In comparison, inoculation with iPEDVPT-P5-96S and iPEDVPT-P96 viruses containing the S gene derived from the attenuated iPEDVPT-P96, induced a delayed onset of clinical symptoms and peak viral shedding. Histological evaluation of piglets inoculated with both viruses (iPEDVPT-P96, n = 3; iPEDVPT-P5-96S, n = 3) at 3 DPI revealed a much milder degree of villous atrophy in the jejunum with conspicuous villous hyperplasia compared to the other two groups. Notably, iPEDVPT-P5-96S-treated piglets exhibited statistically significant severer villous atrophy in jejunum than iPEDVPT-P96-treated piglets (Figure 3). Despite the similar pattern and severity of viral shedding and clinical symptoms, inoculation with iPEDVPT-P5-96S eventually resulted in a higher mortality rate (40% in iPEDVPT-P96 and 80 % and iPEDVPT-P5-96S) and lower average daily weight gain by 10 DPI than those upon inoculation with iPEDVPT-P96. These data suggested that iPEDVPT-P96 fully regained virulence by replacement of the S gene from the virulent PEDVPT 52 strains and the complementary approach can only partially reduce the virulence of the iPEDVPT-P5.