Porcine deltacoronavirus enters ST cells by clathrin-mediated endocytosis and does not require Rab5, Rab7, or Rab11

Porcine deltacoronavirus (PDCoV) is a newly emerged enteric pathogen that causes diarrhea, dehydration, vomiting, and in suckling piglets, a high rate of death. Here, we used endocytosis inhibitors, dominant-negative mutants, and small interfering RNA (siRNA) interference to investigate PDCoV entry into swine testicle (ST) cells. Results showed that chlorpromazine, dynasore, and methyl-beta-cyclodextrin (M beta CD) inhibited PDCoV internalization; immunofluorescence assay (IFA) of dominant-negative (DN) mutants and siRNA interference further demonstrated these results, indicating that PDCoV utilizes clathrin-mediated endocytosis and requires dynamin and cholesterol to enter ST cells. The role of endosomes in PDCoV infection was also investigated and we found that PDCoV does not require Rab5, Rab7, or Rab11 for productive infection, indicating that the transport of PDCoV in ST cells does not require the participation of the endosomal system. Our findings will provide additional understanding of the entry mechanisms of PDCoV and possible antiviral targets.IMPORTANCEPorcine deltacoronavirus (PDCoV) is a newly emerged enteric virus threatening pig industries worldwide. Our previous work showed that PDCoV enters porcine kidney (PK-15) cells through a caveolae-dependent pathway, but the entry mechanism for PDCoV into swine testicle (ST) cells remains unclear. Mechanisms of virus entry can be different with different virus isolates and cell types. Here, we determined that PDCoV enters ST cells via clathrin-mediated endocytosis. Additionally, we found that PDCoV entry does not require Rab5, Rab7, or Rab11. These findings provide additional understanding of the entry mechanisms of PDCoV and possible antiviral targets. Porcine deltacoronavirus (PDCoV) is a newly emerged enteric virus threatening pig industries worldwide. Our previous work showed that PDCoV enters porcine kidney (PK-15) cells through a caveolae-dependent pathway, but the entry mechanism for PDCoV into swine testicle (ST) cells remains unclear. Mechanisms of virus entry can be different with different virus isolates and cell types. Here, we determined that PDCoV enters ST cells via clathrin-mediated endocytosis. Additionally, we found that PDCoV entry does not require Rab5, Rab7, or Rab11. These findings provide additional understanding of the entry mechanisms of PDCoV and possible antiviral targets.