Engineering a TGEV S-trimer chimera with PEDV D0-NTD generates potent neutralizing antibodies against both viruses

Porcine epidemic diarrhea virus (PEDV) and transmissible gastroenteritis virus (TGEV), two economically devastating swine enteric coronaviruses, persistently threaten global pork production via severe clinical morbidity. Vaccination is the most effective means to prevent morbidity and mortality caused by porcine enteric coronavirus infection. Hence, the development of an effective and safe vaccination strategy to protect against both PEDV and TGEV infections is urgently needed. Here, we designed and produced trimerized full-length PEDV and TGEV spike (S) proteins using an efficient mammalian expression vector system in HEK293F cells and reported that compared with the PEDV spike protein (2⁷), the TGEV spike protein induces significantly more potent neutralizing antibodies (2¹²). Moreover, transmission electron microscopy (TEM) with negative staining revealed that the TGEV spike protein exhibited excellent homogeneity. We subsequently used the TGEV S protein trimer as the backbone to display different domains of the PEDV S protein and successfully constructed seven spike protein chimeras. We further screened two chimeric S proteins through expression and mouse experiments, which included TGEV S-PEDV(D0/NTD) and TGEV S-PEDV(D0/NTD/CTD), in which the S protein of TGEV was replaced by the PEDV S D0-NTD and D0-NTD-CTD, respectively. Evaluation of the immunogenicity and efficacy of the chimeric S proteins in piglets confirmed that piglets in the TGEV S-PEDV(D0/NTD)-immunized group generated high levels of neutralizing antibodies (2^13.6 against TGEV; 2^5.8 against PEDV) against both PEDV and TGEV. Our findings suggest that the TGEV S-PEDV(D0/NTD) is a promising candidate for combating both PEDV and TGEV infections.IMPORTANCEThe design strategy of multivalent and multitarget single antigens facilitates the development of vaccines targeting PEDV and TGEV. Optimizing the presentation of PEDV core antigen epitopes on the basis of the S protein trimer structure will provide novel insights into the development of subunit vaccines. Here, we compared the immunogenicities of the TGEV S protein and PEDV S protein and then designed a bivalent chimeric S protein candidate, TGEV S-PEDV(D0/NTD), in which the corresponding segments on the TGEV S protein were replaced with D0-NTD domains from the PEDV S protein. We demonstrated that the TGEV S protein exhibits greater immunogenicity than the PEDV S protein and that TGEV S-PEDV(D0/NTD) induces broad-spectrum neutralization protection against PEDV and TGEV. Our results demonstrate the effectiveness of the chimeric S protein and provide a feasible method for the development of efficient bivalent subunit vaccines against PEDV and TGEV.