Molecular characterization and recombination analysis of porcine reproductive and respiratory syndrome virus emerged in southwestern China during 2012–2016

Abstract

Porcine reproductive and respiratory syndrome virus (PRRSV) is an important swine pathogen causing tremendous economic losses to the swine industry. To investigate the prevalence of PRRSV of genotype 2 (North American type, NA-type) in southwestern China, the Nsp2 hypervariable region (Nsp2 HV) and ORF5 of 61 PRRS viruses collected during 2012-2016 were sequenced and analyzed. All the virus detected clustered into the JXA1-like (52/61), VR-2332-like (7/61), and NADC30-like (2/61) sub-genotypes. Five deletions in Nsp2 HV were detected in addition to the typical 30aa discontinuous deletion in HP-PRRSV, and two of these five were not reported previously. Strikingly, two PRRS virus (SCnj16 and SCcd16) isolated in 2016 contained the classic HP-PRRSV molecular marker in the Nsp2-coding region, but belonged to the NADC30-like sub-genotype on the ORF5 gene. Further recombination and phylogenetic analysis on the two complete genomic sequences revealed that they may have originated from recombination events between the NADC30 and Chinese HP-PRRSV strains. The present study suggests that the endemic PRRSVs in the region have continuously evolved and new vaccine strategies are necessary for more efficient control of the virus.

Genetic Characterization and Pathogenicity of a Novel Recombined Porcine Reproductive and Respiratory Syndrome Virus 2 among Nadc30-Like, Jxa1-Like, and Mlv-Like Strains

“…However, the complex recombination pattern among a NADC30-like, a JXA1-like, and the classical PRRSV-2-derived MLV vaccine strain (RespPRRS MLV) has never been described. In our previous investigation, we found that JXA1-like, NADC30-like, VR-2332-like, and QYYZ-like strains had coexisted in some pig herds in the Sichuan Province since 2016, and the following different recombination patterns were observed in this region: (i) NADC30-like + JXA1-like [ 23 ]; (ii)NADC30-like + JXA1-like + VR-2332-like [ 27 ]; and (iii) NADC30-like + JXA1-like + QYYZ-like [ 28 ]. This evidence thus further supported the possible recombination among the PRRSV strains, NADC30-like and JXA1-like, and the vaccine strain, RespPRRS MLV (VR-2332-like).…”

“…RT-PCR was used to confirm PRRSV positivity in suspensions of lung tissue homogenates, which were then inoculated into PAMs. The PAMs were maintained in RPMI-1640 medium (Transgene, Beijing, China) supplemented with 10% fetal bovine serum (HyClone, Logan, UT, USA) at 37 °C in a humidified 5% CO 2 atmosphere as described previously [ 23 ]. Indirect immunofluorescence assay (IFA) was performed to detect viral antigens in PAMs using GTX129270 (a monoclonal antibody against PRRSV N protein; GeneTex Inc., Irvine, CA, USA).…”

“…cDNA was generated using a PrimeScript TM RT reagent Kit (TaKaRa, Dalian, China) following the supplier’s guidelines. Twelve pairs of overlapped primers specific for the PRRSV-2 genome were utilized for amplifying the complete genome of SCN17, as described previously [ 23 ]. Further, the purified RT-PCR products were ligated to a pMD19-T cloning vector (TaKaRa, Dalian, China), and at least three independent recombinant clones were sequenced by Sangon Biological Technology (Beijing, China).…”

“…Recently, QYYZ-like PRRSV strains were discovered to recombine with JXA1-like strains (GDsg and GD1404) [ 15 , 16 ]. Additionally, NADC30-like strains were found to recombine with JXA1-like strains (HENAN-HEB, JL580, FJ1402, TJnh1501, 15HEN1, HNhx, and SCnj16) [ 17 , 18 , 19 , 20 , 21 , 22 , 23 ] or VR-2332-like strains (Chsx1401, HENAN-XINX, and HNyc15) [ 14 , 20 , 24 ]. Moreover, several complex recombinants among the three lineages of PRRSV-2 strains (SDhz1512, SD17-38, SCcd16, SCcd17, and SCya17) have also been observed in China [ 25 , 26 , 27 , 28 ].…”

Identification of a Novel Recombinant Type 2 Porcine Reproductive and Respiratory Syndrome Virus in China

“…First-strand cDNA was synthesized using random hexamers (TaKaRa, Dalian, China) following the manufacturer’s instructions. The complete genomic sequences of SCcd17 were amplified with fourteen overlapping fragments by reverse transcription-PCR (RT-PCR) as described previously [ 32 ]. The PCR products were purified and cloned into pMD19-T vector (TaKaRa, Dalian, China) following the manufacturer’s instructions, and then sequenced at least three times by the commercial service (Sangon, Shanghai, China) using Sanger sequencing approach.…”