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Porcine reproductive and respiratory syndrome virus (PRRSV) has a strict cell tropism. In addition to the primary alveolar macrophages, PRRSV is strictly cytotropic to African green monkey kidney cells, such as MARC-145 cells; however, MARC-145 cells are not infected by most NADC30-like and NADC34-like PRRSV strains. The essential scavenger receptor CD163 has been proved to mediate productive infection of PRRSV in various non-permissive cell lines. In this study, we systematically tested the porcine CD163 stably expressing 3D4/21 cells for infections with various PRRSV strains. The results showed that the porcine CD163-expressing macrophages support the infections of PRRSV2 of lineages 1, 5, and 8, as evidenced by Western blotting, immunofluorescence assay, quantitative PCR, and virus titration assay. Considering the current prevalence of NADC30-like and NADC34-like PRRSV2 of lineage 1 in China, the CD163-expressing macrophages are very useful for PRRSV research and disease management. [ABSTRACT FROM AUTHOR]

Multiple commercial porcine reproductive and respiratory syndrome (PRRS) modified live vaccines are currently utilized in Chinese swine herds due to the limited cross‐protection of vaccines and coexistence of different PRRS viruses. In this study, an infectious cDNA clone of the highly pathogenic PRRS (HP‐PRRS) vaccine JXA1‐R strain was generated. We successfully rescued the virus from direct in vitro DNA transfection of rJXA1‐R clone, which has similar growth kinetics to the parental JXA1‐R virus in Marc‐145 cells. To further evaluate the potential use of the cloned rJXA1‐R virus as a live vector for foreign gene expression, the enhanced green fluorescent protein (EGFP) was inserted between non‐structural and structural genes. Our results showed that the dynamic expression of EGFP can be visualized by live cell imaging system during the infection in Marc‐145 cells. The availability of our cloned JXA1‐R viruses provides a crucial platform to study the fundamental biology of HP‐PRRS virus vaccine and also serves as a potential effective vector for developing live vector vaccines against swine pathogens. [ABSTRACT FROM AUTHOR]

Porcine reproductive and respiratory syndrome virus 1 (PRRSV1) and 2 (PRRSV2) (including 3 major subtypes: classical (CA‐PRRSV2), highly pathogenic (HP‐PRRSV2) and NADC30‐like (NL‐PRRSV2)) are currently coexisting in Chinese swine herds but with distinct virulence. Reliable detection and differentiation assays are crucial to monitor the prevalence of PRRSV and to adopt effective control strategies. However, current diagnostic methods cannot simultaneously differentiate the four major groups of PRRSV in China. In this study, universal and quadruplex real‐time RT‐PCR assays using TaqMan‐MGB probes were developed for simultaneous detection and differentiation of Chinese PRRSV isolates. The newly developed real‐time RT‐PCR assays exhibited good specificity, sensitivity, repeatability and reproducibility. In addition, the newly developed real‐time RT‐PCR assays were further validated by comparing with a universal PRRSV conventional RT‐PCR assay on the detection of 664 clinical samples collected from 2016 to 2019 in China. Based on the clinical performance, the agreements between the universal and quadruplex real‐time RT‐PCR assays and the conventional RT‐PCR assay were 99.55% and 99.40%, respectively. Totally 90 samples were detected as PRRSV‐positive, including 2 samples that were determined to be co‐infected with NL‐PRRSV2 and HP‐PRRSV2 isolates by the quadruplex real‐time RT‐PCR assay. ORF5 sequencing confirmed the real‐time RT‐PCR results that 2, 6, 27 and 57 of the 92 sequences were PRRSV1, CA‐PRRSV2, NL‐PRRSV2 and HP‐PRRSV2, respectively. This study provides promising alternative tools for simultaneous detection and differentiation of PRRSV circulating in Chinese swine herds. [ABSTRACT FROM AUTHOR]

Porcine reproductive and respiratory syndrome virus (PRRSV) causes huge economic loss to the global swine industry. Even though several control strategies have been applied, PRRS is still not effectively controlled due to the continuous emergence of new variants and limited cross-protection by current vaccines. During the routine epidemiological investigation in 2017, two PRRSV variants were identified from a severe abortion farm and a clinically healthy farm, respectively. The viruses were isolated and denominated as XJ17-5 and JSTZ1712-12. Genomic sequencing indicated that their genomes are both 14,960 bp in length sharing 99.45% nucleotide identity. Sequence alignments identified a discontinuous 30-amino-acid deletion and a continuous 120-amino-acid deletion in nsp2 of both isolates. Genome-based phylogenetic analysis confirmed that XJ17-5 and JSTZ1712-12 belong to the HP-PRRSV subtype but form a new branch with other isolates containing the same 150-amino-acid deletion in nsp2. Pathogenic analysis showed that XJ17-5 is highly virulent causing 60% mortality, while JSTZ1712-12 is avirulent for piglets. Furthermore, fragment comparisons identified 34-amino-acid differences between XJ17-5 and JSTZ1712-12 that might be associated with the distinct virulence. The identification of highly homologous HP-PRRSV variants with new genetic feature and distinct virulence contributes to further analyze the pathogenesis and evolution of PRRSV in the field. [ABSTRACT FROM AUTHOR]

Abstract Classical swine fever virus (CSFV), porcine reproductive and respiratory syndrome virus (PRRSV) and porcine circoviruses (PCV2 and PCV3) are economically important swine viruses that cause reproductive failure and/or respiratory symptoms in pigs. However, the co-infection status of these viruses in Chinese swine herds is not well clarified. In this study, we evaluated the co-infection of these four viruses in 159 pigs collected from 63 herds in eight regions of China from 2016 to 2018. CSFV, PRRSV, PCV2 and PCV3 were detected in 14, 56, 43 and 4 of the pigs, respectively. The percentage of singular infections was 32.71%, while the percentages of dual infections and multiple infections were 15.72% and 3.15%, respectively. The E2 of CSFV, ORF5 of PRRSV, ORF2s of PCV2 and PCV3 from all positive samples were determined and used for phylogenetic analyses. E2-based phylogenetic tree showed that all 14 CSFVs identified in this study belong to 2.1b subtype. ORF5-based phylogenetic tree showed that PRRSV2 is predominant in China while PRRSV1 can also be detected. In addition, 35, 16, 4 and 1 of our PRRSVs are clustered with highly pathogenic PRRSV2, NADC30-like PRRSV2, classical PRRSV2 and PRRSV1, respectively. ORF2-based phylogenetic trees showed that our PCVs are grouped with 2 PCV2 subtypes (PCV2d and PCV2b) and 3 PCV3 subtypes (PCV3a, PCV3b and PCV3c), respectively. Our results provide the latest co-infection status and the diversity of four important swine viruses in Chinese swine herds, which is beneficial for understanding the epidemiology of these viruses. Highlights • The co-infection status of CSFV, PRRSV, PCV2 and PCV3 in eight regions of China was determined. • Both singular infections and different patterns of co-infections were observed. • The diversity of these four viruses was also evaluated by phylogenetic analyses. [ABSTRACT FROM AUTHOR]