-1 strains. The relationship between the parity and the probability of delivering infected piglets and the presence of broadly Nabs examined.ResultsThe proportion of VT events in the two examined farms ranged from 18.9% to 23.0%. Young sows (parity 1–2) were 1.7 times more likely to have VT than older sows (p < 0.05). Despite higher ELISA S/P antibody ratios in younger sows (p < 0.05), NAb against the resident farm strain were at a similar level between sows delivering infected and healthy piglets regardless of age, mostly with low titers (2–3 log2). The titers of NAb against the vaccine virus were also low, and no correlations with VT were observed. When a panel of another 4 strains (1 isolated in the 1990s, and 3 contemporary strains) were used for the neutralization test, most sow sera were not capable of neutralizing the contemporary strains.ConclusionsTiters of NAb could not be correlated with the occurrence of PRRSV VT. The amplitude of NAb present in most vaccinated sows is limited with a considerable proportion unresponsive regarding NAb production.

An attenuated herpesvirus vectored vaccine candidate induces T-cell responses against highly conserved porcine reproductive and respiratory syndrome virus M and NSP5 proteins that are unable to control infection.

Porcine reproductive and respiratory syndrome virus (PRRSV) remains a leading cause of economic loss in pig farming worldwide. Existing commercial vaccines, all based on modified live or inactivated PRRSV, fail to provide effective immunity against the highly diverse circulating strains of both PRRSV-1 and PRRSV-2. Therefore, there is an urgent need to develop more effective and broadly active PRRSV vaccines. In the absence of neutralizing antibodies, T cells are thought to play a central role in controlling PRRSV infection. Herpesvirus-based vectors are novel vaccine platforms capable of inducing high levels of T cells against encoded heterologous antigens. Therefore, the aim of this study was to assess the immunogenicity and efficacy of an attenuated herpesvirus-based vector (bovine herpesvirus-4; BoHV-4) expressing a fusion protein comprising two well-characterized PRRSV-1 T-cell antigens (M and NSP5). Prime-boost immunization of pigs with BoHV-4 expressing the M and NSP5 fusion protein (vector designated BoHV-4-M-NSP5) induced strong IFN-γ responses, as assessed by ELISpot assays of peripheral blood mononuclear cells (PBMC) stimulated with a pool of peptides representing PRRSV-1 M and NSP5. The responses were closely mirrored by spontaneous IFN-γ release from unstimulated cells, albeit at lower levels. A lower frequency of M and NSP5 specific IFN-γ responding cells was induced following a single dose of BoHV-4-M-NSP5 vector. Restimulation using M and NSP5 peptides from PRRSV-2 demonstrated a high level of cross-reactivity. Vaccination with BoHV-4-M-NSP5 did not affect viral loads in either the blood or lungs following challenge with the two heterologous PRRSV-1 strains. However, the BoHV-4-M-NSP5 prime-boost vaccination showed a marked trend toward reduced lung pathology following PRRSV-1 challenge. The limited effect of T cells on PRRSV-1 viral load was further examined by analyzing local and circulating T-cell responses using intracellular cytokine staining and proliferation assays. The results from this study suggest that vaccine-primed T-cell responses may have helped in the control of PRRSV-1 associated tissue damage, but had a minimal, if any, effect on controlling PRRSV-1 viral loads. Together, these results indicate that future efforts to develop effective PRRSV vaccines should focus on achieving a balanced T-cell and antibody response.

Pathogenicity characterization of PRRSV-1 181187-2 isolated in China

PRRSV-1 has caused more clinical infections in pigs in Chinese swine herds in recent years, however, the pathogenicity of PRRSV-1 in China is unclear. In order to study the pathogenicity of PRRSV-1, in this study, a PRRSV-1 strain, 181187-2, was isolated in primary alveolar macrophage (PAM) cells from a farm where abortions had been reported in China. The complete genome of 181187-2 was 14932 bp excluding Poly A, with 54-amino acid continuous deletion in the Nsp2 gene and 1 amino deletion in ORF3 gene compared with LV. Additionally, the piglets inoculated with strain 181187-2 by intranasal and intranasal plus intramuscular injection, animal experiments showed clinical symptoms including transient fever and depression, with no death. The obvious histopathological lesions including interstitial pneumonia and lymph node hemorrhage, and there were no significant differences in clinical symptoms and histopathological lesions with different challenge ways. Our results indicated that PRRSV -1 181187-2 was a moderately pathogenic strain in piglets.

