Abstract
BackgroundPorcine Epidemic Diarrhea Virus (PEDV) is a coronavirus that infects the intestinal tract and causes diarrhea and vomiting in older pigs or extreme dehydration and death that could reach 100% mortality in neonatal piglets. In the US, the first PEDV outbreaks occurred in 2013 and since then US PEDV strains have quickly spread throughout the US and worldwide, causing significant economic and public health concerns. Currently two conditionally approved vaccines exist in the US, but there is no live attenuated vaccine, which is considered the best option in controlling PEDV by inducing transferrable mucosal immunity to susceptible neonatal piglets. In this study, we passaged an US PEDV isolate under various conditions to generate three strains and characterized their growth and antigenicity in cell culture using various assays including Western blot analysis, serum neutralization assay, sequencing analysis and confocal microscopy. Finally, these strains were evaluated for pathogenicity in nursing piglets (1–4 days old).ResultsOne of the PEDV strains generated in this study (designated as PEDV 8aa) is able to replicate in cells without any protease and grows to a high titer of >8 log10 TCID50/ml in cell culture. Interestingly, replication of PEDV 8aa was severely reduced by trypsin and this correlated with the inhibition of virus attachment and entry into the cells. In neonatal nursing piglets, PEDV 8aa (passage number 70 or 105) was found to be fully attenuated with limited virus shedding.ConclusionsThese results suggest that applying selective pressure during viral passages can facilitate attainment of viral attenuation and that PEDV 8aa warrants further investigation as an attenuated vaccine.
Highlights
Porcine Epidemic Diarrhea Virus (PEDV) is a coronavirus that infects the intestinal tract and causes diarrhea and vomiting in older pigs or extreme dehydration and death that could reach 100% mortality in neonatal piglets
The replication of PEDV 8aa was severely reduced by trypsin and this correlated with the inhibition of virus attachment and entry into the cells
Cell culture adaptation of an US PEDV strain Initial attempt to isolate the US PEDV strain required 2 or 3 blind passages with trypsin (1–2 μg/ml) until apparent Cytopathic effect (CPE) appeared in Vero cells
Summary
Porcine Epidemic Diarrhea Virus (PEDV) is a coronavirus that infects the intestinal tract and causes diarrhea and vomiting in older pigs or extreme dehydration and death that could reach 100% mortality in neonatal piglets. In the US, the first PEDV outbreaks occurred in 2013 and since US PEDV strains have quickly spread throughout the US and worldwide, causing significant economic and public health concerns. We passaged an US PEDV isolate under various conditions to generate three strains and characterized their growth and antigenicity in cell culture using various assays including Western blot analysis, serum neutralization assay, sequencing analysis and confocal microscopy These strains were evaluated for pathogenicity in nursing piglets (1–4 days old). The US PEDV strains were reported to have caused outbreaks in Asian and European countries [14,15,16,17,18,19,20,21,22,23], raising significant economic and public health concerns worldwide [24, 25]. Two conditionally approved vaccines exist in the US: alphavirusbased vaccine (Harrisvacccines) and an inactivated vaccine
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