Transmissible gastroenteritis virus (TGEV) is a porcine coronavirus. Lithium chloride (LiCl) has been found to be effective against several DNA viruses, such as Herpes simplex virus and vaccinia virus. Recently, we and others have reported the inhibitory effect of LiCl on avian infectious bronchitis coronavirus (IBV) infection, an RNA virus. In the current study, the action mechanism of LiCl on cell infection by TGEV was investigated. Plaque assays and 3-(4,5)-dimethylthiahiazo(-z-y1)-3,5-di-phenyl tetrazoliumbromide (MTT) assays showed that the cell infection by TGEV was inhibited in a dose-dependent manner, when LiCl was added to virus-infected cells; the cell infection was not affected when either cells or viruses were pretreated with the drug. The inhibition of TGEV infection in vitro by LiCl was observed at different virus doses and with different cell lines. The inhibitory effect of LiCl against TGEV infection and transcription was confirmed by RT-PCR and real-time PCR targeting viral S and 3CL-protease genes. The time-of-addition effect of the drug on TGEV infection indicated that LiCl acted on the initial and late stage of TGEV infection. The production of virus was not detected at 36 h post-infection due to the drug treatment. Moreover, immunofluorescence (IF) and flow cytometry analyses based on staining of Annexin V and propidium iodide staining of nuclei indicated that early and late cell apoptosis induced by TGEV was inhibited efficiently. The ability of LiCl to inhibit apoptosis was investigated by IF analysis of caspase-3 expression. Our data indicate that LiCl inhibits TGEV infection by exerting an anti-apoptotic effect. The inhibitory effect of LiCl was also observed with porcine epidemic diarrhea coronavirus. Together with other reports concerning the inhibitory effect of lithium salts on IBV in cell culture, our results indicate that LiCl may be a potent agent against porcine and avian coronaviruses.

The effects of glycyrrhizin diammonium (GD) and lithium chloride (LiCl) on cell infection by avian infectious bronchitis virus (IBV) were investigated using cytopathic effect observation, plaque-reduction assay and reverse transcriptase-polymerase chain reaction. The anti-viral effect of GD and LiCl on virus, on virus-infected cells or on cells pre-treated by both drugs was analysed, respectively. Our results showed that GD had a direct antiviral activity, leading to complete inhibition of cell infection. The cell infection was not alleviated by either pre-treatment of cells with GD or addition of the drug post infection, confirming that the inhibitory effect of GD, unlike LiCl, on IBV is a viral factor, rather than a cellular factor. The inhibitory effect of both drugs was confirmed by infecting primary chicken embryo kidney cells. In addition, apoptosis of infected cells was positively related with cytopathic effect and could be inhibited by effective drug treatment. Our data indicate that GD and LiCl have potential to prevent IBV infection in vitro through different antiviral mechanisms. The data are helpful for using antivirals efficiently.

Porcine transmissible gastroenteritis virus (TGEV) and porcine epidemic diarrhea virus (PDEV) can cause severe diarrhea in pigs. Development of effective vaccines against TGEV and PEDV is one of important prevention measures. The spike (S) protein is the surface glycoprotein of TGEV and PEDV, which can induce specific neutralization antibodies and is a candidate antigen for vaccination attempts. In this study, the open reading frames of the TGEV S1 protein and in addition of the S or S1 proteins of PEDV were inserted into the eukaryotic expression vector, pIRES, resulting in recombinant plasmids, pIRES-(TGEV-S1-PEDV-S1) and pIRES-(TGEV-S1-PEDV-S). Subsequently, 6–8 weeks old Kunming mice were inoculated with both DNA plasmids. Lymphocyte proliferation assay, virus neutralization assay, IFN-γ assay and CTL activity assay were performed. TGEV/PEDV specific antibody responses as well as kinetic changes of T lymphocyte subgroups of the immunized mice were analyzed. The results showed that the recombinant DNA plasmids increased the proliferation of T lymphocytes and the number of CD4+ and CD8+ T lymphocyte subgroups. In addition, the DNA vaccines induced a high level of IFN-γ in the immunized mice. The specific CTL activity in the pIRES-(TGEV-S1-PEDV-S) group became significant at 42 days post-immunization. At 35 days post-immunization, the recombinant DNA plasmids bearing full-length S genes of TGEV and PEDV stimulated higher levels of specific antibodies and neutralizing antibodies in immunized mice.

Avian infectious bronchitis virus (IBV), a coronavirus, causes infectious bronchitis leading to enormous economic loss in the poultry industry worldwide. Houttuynia cordata (Saururaceae) (HC) is a traditional Chinese medicine used in China. In the present study, the effect of HC on cell infection by IBV was determined using plaque assay and reverse transcription-polymerase chain reaction. The inhibitory effect of HC on IBV infection in ovo and in vivo was analysed using specific pathogen free (SPF) chicken embryos and chickens. Moreover, the effect of HC on cell apoptosis induced by IBV was investigated. Results showed that HC had more than 90% inhibition rate against IBV infection in Vero cells and chicken embryo kidney cells, and decreased more than 90% apoptotic cells caused by IBV. HC fully protected the SPF embryos, and had more than 50% protection rate in SPF chickens, against IBV challenge.