Genomic Characterization and Pathogenicity of BJEU06-1-Like PRRSV-1 ZD-1 Isolated in China

Porcine reproductive and respiratory syndrome virus (PRRSV)-1 and PRRSV-2 have long been cocirculating in China. To date, all PRRSV-1 strains in China have been classified as subtype 1. We investigated the prevalence of PRRSV-1 in several areas of China from 2016 to 2022 and found that BJEU06-1-like strains comprised the main epidemic branch of PRRSV-1. Pathogenicity data for this subgroup are currently lacking. In this study, the Chinese BJEU06-1-like PRRSV-1 strain ZD-1 was isolated from primary alveolar macrophages (PAMs). ZD-1 has undergone no recombination and has a 5-aa discontinuous deletion in the Nsp2 protein, similar to other BJEU06-1-like strains; additionally, ZD-1 has a 26 aa C-terminal truncation in the GP3 gene. Pathogenicity studies revealed that ZD-1 causes obvious clinical symptoms: prolonged fever; reduced body weight; alveolar epithelial proliferation and moderate alveolar diaphragm widening in the lungs; diffuse lymphocytic hyperplasia in the lymph nodes; high levels of viremia in the serum; and elevated viral loads in the lungs, lymph nodes, and tonsils. These results suggested that the BJEU06-1-like PRRSV-1 strain ZD-1 is moderately pathogenic to piglets. This is the first study to evaluate the pathogenicity of the BJEU06-1-like branch in China, enriching the understanding of PRRSV-1 in China.

Quantitative Analysis of Inflammatory Uterine Lesions of Pregnant Gilts with Digital Image Analysis Following Experimental PRRSV-1 Infection.

Reproductive disorders caused by porcine reproductive and respiratory syndrome virus-1 are not yet fully characterized. We report QuPath-based digital image analysis to count inflammatory cells in 141 routinely, and 35 CD163 immunohistochemically stained endometrial slides of vaccinated or unvaccinated pregnant gilts inoculated with a high or low virulent PRRSV-1 strain. To illustrate the superior statistical feasibility of the numerical data determined by digital cell counting, we defined the association between the number of these cells and endometrial, placental, and fetal features. There was strong concordance between the two manual scorers. Distributions of total cell counts and endometrial and placental qPCR results differed significantly between examiner1's endometritis grades. Total counts' distribution differed significantly between groups, except for the two unvaccinated. Higher vasculitis scores were associated with higher endometritis scores, and higher total cell counts were expected with high vasculitis/endometritis scores. Cell number thresholds of endometritis grades were determined. A significant correlation between fetal weights and total counts was shown in unvaccinated groups, and a significant positive correlation was found between these counts and endometrial qPCR results. We revealed significant negative correlations between CD163+ counts and qPCR results of the unvaccinated group infected with the highly virulent strain. Digital image analysis was efficiently applied to assess endometrial inflammation objectively.

PRRSV-1 induced lung lesion is associated with an imbalance between costimulatory and coinhibitory immune checkpoints.

Porcine reproductive and respiratory syndrome virus (PRRSV) induces a dysregulation on the innate and adaptive immune responses. T-cell activation requires a proper interaction and precise balance between costimulatory and coinhibitory molecules, commonly known as immune checkpoints. This study aims to evaluate the expression of immune checkpoints in lung and tracheobronchial lymph node from piglets infected with two PRRSV-1 strains of different virulence during the early stage of infection. Seventy 4-week-old piglets were grouped into three experimental groups: (i) control, (ii) 3249-infected group (low virulent strain), and (iii) Lena-infected group (virulent strain) and were euthanized at 1, 3, 6, 8, and 13 days post-infection (dpi). Lung and tracheobronchial lymph node were collected to evaluate histopathological findings, PRRSV viral load and mRNA expression of costimulatory (CD28, CD226, TNFRSF9, SELL, ICOS, and CD40) and coinhibitory (CTLA4, TIGIT, PD1/PDL1, TIM3, LAG3, and IDO1) molecules through RT-qPCR. Our findings highlight a mild increase of costimulatory molecules together with an earlier and stronger up-regulation of coinhibitory molecules in both organs from PRRSV-1-infected animals, especially in the lung from virulent Lena-infected animals. The simultaneous expression of coinhibitory immune checkpoints could work in synergy to control and limit the inflammation-induced tissue damage. Further studies should be addressed to determine the role of these molecules in later stages of PRRSV infection.

