key: cord-342766-ndzhlf3k
authors: Ku, X.; Chen, F.; Li, P.; Wang, Y.; Yu, X.; Fan, S.; Qian, P.; Wu, M.; He, Q.
title: Identification and genetic characterization of porcine circovirus type 3 in China
date: 2017-03-19
journal: Transbound Emerg Dis
DOI: 10.1111/tbed.12638
sha: 
doc_id: 342766
cord_uid: ndzhlf3k

A novel circovirus called porcine circovirus type 3 (PCV3) was recently reported to exist in the USA. This circovirus is associated with porcine dermatitis, nephropathy syndrome and reproductive failure. This study reports on the first identification, widely epidemic, different phylogenetic clusters, potential role in sow reproductive failure and possible origins of PCV3 in China.

Since 2010, new circoviruses from porcine samples were only initially characterized in some studies (Li et al., 2010; Zhang, Li, Deng, Kapusinszky, & Delwart, 2014) . In 2016, a novel circovirus, called PCV type 3 (PCV3), was reported to exist in the USA (Palinski et al., 2016) . PCV3 is associated with porcine dermatitis, nephropathy syndrome and reproductive failure (Palinski et al., 2016) and cardiac and multisystemic inflammation (Phan et al., 2016) . 

A total of 356 sows at three farms in the Liaoning and Jiangxi Provinces and Chongqin City have suffered from reproductive failure and acute loss of neonatal piglets since March 2016. The stillborn rate of the delivery sows ranged from 5.2% to 20.1% at these three farms, while sow mortality ranged from 5.4% to 10.5%. The general pig pathogens, such as PRV, PCV2, PRRSV, PEDV, TGEV, RV and HCV, were excluded by reverse transcription polymerase chain reaction (PCR) or simple PCR methods (Li, Li, Yan, Chen, & He, 2009; Yu et al., 2016; Zhang & He, 2010) . PCV3 was the only detected † These authors contributed equally to this article. pathogen in these cases. To better understand the infection status, epidemic status, geographical distribution, potential pathogenicity and genetic characteristics of PCV3 in China, a total of 222 samples (i.e., stillborn, tissues, semen and serum) were collected from 35 farms in 11 provinces or districts (i.e., Anhui, Chongqing, Fujian, Hebei, Henan, Hunan, Jiangsu, Jiangxi, Liaoning, Shenyang and Zhejiang) in China. These samples were subjected to pathogen detection in the Diagnostic Center of Animal Epidemic Diseases of Huazhong Agricultural University. The aforementioned samples were individually collected, stored, distilled in 2-or 5-ml EP tubes or clear package bags and then transported utilizing ice boxes. The stillborn and tissues samples were homogenized and diluted 10-fold with phosphate-buffered saline (PBS; 0.1 M, pH 7.4). The semen and serum samples were diluted 10-fold with PBS (0.1 M, pH 7.4). All the samples were frozen and thawed twice to release, further subjected to a vortex for 5 min, and centrifuged at 11,000 rpm (Eppendorf, Germany) for 8 min at 4°C. The supernatants were utilized for DNA and RNA extraction immediately or stored at À80°C refrigerator for further usage. A pair of primers was designed to detect PCV3 ( Table 1 ).

The detection limit of this method was 20.5 pg nuclear acid. The primers have no cross-reactions with the PR, PCV2, TGEV, TGEV, RV, PRRSV, porcine parvovirus and deltacoronavirus temples stored in our laboratory. Two pairs of primers were utilized for the entire genome sequencing ( Table 1) .

The reaction conditions were as follows: pre-denaturation at 94°C for 5 min, 35 cycles of denaturation at 94°C for 30 s, annealing at 55°C for 30 s, extension at 72°C for 1 min and a final extension at 72°C for 10 min. The amplicons for the genome sequencing were gel-purified, cloned into pMD â 18-T Vector (Takara, Japan) and also plays a role in these cases must be further researched. The singular infection rate of PCV3 in the samples was 18.9% (42/222 belonged to cluster 3a. The sequence similarity of these CV isolates is plotted in Figure 2 to study the genetic relationship of the Chinese PCV3 isolates with the Chinese Bat CVs. Two recombination regions were identified between the bat CVs and PCV3 isolates in 264-564nt

and 714-1148nt ( Figure 2 ). These results suggest that PCV3 can be the recombination results of the bat CVs.

PCV3 has been poorly understood because it was first identified in the USA in 2010 (Li et al., 2010) . PCV3 has been identified as a pathogen agent associated with pig diseases (Palinski et al., 2016 

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Similarity plot of the whole genome of 10 PCV3 isolates with 5 Bat circovirus isolates. The Chinese PCV3 isolates, PCV3/CN/ Fujian-5/2016, was set as query strain. The similarity plot was constructed using the two-parameter (Kimura) distance model with a sliding window of 200 bp and step size of 20 bp. The vertical and horizontal axes indicate the nucleotide similarity per cent and nucleotide position (bp) in the graph, respectively

PCV3/CN/Fujian-5/2016 KY075986.1 China

Complete genome analysis

Complete genome analysis

Complete genome analysis

Complete genome analysis

Complete genome analysis

Complete genome analysis

Complete genome analysis

KY075995.1 China:Anhui 2016 Partial capsid gene analysis 11 PCV3/CN/Fujian-5/201608 KY075996.1 China:Fujian 2016 Partial

China:Fujian 2016 Partial capsid gene analysis

KY076006.1 China:Liaoning 2016 Partial


