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Isolation and characterization of porcine epidemic diarrhea virus with a novel continuous mutation in the S1⁰ domain.

ABSTRACT: Porcine epidemic diarrhea virus (PEDV), which re-emerged in China in 2010, has caused severe economic losses to the global pig industry. In this study, a PEDV strain, designated PEDV WMB, was isolated from piglets with severe diarrhea on a pig farm in Henan Province of China. Whole-genome sequencing and analysis revealed that the PEDV WMB strain belongs to subtype G2c and has a unique continuous mutation in the S1⁰ antigenic epitope of the S protein. Moreover, the virus-neutralization (VN) test indicated that polyclonal antibodies against the S1⁰ protein of other G1 and G2 strains showed reduced VN reactivity to PEDV WMB. The pathogenicity of PEDV WMB was further investigated in 3 day-old piglets. PEDV infection-related clinical symptoms and morphological lesions were observed and confirmed by histopathological and immunohistochemical examination (IHC). These results illustrated that continuous mutation of the S1⁰ epitope might affect the immunogenicity or pathogenicity of PEDV, providing evidence of the need to monitor the genetic diversity of the virus and develop effective measures to prevent and control PEDV.

SUBMITTER: Han X

PROVIDER: S-EPMC10232790 | biostudies-literature | 2023

REPOSITORIES: biostudies-literature

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Isolation and characterization of porcine epidemic diarrhea virus with a novel continuous mutation in the S1<sup>0</sup> domain.

Han Xueying X Liu Yangkun Y Wang Yan Y Wang Tiejun T Li Ning N Hao Feng F Yao Lunguang L Guo Kangkang K

Frontiers in microbiology 20230518

Porcine epidemic diarrhea virus (PEDV), which re-emerged in China in 2010, has caused severe economic losses to the global pig industry. In this study, a PEDV strain, designated PEDV WMB, was isolated from piglets with severe diarrhea on a pig farm in Henan Province of China. Whole-genome sequencing and analysis revealed that the PEDV WMB strain belongs to subtype G2c and has a unique continuous mutation in the S1<sup>0</sup> antigenic epitope of the S protein. Moreover, the virus-neutralization ...[more]

PMID: 37275149

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Project description:BackgroundPorcine epidemic diarrhea virus (PEDV) is a highly contagious virus infecting pigs of all ages with high morbidity and mortality among newborn piglets. Currently, there is no effective vaccine available to protect the pigs from PEDV. The N-terminal subunit of spike protein (S1) is responsible for virus binding to the cellular receptor and contains a number of neutralizing antibody epitopes. Thus, we expressed and produced recombinant S1 protein to protect newborn piglets by immunization of sows.MethodsAffinity tagged PEDV S1 protein was expressed in a secretory form in yeast, insect and mammalian cells to identify the most suitable production system. Purified recombinant protein was analysed by SDS-PAGE, Western blot and deglycosylation assay. A pregnant sow was intramuscularly immunized three times with adjuvanted recombinant protein prior to farrowing. PEDV-specific immune responses in sera and colostrum of the sow and piglets were assayed by ELISA and virus neutralization assays. Piglets were challenged orally with PEDV, and clinical parameters were monitored for 6 days post-challenge.Results and conclusionOf three eukaryotic expression systems tested (yeast, insect-cell, and mammalian), expression by HEK-293 T cells gave the highest yield of protein that was N-glycosylated and was the most appropriate candidate for vaccination. Administration of the subunit vaccine in a sow resulted in induction of S1-specific IgG and IgA that were passively transferred to the suckling piglets. Also, high virus neutralization titres were observed in the serum of the vaccinated sow and its piglets. After PEDV challenge, piglets born to the vaccinated sow exhibited less severe signs of disease and significantly lower mortality compared to the piglets of a control sow. However, there were no significant differences in diarrhea, body weight and virus shedding. Thus, vaccination with S1 subunit vaccine failed to provide complete protection to suckling piglets after challenge exposure, and further improvements are needed for the development of a subunit vaccine that fully protects against PEDV infection.

Project description:BackgroundPorcine Epidemic Diarrhea (PED) is an acute and highly contagious enteric disease caused by PED virus (PEDV), characterized by vomitting, watery diarrhea and fatal dehydration with high mortality in sucking piglets of one week of age. Although PEDV induced cell apoptosis has been established in vitro and in vivo, the functional protein that contributes to this event remains unclear.MethodsThe activation or cleavage of main apoptosis-associated molecular such as AIFM1, caspase-3, caspase-8, caspase-9 and PARP in PEDV infected host cells were analyzed by western blotting. The nuclear change of infected cell was monitored by confocal immunofluorescence assay. The overexpressing plasmids of 16 non-structural proteins (Nsp1-16) and 6 structural proteins (M, N, E, ORF3, S1 and S2) were constructed by cloning. Cell apoptosis induced by PEDV or overexpression non-structural or structural proteins was measured by the flow cytometry assay.ResultsPEDV could infect various host cells including Vero, Vero-E6 and Marc-145 and cause obvious cytopathic effects, including roundup, cell fusion, cell membrane vacuolation, syncytium formation and cause apparent apoptosis. In infected cells, PEDV-induced apoptosis is accompanied by nuclear concentration and fragmentation as a result of caspase-3 and caspase-8 activation and AIFM1 and PARP cleavage. Overexpression of S1 Spike protein of PEDV SM98 strain effectively induced host cell apoptosis, while the expression of the other non-structure proteins (Nsp1-16) and structural proteins (M, N, E, S2 and ORF3) has no or less effect on cell apoptosis. Similarly, expression of S1 protein from wild-type strain BJ2011 or cell-adapted strain CV777, also induce apoptosis in transfected cells. Finally, we demonstrated that the S1 proteins from various coronavirus family members such as TGEV, IBV, CCoV, SARS and MERS could also induce Vero-E6 cells apoptosis.ConclusionS1 Spike protein is one of the most critical functional proteins that contribute to cell apoptosis. Expression of S1 proteins of the coronavirus tested in this study could all induce cell apoptosis suggesting S1 maybe is an effective inducer in Coronavirus-induced cell apoptosis and targeting S1 protein expression probably is a promising strategy to inhibit coronavirus infection and thus mediated apoptosis on host cells.

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Isolation and characterization of porcine epidemic diarrhea virus with a novel continuous mutation in the S1⁰ domain.

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Isolation and characterization of porcine epidemic diarrhea virus with a novel continuous mutation in the S1<sup>0</sup> domain.

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Isolation and characterization of porcine epidemic diarrhea virus with a novel continuous mutation in the S10 domain.

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Isolation and characterization of porcine epidemic diarrhea virus with a novel continuous mutation in the S1<sup>0</sup> domain.

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OmicsDI is part of the ELIXIR infrastructure

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Isolation and characterization of porcine epidemic diarrhea virus with a novel continuous mutation in the S1⁰ domain.