Genome characterization of two NADC30-like porcine reproductive and respiratory syndrome viruses in China
- Xiangdong Li†2,
- Jiajun Wu†3,
- Feifei Tan2,
- Yingying Li2,
- Guobiao Ji2,
- Jinshan Zhuang2,
- Xinyan Zhai3Email author and
- Kegong Tian1, 2, 3Email author
© The Author(s) 2016
Received: 4 February 2016
Accepted: 20 September 2016
Published: 29 September 2016
The recent emergence of NADC30-like porcine reproductive and respiratory syndrome virus (PRRSV) in vaccinated pigs arose more attentions for the high incidents of mutation and recombination of PRRSVs.
In this study, we determined full-length genome sequences of two NADC30-like PRRSV isolates from recent PRRSV outbreaks in China. Phylogenetic analysis showed that these two isolates were clustered in an independent branch together with NADC30, an American isolate in 2008. Genetically, HNjz15 shared 95.6 % nucleotide similarity to NADC30 without any exotic gene insertion. By contrast, HNyc15 shared 93.8 % similarity to NADC30 with recombination with VR-2332 and CH-1a. Two more previously reported NADC30-like PRRSVs were also analyzed and had exotic gene insertions with different PRRSV strains in their nonstructural protein genes.
The above results showed the increased mutation and recombination rates of NADC30-like PRRSV under current vaccination pressure and a more pressing situation for the PRRSV eradication and control in China.
KeywordsPRRSV NADC30-like Recombination HNjz15 HNyc15
Chinese pig industry has been obsessed by porcine reproductive and respiratory syndrome (PRRS) for decades, especially since the outbreak of highly pathogenic PRRS (HP-PRRS) in 2006 (Tian et al. 2007). The causative agent porcine reproductive and respiratory syndrome virus (PRRSV) belongs to the Order Nidovirales, family Arteriviridae (Conzelmann et al. 1993). PRRSV can be divided into European genotype 1 and North American genotype 2 with VR-2332 and Lelystad as prototypical strains, respectively. PRRSV genome is about 15 kb in length and contains at least 11 open reading frames. ORF1a and ORF1b constitute nearly 75 % of the viral genome and are cleaved into at least 14 nonstructural proteins that are responsible for genome replication and transcription (Zhou et al. 2015b). Three membrane-associated proteins, GP2a, GP3, and GP4 formed a hetero-trimer complex and are involved in virus entry (Li et al. 2016). Three major structural proteins GP5, unglycosyulated membrane protein M, and nucleocapsid protein N locate at 3′ end of genome, and are indispensable for both virion formation and viral infectivity (Hu and Zhang 2014).
Recently, several field isolates of PRRSV had a very unique genetic background and showed the highest nucleotide similarity to a group represented by NADC30, a type 2 PRRSV that has been isolated in Unite States of America in 2008 (Zhao et al. 2015; Brockmeier et al. 2012; Zhou et al. 2015a). These viruses therefore were designated as NADC30-like PRRSV in China. The first two NADC-30 like PRRSVs, HENAN-XINX (access number KF611905) and HENAN-HEB (KJ143621), were isolated in Henan province and their whole genome became available on NCBI in 2013. The clinical symptoms of NADC30-like PRRSV infection were characterized by respiratory disorders of piglets and abortions of pregnant sows in vaccinated pigs which indicate the inability of current commercial PRRSV vaccine to protect NADC30-like PRRSV infection. So far, the disease has been reported to be widely spread in several provinces and led to huge amount of economic losses in China since 2014 (Zhou et al. 2015a).
Results and discussion
Comparison of complete genomes of HNjz15 and HNyc15 with NADC30 and JXA1
HNjz15 to JXA1
HNyc15 to JXA1
HNjz15 to NADC30
HNyc15 to NADC30
An unusual phenomenon of these NADC30-like PRRSVs as compared with other PRRSV variants is the unparalleled incidence of genome-wide recombination with other strains of PRRSV including both classical type 2 PRRSV such as VR-2332 and HP-PRRSV (Zhao et al. 2015). In Zhao’s study, JL580 NADC30-like PRRSV has 6 recombination breakpoints between NADC30 and a HP-PRRSV 09HEN1 (two locate in nsp2, others locate in nps3, nsp7, ORF2a, and ORF4) (Zhao et al. 2015). To explore the recombination of our NADC30-like isolates together with two previously reported two NADC30-like strains (HENAN-XINX and HENAN-HEB), recombination incidences were analyzed by performing similarity within 500-bp window sliding along the genome alignment with 20 bp step size (SimPlot v3.5.1). The analysis results showed that HNjz15 shared 95.6 % nucleotide similarity with NADC30 without any exotic gene insertion, while HNyc15 shared 93.8 % with NADC30 with recombination with VR-2332 and CH-1a between ORF2 and ORF4 (Additional file 1: Appendix Fig. S1A, B). However, it was possible the recombination could occur in cell culture since HNyc15 was propagated on PAM before it was subjected to genome sequencing. By contrast, HENAN-XINX has recombination between NADC30 and VR-2332 in nsp2–5 (Additional file 1: Appendix Fig. S1C), and HENAN-HEB has recombination between NADC30 and JXA1 in nsp2 (Additional file 1: Appendix Fig. S1D). Therefore, unlike the previous PRRSV field isolates, NADC30-like PRRSVs have higher incidences of recombination with both vaccine strains and field isolates.
The recombination of gene segments among different PRRSV strains may lead to the change of virulence (Zhao et al. 2015). In Zhao’ study, the JL580 PRRSV strain was a mosaic NADC30-like virus with HP-PRRSV 09HEN1 recombination at six different sites spanned the genome (Zhao et al. 2015). The pathogenicity of JL580 was tested on 6-week-old pigs and was much higher than that of parental strain NADC-30. One limitation of this study was that we did not perform the animal experiments on non-vaccinated and vaccinated pigs to test the pathogenicity of these two NADC30-like PRRSVs. However, the different recombination patterns of HNjz15 (without recombination), HNyc15 (recombination with VR-2332 and CH-1a), HENAN-HEB (recombination with JXA1), and HENEN-XING (recombination with VR-2332) summarized in this study may provide valuable virus resources to study the change of pathogenicity of PRRSVs through different recombination patterns and genomic sites.
Two NADC30-like PRRSVs were isolated and analyzed based on the genome level. Phylogenic analysis showed that they are most closed to the NADC30 strain. Genome analysis revealed they undergone unusual frequency of recombination events and became prevalent in China even though the HP-PRRSV still is the dominating strain in Chinese pig herds. The recombination events of these NADC30-like PRRSVs with other PRRSVs are more complex than we expected, and these mosaic PRRSVs show obvious distinct pathogenicity according to the strain they exchanged. The different recombination patterns and pathogenicity of these NADC30-like PRRSVs may lead to more pressing situation for PRRSV control in China.
KT and XZ conceived and designed the experiments. XL, JW, FT, YL, GJ, and JZ performed the experiments and analyzed the data. XL and KT wrote the paper. All authors read and approved the final manuscript.
This study was funded by grant from The National Natural Science Foundation of China (Grant No. 3149600031), Major Science and Technology Program in Henan Province (Grant No. 131100110200), Innovation Scientists and Technicians Troop Construction Projects of Henan Province (Grant No. 142101510001), and Talents Plan for Scientific and Technological Innovation in Henan Province (Grant No. 144200510002).
The authors declare that they have no competing interests.
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