miR-541-3p Promoted Porcine Reproductive and Respiratory Syndrome Virus 2 (PRRSV-2) Replication by Targeting Interferon Regulatory Factor 7

Porcine reproductive and respiratory syndrome (PRRS) is a disease caused by PRRS virus (PRRSV), which seriously harms the pig industry. Revealing the mechanism by which PRRSV inhibits immune response will help prevent and control PRRS. Here, we found that PRRSV-2 may hijack host miR-541-3p to inhibit host innate immune response. Firstly, this work showed that miR-541-3p mimics could facilitate the replication of PRRSV-2 and the results of the quantitative real time polymerase chain reaction (qRT-PCR) showed that PRRSV-2 could up-regulate the expression of miR-541-3p in MARC-145 cells. Since previous studies have shown that type I interferon could effectively inhibit the replication of PRRSV-2, the present work explored whether miR-541-3p regulated the expression of type I interferon and found that miR-541-3p could negatively regulate the transcription of type I interferon by targeting interferon regulatory factor 7 (IRF7). More importantly, PRRSV-2 infection could down-regulate the expression of IRF7 and over-expression of IRF7 could down-regulate the replication of PRRSV-2 in MARC-145 cells. In conclusion, PRRSV-2 infection up-regulated the expression of miR-541-3p to promote its replication in MARC-145 cells, since miR-541-3p can negatively regulate the transcription of type I interferon by targeting IRF7.

Evaluation of the Cross-Protective Efficacy of a Chimeric PRRSV Vaccine against Two Genetically Diverse PRRSV2 Field Strains in a Reproductive Model

Despite the routine use of porcine reproductive and respiratory syndrome (PRRS)-modified live vaccines, serious concerns are currently being raised due to their quick reversion to virulence and limited cross-protection against divergent PRRS virus (PRRSV) strains circulating in the field. Therefore, a PRRS chimeric vaccine (JB1) was produced using a DNA-launched infectious clone by replacing open reading frames (ORFs) 3–6 with those from a mixture of two genetically different PRRSV2 strains (K07–2273 and K08–1054) and ORF1a with that from a mutation-resistant PRRSV strain (RVRp22) exhibiting an attenuated phenotype. To evaluate the safety and cross-protective efficacy of JB1 in a reproductive model, eight PRRS-negative pregnant sows were purchased and divided into four groups. Four sows in two of the groups were vaccinated with JB1, and the other 4 sows were untreated at gestational day 60. At gestational day 93, one vaccinated group and one nonvaccinated group each were challenged with either K07–2273 or K08–1054. All of the sows aborted or delivered until gestation day 115 (24 days post challenge), and the newborn piglets were observed up to the 28th day after birth, which was the end of the experiment. Overall, pregnant sows of the JB1-vaccinated groups showed no meaningful viremia after vaccination and significant reductions in viremia with K07–2273 and K08–1054, exhibiting significantly higher levels of serum virus-neutralizing antibodies than non-vaccinated sows. Moreover, the JB1-vaccinated groups did not exhibit any abortion due to vaccination and showed improved piglet viability and birth weight. The piglets from JB1-vaccinated sows displayed lower viral concentrations in serum and fewer lung lesions compared with those of the piglets from the nonvaccinated sows. Therefore, JB1 is a safe and effective vaccine candidate that confers simultaneous protection against two genetically different PRRSV strains.

Characterization of Two Immunodominant Antigenic Peptides in NSP2 of PRRSV-2 and Generation of a Marker PRRSV Strain Based on the Peptides

Porcine reproductive and respiratory syndrome (PRRS) is a widespread disease with great economic importance in the pig industry. Although vaccines against the PRRS virus (PRRSV) have been employed for more than 20 years, differentiating infected from vaccinated animal remains challenging. In this study, all 907 non-structural protein 2 (NSP2) full-length sequences of PRRSV-2 available from GenBank were aligned. Two peptides, at positions 562–627 (m1B) and 749–813 (m2B) of NSP2, were selected, and their potential for use in differential diagnosis was assessed. Both m1B and m2B were recognized by PRRSV-positive pig serum in peptide-coated enzyme-linked immunosorbent assays. Further epitope identification yielded five overlapping short peptides for the immunodominant regions of m1B and m2B. Using the infectious clone of PRRSV HuN4-F112 as a template, the deletion mutants rHuN4-F112-m1B, rHuN4-F112-m2B, and rHuN4-F112-C5-m1B-m2B were generated and successfully rescued in Marc-145 cells. Growth kinetics revealed that deletion of m1B and m2B did not significantly affect virus replication. Hence, m1B and m2B show potential as molecular markers for developing a PRRSV vaccine.