Introduction of a PRRSV-1 strain of increased virulence in a pig production structure in Spain: virus evolution and impact on production

BackgroundA strain of Porcine reproductive and respiratory syndrome virus (PRRSV), showing characteristics of enhanced virulence, affected a pyramidal production system from Spain with 7600 sows in 4 genetic nuclei and 13,000 sows in multipliers. Different PRRSV strains circulating in this production system from December 2020 to October 2021 were detected and sequenced. The spread of each isolate was examined and their impact on health and production in three of the affected farms was evaluated.ResultsThe newly emerged PRRSV isolate with enhanced virulence entered the system before the onset of the study (January 2020) and afterwards four significantly different clades were detected during the study period in different farms, probably because of independent introduction events. The diversification of the enhanced virulence strain was higher for those clades (substitution rates up to 1.1% nucleotides/year) compared to other PRRSV strains present in the production system (up to 0.17%), suggesting a faster spread and adaptation. The impact of the infection in the first affected farm was dramatic, with an average abortion rate above 27% during 17 weeks before returning to the baseline production. Fertile sow mortality reached 6.5% for 39 weeks. In two farms infected later by other clades of this enhanced virulence strain, the impact was less acute; despite the fact that for parameters such as the proportion of stillbirths or mummies, more than 10 months were needed to recover pre-outbreak values. In the examined nurseries, mortalities reached peaks between 28 and 50% and several months were needed to return to normality.ConclusionIntroduction of a PRRSV strain of enhanced virulence in a production system where several farms were previously positive for other PRRSV strains, resulted in a fast spread such as would be observed in naïve farms. The productive and health impact was very high taking several months to return to normality.

Predicting Antigenic Distance from Genetic Data for PRRSV-Type 1: Applications of Machine Learning.

The control of porcine reproductive and respiratory syndrome (PRRS) remains a significant challenge due to the genetic and antigenic variability of the causative virus (PRRSV). Predominantly, PRRSV management includes using vaccines and live virus inoculations to confer immunity against PRRSV on farms. While understanding cross-protection among strains is crucial for the continued success of these interventions, understanding how genetic diversity translates to antigenic diversity remains elusive. We developed machine learning algorithms to estimate antigenic distance in silico, based on genetic sequence data, and identify differences in specific amino acid sites associated with antigenic differences between viruses. First, we obtained antigenic distance estimates derived from serum neutralization assays cross-reacting PRRSV monospecific antisera with virus isolates from 27 PRRSV1 viruses circulating in Europe. Antigenic distances were weakly to moderately associated with ectodomain amino acid distance for open reading frames (ORFs) 2 to 4 (ρ < 0.2) and ORF5 (ρ = 0.3), respectively. Dividing the antigenic distance values at the median, we then categorized the sera-virus pairs into two levels: low and high antigenic distance (dissimilarity). In the machine learning models, we used amino acid distances in the ectodomains of ORFs 2 to 5 and site-wise amino acid differences between the viruses as potential predictors of antigenic dissimilarity. Using mixed-effect gradient boosting models, we estimated the antigenic distance (high versus low) between serum-virus pairs with an accuracy of 81% (95% confidence interval, 76 to 85%); sensitivity and specificity were 86% and 75%, respectively. We demonstrate that using sequence data we can estimate antigenic distance and potential cross-protection between PRRSV1 strains. IMPORTANCE Understanding cross-protection between cocirculating PRRSV1 strains is crucial to reducing losses associated with PRRS outbreaks on farms. While experimental studies to determine cross-protection are instrumental, these in vivo studies are not always practical or timely for the many cocirculating and emerging PRRSV strains. In this study, we demonstrate the ability to rapidly estimate potential immunologic cross-reaction between different PRRSV1 strains in silico using sequence data routinely collected by production systems. These models can provide fast turn-around information crucial for improving PRRS management decisions such as selecting vaccines/live virus inoculation to be used on farms and assessing the risk of outbreaks by emerging strains on farms previously exposed to certain PRRSV strains and vaccine development among others.

