During investigations into an outbreak of egg production decline, retarded growth, and even death among ducks in Southeast China, a novel Tembusu virus strain named Tembusu virus Fengxian 2010 (FX2010) was isolated. This virus replicated in embryonated chicken eggs and caused embryo death. In cross-neutralization tests, antiserum to the partial E protein of Tembusu virus Mm1775 strain neutralized FX2010, whereas antiserum to Japanese encephalitis virus did not. FX2010 is an enveloped RNA virus of approximately 45-50 nm in diameter. Sequence analysis of its E and NS5 genes showed that both genes share up to 99.6% nucleotide sequence identity with Baiyangdian virus, and up to 88% nucleotide sequence identity with their counterparts in Tembusu virus. FX2010 was transmitted without mosquito, and caused systemic infection and lesions in experimentally infected ducks. These results indicate that FX2010 and BYD virus are newly emerged Tembusu virus strains that cause an infectious disease in ducks.

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A B S T R A C TIn recent years, the outbreaks of porcine epidemic diarrhea (PED) caused by the highly virulent porcine epidemic diarrhea virus (PEDV) variants occurred frequently in China, resulting in severe economic impacts to the pork industry. In this study, we selected and analyzed the genetic evolution of 15 PEDV representative strains that were identified in fecal samples of diarrheic piglets in 10 provinces and cities during 2011-2017. The phylogenetic analysis indicated that all the 15 PEDV isolates clustered into G2 genotype associated with the current circulating strains. Compared with the genome of the prototype strain CV777, these strains had 103-120 amino acid mutations in their S proteins, most of which were in the N terminal domain of S1 (S1-NTD). We also found 37 common mutations in all these 15 strains, although these strains shared 96.9-99.7% nucleotide homology and 96.3-99.8% amino acid homology in the S protein compared with the other original pandemic strains. Computational analysis showed that these mutations may lead to remarkable changes in the conformational structure and asparagine (N)-linked glycosylation sites of S1-NTD, which may be associated with the altered pathogenicity of these variant PEDV strains. We evaluated the pathogenicity of the PEDV strain FJzz1 in piglets through oral and intramuscular infection routes. Compared with oral infection, intramuscular infection could also cause typical clinical signs but with a slightly delayed onset, confirming that the variant PEDV isolate FJzz1 was highly pathogenic to suckling piglets. In conclusion, we analyzed the genetic variation and pathogenicity of the emerging PEDV isolates of China, indicating that G2 variant PEDV strains as the main prevalent strains that may mutate continually. This study shows the necessity of monitoring the molecular epidemiology and the etiological characteristics of the epidemic PEDV isolates, which may help better control the PED outbreaks. Stevenson et al., 2013;Vlasova et al., 2014). Subsequently, the outbreak of PED occurred in Okinawa, Japan in October 2013, followed by https://doi.

The spike (S) protein of severe acute respiratory syndrome-coronavirus (SARS-CoV) is a major virion structural protein. It plays an important role in interaction with receptor and inducing neutralizing antibodies. In the study, six tentative antigenic epitopes (S1 S2 S3 S4 S5 S6) of the spike protein of SARS-CoV were predicted by bio-informatics analysis, and a multi-epitope chimeric gene of S1-S2-S3-S4-S5-S6 was synthesized and fused to downstream GST gene in pGEX-6p-1. The Western blotting demonstrated that SARS patient convalescent serum could recognize the recombinant fusion protein. A number of monoclonal antibodies were developed against the fusion protein. In further, the six predicted epitope genes were individually fused to GST of pGEX-6p-1 and expressed in Escherichia coli BL21, respectively. Among six fusion peptides, S5 reacted with monoclonal antibody D3C5 and S2 reacted with monoclonal antibody D3D1 against spike protein of SARS-CoV. The epitopes recognized by monoclonal antibodies D3C5 and D3D1 are linear, and correspond to 447-458 and 789-799 amino acids of spike protein of SARS-CoV, respectively. Identification of antigenic epitope of spike protein of SARS-CoV could provide the basis for the development of immunity-based prophylactic, therapeutic, and diagnostic techniques for the control of severe acute respiratory syndrome.

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Porcine epidemic diarrhea virus (PEDV) is a highly contagious coronavirus that causes severe diarrhea and death, particularly in neonatal piglets. The nucleocapsid protein (N protein) of PEDV presents strong immunogenicity and contributes to the cross-reactivity between PEDV and TGEV. However, the characterization of epitopes on the PEDV N protein remains largely unknown. Here, two monoclonal antibodies (MAbs) specific to the N protein of a PEDV strain, FJzz1/2011, were generated and screened against a partially overlapping library of 24 GST-fusion N protein-truncated constructs. We confirmed that residues 18–133 (designated NEP-D4) and residues 252–262 (designated NEP-D6) were the epitopes targeted by MAbs PN-D4 and PN-D6, respectively. Sequence analysis revealed that these two epitopes were highly conserved among PEDV strains but were significantly different from other members of the Coronavirinae subfamily. Western blot analysis showed that they could be specifically recognized by PEDV antisera but could not be recognized by TGEV hyperimmune antisera. Indirect immunofluorescence (IFA) assays confirmed no cross-reaction between these two MAbs and TGEV. In addition, the freeze-thaw cycle and protease treatment results indicated that NEP-D4 was intrinsically disordered. All these results suggest that these two novel epitopes and their cognate MAbs could serve as the basis for the development of precise diagnostic assays for PEDV.

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