In Situ Hybridization of PRRSV-1 Combined with Digital Image Analysis in Lung Tissues of Pigs Challenged with PRRSV-1

Betaarterivirus suid 1 and 2 are the causative agents of porcine reproductive and respiratory syndrome (PRRS), which is one of the most significant diseases of the swine industry, causing significant economic losses in the main pig producing countries. Here, we report the development of a novel, RNA-based in situ hybridization technique (RNAscope) to detect PRRS virus (PRRSV) RNA in lung tissues of experimentally infected animals. The technique was applied to lung tissues of 20 piglets, which had been inoculated with a wild-type, highly pathogenic PRRSV-1 strain. To determine the RNAscope’s applicability as a semi-quantitative method, we analysed the association between the proportion of the virus-infected cells measured with an image analysis software (QuPath) and the outcome of the real-time quantitative reverse transcription polymerase chain reaction (qRT-PCR) tests performed in parallel. The results of the quantitative approach of these two molecular biological methods show significant association (pseudo R2 = 0.3894, p = 0.004). This is the first time RNAscope assay has been implemented for the detection of PRRSV-1 in experimental animals.

Impacts of Quarterly Sow Mass Vaccination with a Porcine Reproductive and Respiratory Syndrome Virus Type 1 (PRRSV-1) Modified Live Vaccine in Two Herds

In recent years, there has been a considerable increase in the use of Modified Live PRRSV Vaccines (MLV) for mass vaccination in Denmark. The potential risks and negative impact of this strategy have been sparsely studied. The aim of this study was to investigate the impact of quarterly sow mass vaccination in two Danish sow herds. The study was performed as an observational prospective cohort of 120 sows in each of two commercial breeding herds in a paired design. Blood samples were taken from sows and oral fluid samples from nursery pigs (four to ten weeks old) before and after vaccination. The presence of PRRSV-1 RNA was measured by real time quantitative reverse transcription-polymerase chain reaction (RT-qPCR), and the level of PRRSV-1 specific antibodies was measured by two different serological assays. PRRS virus was not detected in the sow herds two days before and two weeks after vaccination, but the vaccine strain virus was detected in the nursery pigs. The prevalence of sows without antibodies towards PRRSV-1 went from 6–15% before vaccination to 1–4% after vaccination depending on the serological assay used, despite the fact that they had previously been repeatedly vaccinated. Four sows tested negative for antibodies in both assays after vaccination.

Host Genetics of Response to Porcine Reproductive and Respiratory Syndrome in Sows: Antibody Response as an Indicator Trait for Improved Reproductive Performance

Antibody response to porcine reproductive and respiratory syndrome (PRRS) virus (PRRSV) infection, measured as sample-to-positive (S/P) ratio, has been proposed as an indicator trait for improved reproductive performance during a PRRS outbreak in Landrace sows. However, this result has not yet been validated in Landrace sows or evaluated in terminal sire lines. The main objectives of this work were to validate the use of S/P ratio as an indicator trait to select pigs during a PRRS outbreak and to explore the genetic basis of antibody response to PRRSV. Farrowing data included 2,546 and 2,522 litters from 894 Duroc and 813 Landrace sows, respectively, split into pre-PRRS, PRRS, and post-PRRS phases. Blood samples were taken from 1,231 purebred sows (541 Landrace and 690 Duroc) following a PRRS outbreak for subsequent PRRSV ELISA analysis for S/P ratio measurement. All animals had high-density genotype data available (29,799 single nucleotide polymorphisms; SNPs). Genetic parameters and genome-wide association studies (GWAS) for S/P ratio were performed for each breed separately. Heritability estimates (± standard error) of S/P ratio during the PRRS outbreak were moderate, with 0.35 ± 0.08 for Duroc and 0.34 ± 0.09 for Landrace. During the PRRS outbreak, favorable genetic correlations of S/P ratio with the number of piglets born alive (0.61 ± 0.34), number of piglets born dead (−0.33 ± 0.32), and number of stillborn piglets (−0.27 ± 0.31) were observed for Landrace sows. For Duroc, the GWAS identified a major quantitative trait locus (QTL) on chromosome (Chr) 7 (24-15 megabases; Mb) explaining 15% of the total genetic variance accounted for by markers (TGVM), and another one on Chr 8 (25 Mb) explaining 2.4% of TGVM. For Landrace, QTL on Chr 7 (24–25 Mb) and Chr 7 (108–109 Mb), explaining 31% and 2.2% of TGVM, respectively, were identified. Some of the SNPs identified in these regions for S/P ratio were associated with reproductive performance but not during the PRRS outbreak. Genomic prediction accuracies for S/P ratio were moderate to high for the within-breed analysis. For the between-breed analysis, these were overall low. These results further support the use of S/P ratio as an indicator trait for improved reproductive performance during a PRRS outbreak in Landrace sows.

