Introduction
SADS-CoV, also name PEAV (Gong et al., 2017) and SeACoV (Y. Pan et al., 2017), as the newest strain of porcine coronaviruses (CoVs), was first detected in pig herds with diarrhea-outbreak in Guangdong, China in 2017 (P. Zhou et al., 2018). SADS-CoV belongs to the genus Alphacoronavirus of the family Coronaviridae (P. Zhou et al., 2018), which also includes Transmissible gastroenteritis virus (TGEV) (Doyle & Hutchings, 1946), Porcine epidemic diarrhea virus (PEDV) (Pensaert & de Bouck, 1978) and Porcine deltacoronavirus (PDCoV) (Woo et al., 2012). SADS-CoV genome is an enveloped, single-stranded, positive-sense RNA virus, which is approximately 27 kb in length, and arranged in the order of: 5’ UTR-ORF1a/1b-S-NS3-E-M-N-NS7a-3’ UTR, encoding 16 non-structural proteins and 4 structural proteins (Gong et al., 2017; Y. Pan et al., 2017; P. Zhou et al., 2018). SADS-CoV caused clinical symptoms similar to other porcine enteric pathogens, such as TGEV, PEDV and PDCoV, characterized by vomiting, diarrhea, dehydration, and a mortality rate as high as 90% in piglets (Xu et al., 2019; P. Zhou et al., 2018). Since SADS-CoV was first reported in southern China in 2017 (Gong et al., 2017), the retrospective investigation of 236 samples from 45 swine farms showed that the prevalence of PEAV in pigs was reported to be 43.53% in Guangdong, China in August 2016 to May 2017 (L. Zhou, Sun, et al., 2019). And yet from May 2017 to January 2019, there were no new SADS cases reported in pig herds in Guangdong, but the re-emerging of SADS-CoV infection in pig herds in Guangdong on February 2019 (L. Zhou, Li, et al., 2019), indicating a continuing threat to pig farms. Apart from Guangdong, a SADS-CoV-like strain, CN/FJWT/2018, was recently discovered in Fujian, China (K. Li et al., 2018). Recently, it was reported that SADS-CoV could infect human cell lines (Edwards et al., 2020; Yang et al., 2019) and in view of the great harm of SARS-CoV2 (Hu, Guo, Zhou, & Shi, 2020), which same cross-species infection likely originate from bats as SADS-CoV (Hu et al., 2020; P. Zhou et al., 2018), reveal that the monitoring of epidemic situation of SADS-CoV in pig herds might have important public health significance.
Since first detection in swine herds in 2017, multiple detection methods including real time reverse transcription loop-mediated isothermal amplification method (RT-LAMP) (H. Wang et al., 2018), SYBR green-based real-time PCR (Ma et al., 2019), multiplexTaq Man-probe-based real-time PCR (Z. Pan et al., 2020)), microfluidic-RT-LAMP chip (L. Zhou et al., 2020) are developed and widely used to detect SADS-CoV infection. However, these detection methods only target viral nucleic acid, serological assays for SADS-CoV to undertake epidemiological studies are not available to date. Enzyme-linked immunosorbent assay (ELISA), which mainly including indirect ELISA, double-antibody sandwich ELISA and blocking ELISA, has strong specificity and high sensitivity and is widely used in pathogens antibodies detection and vaccines evaluation (Knuchel, Ackermann, Muller, & Kihm, 1992). The basic principle of ELISA is the specific response between antigen and antibody. Therefore, for pathogen specific antibody detection, it is critical to select the appropriate detect antigen. S protein of CoVs belong to type I membrane fusion protein, locate on the envelope of the virus particle (Yu, Qiao, Guo, & Wang, 2020). It is known that the S protein, which can be divided into domain 1 (S1) and domain 2 (S2), plays an important role in virus attachment and entry, and induction of neutralizing antibodiesin vivo (Wen et al., 2018). In addition, the S protein has higher antigenicity than any of the other CoVs’ proteins, and anti-S antibodies persist longer than anti- nucleocapsid (N) antibodies in vivo(Knuchel et al., 1992). Therefore, Many studies have selected the whole or part of S protein as a coating antigen to develop indirect ELISA methods for the detection of TGEV (Sestak, Zhou, Shoup, & Saif, 1999), PEDV (Lin et al., 2018) and PDCoV (Lu et al., 2020) antibodies. Although SADS-CoV has the smallest S protein of only 1130 amino acids (Gong et al., 2017) compared to other reported CoVs, the structural features of SADS-CoV S protein are evolutionarily related to other CoVs genera (Guan et al., 2020), indicating that S protein of SADS-CoV can also be used as detect antigen.
SADS-CoV is an important enteropathogen in pigs, but currently no suitable serology assays to detect the specific antibodies for SADS-CoV. The objective of this study was to develop an anti-SADS-CoV IgG indirect ELISA based on recombinant S protein and utilize this ELISA to investigate the prevalence rates of SADS-CoV in the pig farms of China, which might provide a tool for advancing understanding of the epidemiology of SADS-CoV.