Isolation and characterization of the first porcine Getah virus strain HNJZ-S1 from an aborted piglet in China
Running Head: the first porcine Getah virus strain in China
Feng Zhou1, Aojie Wang1, Hongtao Chang1, Dandan Cui1, Xingang Wang1, Lu Chen1*, Chuanqing Wang1*
1College of animal science and veterinary medicine, Henan Agricultural University, Agricultural Road No. 63, Zhengzhou, Henan, China (450002)
*Address correspondence to: Chuanqing Wang, E-mail:wchuanq@163.com
Lu Chen, E-mail: chenluhau@126.com
ABSTRACT
Severity of porcine reproductive and respiratory syndrome (PRRS) implicated co-infection with other pathogens in pig herd in China, which normally were found Japanese encephalitis virus (JEV), pseudorabies virus (PRV), porcine reproductive and respiratory syndrome virus (PRRSV), porcine circovirus, porcine parvovirus and classical swine fever virus (CSFV). Here we verified a specific fragment which was similar to Getah virus (GETV) by RT-PCR and sequencing from aborted fetuses in Henan province, China. Then we isolated and purified the stain virus. We named it as HNJZ-S1. Furthermore, we characterized HNJZ-S1 by passage and plaque titer, whole genome sequencing, Electron microscopy and animal infection experiments with mice and piglets. The data show that the full-length genome sequence of HNJZ-S1was 11689 bp with homologous 97.4%–99.3% identical to the GETV sequence available in GenBank. Phylogenetic tree analysis showed that HNJZ-S1 was closely related to mosquito isolate 12IH26 from Japan, horse isolates 14-I-605-C1 and 14- I-605-C2, and mosquito isolate HB0234 from China. Animal experiments showed that the isolate HNJZ-S1 caused abortion in pregnant mice and was lethal postnatal day 3 mice and piglets. This study provides important information on the tropism, infection, pathogenicity, environmental microbiology and public health implications of pig derived GETV and will also serve as a source of reliable research materials.
Keywords: Swine, Getah virus, Abortive, Virus isolation, Pathogenicity
The mosquito borne Geta virus (GETV) belongs to the genus Alphavirus , family Otariidae . The viral genome of GETV is composed of positive single-stranded RNA including a 5’-cap structure, two open reading frames (ORFs) and a 3’-poly(A) tail. The two ORFs encodes the structural (C/capsid, E3, E2, 6K, E1) and non-structural proteins (NSP1 to NSP4) respectively, which are flanked by a 44 nt 26S junction region associated with transcription of an intracellular subgenomic 26S RNA(Strauss et al., 1994). Previous serological studies showed that this virus can infect multiple species, including humans(Li et al., 1992; Marchette et al., 1980 ), horses, pigs, cattle, monkeys and birds(Li et al., 2007). The symptom of infection includes rash, fever and joint edema in horses(KAMADA et al.,1991; KAMADA et al.,1991), reproductive disorders in pregnant sows and tremors in piglets, deaths only occur in pigs in the fetal stage(Yago et al., 1987; Kawamura et al., 1987).
GETV was well known that the first isolated from Culex  sp. mosquitoes in Malaysia in 1955 and is now widely distributed across Asia (Ksiazek et al., 1981) (Morita and Igarashi, 1984)(Ksiazek et al., 1981; Morita et al., 1984; Leake et al., 1986; Bryant et al., 2005), Europe and Australia based on viral isolation and/or molecular epidemiological investigations(Fukunaga et al., 2000). In 1964, GETV was first isolated from mosquitoes on Hainan Island in China (Li et al., 1992) and was later detected in mosquitoes in several provinces and cities (Zhai et al., 2008). In addition, recently two high pathogenic swine GETV strains were detected in blue foxes in China (Shi et al., 2019).
In November 2016, during attempt to detect pathogens in aborted fetuses from a pig farm in Henan Province, we found that classical swine fever virus (CSFV), porcine reproductive and respiratory syndrome virus (PRRSV), Japanese encephalitis virus (JEV) and pseudorabies virus (PRV) were negative. Interestingly, a non-specific band was amplified using CSFV primers. The non-specific band was sequenced and analyzed by BLAST search against the GenBank database. It turned out that the amplicon had high homology to GETV. GETV was isolated by cell passage and plaque purification, and was subjected to electron microscopy observation, genome sequencing. Moreover, we determined the pathogenicity of the strain for pregnant and neonatal mice, and the piglets.