Abstract
A highly virulent porcine epidemic diarrhea virus (PEDV) appeared in China and spread rapidly to neighbor countries, which have led to great economic losses to the pig industry. In the present study, we isolated a PEDV using Vero cells and serially propagated 100 passages. PEDV SDSX16 was characterized in vitro and in vivo. The viral titers increased to 107.6 TCID50/mL (100th) by serial passages. The spike (S) gene and the whole gene of the SDSX16 virus was fully sequenced to assess the genetic stability and relatedness to previously identified PEDV. Along with successive passage in vitro, there were 18 nucleotides (nt) deletion occurred in the spike (S) gene resulting in a deletion of six amino acids when the SDSX16 strain was passaged to the 64th generation, and this deletion was stable until the P100. However, the ORF1a/b, M, N, E, and ORF3 genes had only a few point mutations in amino acids and no deletions. According to growth kinetics experiments, the SDSX16 deletion strain significantly enhanced its replication in Vero cells since it was passaged to the 64th generation. The animal studies showed that PEDV SDSX16-P10 caused more severe diarrhea and vomiting, fecal shedding, and acute atrophic enteritis than SDSX16-P75, indicating that SDSX16-P10 is enteropathogenic in the natural host, and the pathogenicity of SDSX16 decreased with successive passage in vitro. However, SDSX16-P10 was found to cause lower levels of cytokine expression than SDSX16-P75 using real-time PCR and flow cytometry, such as IL1β, IL6, IFN-β, TNF-α, indicating that SDSX16-P10 might inhibit the expression of cytokines. Our data indicated that successive passage in vitro resulted in virulent attenuation in vivo of the PEDV variant strain SDSX16.
Highlights
Porcine epidemic diarrhea (PED) is a highly contagious swine disease
The animal studies showed that porcine epidemic diarrhea virus (PEDV) SDSX16-P10 caused more severe diarrhea and vomiting, fecal shedding, and acute atrophic enteritis than SDSX16-P75, indicating that SDSX16-P10 is enteropathogenic in the natural host, and the pathogenicity of SDSX16 decreased with successive passage in vitro
The isolated PEDV strain SDSX16 was serially passaged in Vero cells for a total of 100 passages (P1 to P100)
Summary
Porcine epidemic diarrhea (PED) is a highly contagious swine disease. The main clinical signs include severe vomiting, diarrhea, and dehydration in suckling piglets [1,2,3]. The most of sow herds had immunized with a CV777-based vaccine, large-scale outbreaks of PED were still reported in China since late 2010. It resulted in high rates of morbidity and mortality in suckling piglets and substantial economic losses in the country. Both modified live vaccines and killed vaccines were commercially available in China, but the current vaccines were less effective against the PEDV variants, and the PED epidemics were not properly controlled. The need for the development of better vaccines against the novel
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