Fetale Hypoxie und Apoptose nach maternaler Infektion mit dem PRRS-Virus

Malgarin CM, Moser F, Pasternak JA et al. Fetal hypoxia and apoptosis following maternal porcine reproductive and respiratory syndrome virus (PRRSV) infection. BMC Vet Res 2021; 17: 182 Das Porzine Reproduktive und Respiratorische Syndrom gehört zu den bedeutendsten viralen Schweineerkrankungen und geht mit hohen wirtschaftlichen Verlusten einher. Etwa die Hälfte dieser Verluste ist durch Aborte, totgeborene oder lebensschwache Ferkel sowie Kümmerer auf die reproduktive Form der Erkrankung bei maternaler Infektion in der Spätträchtigkeit zurückzuführen. Die Mechanismen für den fetalen Tod nach maternaler Infektion sind noch weitgehend ungeklärt. Vermutet wird eine Hypoxie infolge von Nabelschnurläsionen oder Plazentaablösung durch Apoptose. Ziel der Studie war der Nachweis von Anzeichen für Apoptose sowie Hypoxie im fetalen Gewebe.

The Ability of Porcine Reproductive and Respiratory Syndrome Virus Isolates to Induce Broadly Reactive Neutralizing Antibodies Correlates With In Vivo Protection

Porcine reproductive and respiratory syndrome (PRRS) is considered one of the most relevant diseases of swine. The condition is caused by PRRS virus (PRRSV), an extremely variable virus of the Arteriviridae family. Its heterogeneity can be responsible, at least partially, of the poor cross-protection observed between PRRSV isolates. Neutralizing antibodies (NAs), known to play a role in protection, usually poorly recognize heterologous PRRSV isolates, indicating that most NAs are strain-specific. However, some pigs develop broadly reactive NAs able to recognize a wide range of heterologous isolates. The aim of this study was to determine whether PRRSV isolates that induce broadly reactive NAs as determined in vitro are able to confer a better protection in vivo. For this purpose two in vivo experiments were performed. Initially, 40 pigs were immunized with a PRRSV-1 isolate known to induce broadly reactive NAs and 24 additional pigs were used as controls. On day 70 after immunization, the pigs were divided into eight groups composed by five immunized and three control pigs and exposed to one of the eight different heterologous PRRSV isolates used for the challenge. In the second experiment, the same experimental design was followed but the pigs were immunized with a PRRSV-1 isolate, which is known to generate mostly strain-specific NAs. Virological parameters, specifically viremia and the presence of challenge virus in tonsils, were used to determine protection. In the first experiment, sterilizing immunity was obtained in three groups, prevention of viremia was observed in two additional groups, although the challenge virus was detected occasionally in the tonsils of immunized pigs, and partial protection, understood as a reduction in the frequency of viremia compared with controls, was recorded in the remaining three groups. On the contrary, only partial protection was observed in all groups in the second experiment. The results obtained in this study confirm that PRRSV-1 isolates differ in their ability to induce cross-reactive NAs and, although other components of the immune response might have contributed to protection, pigs with cross-reactive NAs at the time of challenge exhibited better protection, indicating that broadly reactive NAs might play a role in protection against heterologous reinfections.