Molecular Characterization of the Nsp2 and ORF5s of PRRSV Strains in Sichuan China during 2012–2020

Porcine reproductive and respiratory syndrome virus (PRRSV) is an important pathogen that poses a serious threat to the global pig industry. Sichuan Province is one of the largest pig breeding provinces in China. There is a lack of reports on the continuous surveillance and systematic analysis of prevalent strains of PRRSV in Sichuan Province in recent years. To fill this gap, a total of 539 samples were collected from 13 breeding regions in Sichuan during 2012-2020. The detection result showed that the positive rate of PRRSV was 52.32% (282/539). The ORF5s and Nsp2 were obtained and further analyzed, with Chinese reference strains downloaded from the GenBank. Phylogenetic analysis showed that the PRRSV strains sequenced in this study belonged to PRRSV-1 and PRRSV-2 (lineage 1, 3, 5 and 8). In total, 168 PRRSV-2 strains were selected for ORF5 analyses, and these strains were classified into sub-lineage 8.7 (HP-PRRSV), sub-lineage 5.1 (classical PRRSV), sub-lineage 1.8 (NADC30-like), sub-lineage 1.5 (NADC34-like) and sub-lineage 3.5 (QYYZ-like), accounting for 60.71% (102/168), 11.31% (19/168), 18.45% (31/168), 2.97% (5/168) and 6.55% (11/168) of the total analyzed strains, respectively. The Nsp2 of identified PRRSV strains exhibited a nucleotide identity of 44.5-100%, and an amino acid identity of 46.82-100%. The ORF5 of the identified PRRSV strains exhibited a nucleotide identity of 81.3-100%, and an amino acid identity of 78.5-100%. A sequence analysis of ORF5 revealed that the mutation sites of GP5 were mainly concentrated in HVR1 and HVR2 and the virulence sites. In summary, the HP-PRRSV, NADC30-like PRRSV, Classic-PRRSV, QYYZ-like PRRSV, NADC34-like PRRSV and PRRSV-1 strains exist simultaneously in pigs in Sichuan. NADC30-like PRRSV was gradually becoming the most prevalent genotype currently in Sichuan province. This study suggested that PRRSV strains in Sichuan were undergoing genomic divergence.

Genomic Analysis of Porcine Reproductive and Respiratory Syndrome Virus 1 Revealed Extensive Recombination and Potential Introduction Events in China

Simple SummaryPorcine reproductive and respiratory syndrome, caused by the porcine reproductive and respiratory syndrome virus, is considered one of the most devastating swine diseases worldwide. Porcine reproductive and respiratory syndrome virus 1 was first isolated in China in 2006, and there have been few reports concerning its genetic characteristics in China. We hope to find out the regularity of genetic diversity, recombination, and evolution of the virus by analyzing all available genomic sequences during 1991–2018. We found that high-frequency recombination regions were concentrated in non-structural protein 2 and structural proteins 2 to 4 and extensive deletions in non-structural protein 2; phylogenetic analysis revealed four independent introductions in China. Our results suggest that attention should be paid to the prevention and control of porcine reproductive and respiratory syndrome virus 1 and the rational use of vaccine strains. These results will help us to understand the recombination of porcine reproductive and respiratory syndrome virus and strengthen viral inspection before mixing herds of swine to reduce the probability of novel recombinant variants. Moreover, our study might form the basis of monitoring and control measures to prevent the spread of this economically important virus.Porcine reproductive and respiratory syndrome (PRRS), caused by the PRRS virus (PRRSV), is considered one of the most devastating swine diseases worldwide. PRRSV-1 was first isolated in China in 2006. However, there were few reports concerning the genetic characteristics of PRRSV-1 in China. In this study, three PRRSV-1 strains (HL85, HeB3, and HeB47) were detected by a general RT-qPCR method from clinical samples in 2018. HeB47 was identified as a recombinant between the BJEU06-1 and CReSA228-like strains. To further analyze the recombination and deletion features of PRRSV-1, all the available 88 complete genome sequences (isolated in 19 countries) from 1991 to 2018 in GenBank were analyzed. The high-frequency recombination regions were concentrated in NSP2 and GP2 to GP4. More importantly, phylogenetic analysis of PRRSV-1 revealed four independent introductions in China. Therefore, it is necessary to strengthen the important monitoring of breeding pigs and pork products and epidemiological surveys on pig farms to prevent the further spread of PRRSV-1.

