GENOME ANNOUNCEMENT
Complete Genome Sequence of a Highly Pathogenic Porcine Reproductive and Respiratory Syndrome Virus NM1 Strain from Northern China Zhenguang Li,a Xue Leng,a Qiaoling Qi,b Fengxue Wang,a Yongjun Wen,a Bin Tan,a Yanliang He,b Mingqi Xia,b Wei Lu,b Lizhi Chen,a Shipeng Cheng,a and Hua Wua,b Division of Zoonoses, Institute of Special Economic Animal and Plant Sciences, Chinese Academy of Agricultural Sciences, Jilin, China,a and Sinovet (Beijing) Biotechnology Co., Ltd., Beijing, Chinab
NM1 is a highly pathogenic North American-type porcine reproductive and respiratory syndrome virus (PRRSV). The complete genome sequence shows that NM1 shares high sequence identity (99.2 to 99.4%) to other HP-PRRSV isolates, containing two discontinuous deletions, a 1-amino-acid deletion at position 481 and a 29-amino-acid deletion at positions 533 to 651, in nonstructural protein 2.
P
orcine reproductive and respiratory syndrome virus (PRRSV) is the causative agent of porcine reproductive and respiratory syndrome (PRRS), which is characterized by respiratory distress in piglets and reproductive failure in sows. PRRS results in great economic losses to the swine industry (7). PRRSV is a positivestrand RNA virus that belongs to the family Arteriviridae (2). PRRS, first reported in 1987 in the United States (5) and in 1996 in China (4), has become a well-recognized global swine disease (1, 12). The highly pathogenic PRRS (HP-PRRS) first emerged in Southern China in 2006. It was characterized by high fever and high mortality in pigs of all ages. The etiological agent of the disease is highly pathogenic PRRSV (HP-PRRSV), which contains a discontinuous 30-amino-acid deletion in nonstructural protein 2 (Nsp2) (10, 11). The virus is currently widespread in China and several Southeast Asian countries (3, 8). Here, we report the outbreak of the HP-PRRS in herds of pigs in Inner Mongolia, China. Sera and tissue samples were collected from diseased pigs suspected of being infected with PRRSV in 2008. The strain, named NM1, replicated well in Marc-145 cells and caused cytopathogenic effect (CPE) on the cell culture. Experimental animal infection revealed that all piglets infected with NM1 developed typical clinical symptoms of PRRS. Three of the five piglets died in the test, suggesting that the NM1 strain had a strong pathogenicity. To understand the diversity and the evolutionary characteristics of NM1, we determined its complete genome sequence. Fourteen pairs of primers were designed for amplifying 14 overlapping fragments of the NM1 isolate full-length genome based on the sequences of PRRSV strains VR-2332 and JXA1 (6). The PCR products were purified, cloned into the pMD18-T vector (TaKaRa), sequenced three times on an ABI Prism 3730 sequencer (Applied Biosystems), and subsequently assembled using DNAStar version 7.0 to obtain the complete genome sequence. The terminal sequences were acquired by using a kit for rapid amplification of cDNA ends (RACE; Clontech, Japan). Sequence alignment was performed using ClustalX 2.1 (9), and a phylogenetic tree was constructed using MEGA 4. The complete genome of the NM1 strain was 15,356 nucleotides in length, including the poly(A) tail. Phylogenetically, the NM1 shared 89.6% and 61.6% nucleotide identity with prototype North American strain VR-2332 and European strain Lelystad, respectively. Meanwhile, NM1 exhibited 99.4%, 99.2%, and 99.4% nucleotide homologies with the three other HP-PRRSV variants, TJ, JXA1, and
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HuN4 (GenBank accession numbers EU860248, EF112445, and EF635006, respectively). Multiple sequence alignment based on the Nsp2-deduced amino acid sequence showed that NM1 had two deletions. One site, corresponding to position 481 in PRRSV strain VR2332, had a single amino acid deletion, while the other deletion site, corresponding to position 533 to 561, had a 29-amino-acid deletion. These calculations concluded that NM1 was highly similar to other HP-PRRSV strains and demonstrated that a new highly pathogenic Northern American-type PRRSV had spread widely in northern China. Nucleotide sequence accession number. The genome sequence of PRRSV strain NM1 has been deposited in GenBank under accession number EU860249. ACKNOWLEDGMENTS This work was supported by a grant from the national “Eleventh FiveYear” science and technology support program (2009BADB4B00) from the Ministry of Sciences and Technology of China. The animal experiments herein described have been reviewed and approved by the local Animal Care & Use Committee of the Institute of Special Wild Economic Animal and Plant Sciences, Chinese Academy of Agricultural Sciences, Jilin, China.
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Received 27 September 2012 Accepted 28 September 2012 Address correspondence to Hua Wu,
[email protected]. Z.L. and X.L. contributed equally to this paper. Copyright © 2012, American Society for Microbiology. All Rights Reserved. doi:10.1128/JVI.02642-12
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