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.

Heterologous Challenge with PRRSV-1 MLV in Pregnant Vaccinated Gilts: Potential Risk on Health and Immunity of Piglets.

Simple SummaryModified live virus (MLV) vaccines are considered as the key component to control the Porcine Reproductive and Respiratory Syndrome Virus (PRRSV). The majority of pig farms apply the ‘mass’ vaccination strategy in breeding female animals. However, this PRRS MLV vaccination protocol involves the risk of inoculation of sows in the last stage of gestation, resulting in possible infection of the fetus as the virus can efficiently cross the placenta during the last period of pregnancy. Thus, we evaluated the ability of the vaccine virus to act as a pathogenic strain, to be transmitted to fetuses and to affect the health status of neonatal piglets. The results indicated that the study gilts transmitted the vaccine virus to their offspring, as well as that the PRRSV-infected piglets showed a poor clinical performance. Consequently, the pig farms that apply PRRS MLV vaccination in a routine blanket vaccination strategy must avoid inoculating pregnant gilts the last week before their parturition.The objective of the present study was to evaluate the potential risks of the four commercial PRRS-1 MLV vaccines in pregnant vaccinated gilts at the last stage of gestation under field conditions. The study was conducted at four pig farms, including 25 gilts from each farm (25 × 4 = 100 gilts), which were equally allocated to five different study groups. A PRRS-1 MLV vaccination was applied on the 100th day of their pregnancy with the different commercial vaccines that are available in the Greek market. The results indicated virus congenital infection and viremia in piglets (20/200 = 10% PRRSV infected piglets), and detection of PRRSV-specific antibodies (181/200 = 90.5% piglets found with PRRSV antibodies). The subsequent phylogenetic analyses revealed high percentages of similarity between the PRRSV-1 strain detected in infected litters and the PRRSV-1 vaccine strain to which the study gilts had been previously exposed to. Health status analyses of trial piglets resulted in differences between litters from vaccinated sows and litters from non-vaccinated sows at 110th day of gestation as regards the number of weak-born piglets, mummies, and piglets with splay-leg and/or respiratory symptoms. The current study’s results indicate several potential dangers of the PRRS MLV vaccination in late gestation.

Activation of T-bet, FOXP3, and EOMES in Target Organs From Piglets Infected With the Virulent PRRSV-1 Lena Strain.

Transcription factors (TFs) modulate genes involved in cell-type-specific proliferative and migratory properties, metabolic features, and effector functions. Porcine reproductive and respiratory syndrome virus (PRRSV) is one of the most important pathogen agents in the porcine industry; however, TFs have been poorly studied during the course of this disease. Therefore, we aimed to evaluate the expressions of the TFs T-bet, GATA3, FOXP3, and Eomesodermin (EOMES) in target organs (the lung, tracheobronchial lymph node, and thymus) and those of different effector cytokines (IFNG, TNFA, and IL10) and the Fas ligand (FASL) during the early phase of infection with PRRSV-1 strains of different virulence. Target organs from mock-, virulent Lena-, and low virulent 3249-infected animals humanely euthanized at 1, 3, 6, 8, and 13 days post-infection (dpi) were collected to analyze the PRRSV viral load, histopathological lesions, and relative quantification through reverse transcription quantitative PCR (RT-qPCR) of the TFs and cytokines. Animals belonging to both infected groups, but mainly those infected with the virulent Lena strain, showed upregulation of the TFs T-bet, EOMES, and FOXP3, together with an increase of the cytokine IFN-γ in target organs at the end of the study (approximately 2 weeks post-infection). These results are suggestive of a stronger polarization to Th1 cells and regulatory T cells (Tregs), but also CD4+ cytotoxic T lymphocytes (CTLs), effector CD8+ T cells, and γδT cells in virulent PRRSV-1-infected animals; however, their biological functionality should be the object of further studies.