Porcine reproductive and respiratory syndrome virus enters cells through a low pH-dependent endocytic pathway

The entry pathway of porcine reproductive and respiratory syndrome virus (PRRSV) into MARC-145 cells was investigated using a variety of drugs that interfere with the pH of intracellular vesicles by different mechanisms. Virus entry was assessed by measuring viral RNA replication or production of infectious virus. Chloroquine, ammonium chloride and bafilomycin A1 inhibited RNA replication or production of infectious virus in a dose-dependent manner. These drugs inhibited virus replication when added to the cells prior to, at infection or soon after infection. Moreover, the effect of chloroquine on PRRSV replication was reversible under acidic conditions of the media. Taken together, these results indicated that a low pH was required during virus entry. Electron microscopic data showed virus particles at the cell surface or within small vesicles which were circumscribed by a clathrin-like zone. In addition, the number of PRRSV-infected cells was decreased in the presence of cytochalasin B and phenylarsine oxide. Thus, we concluded that PRRSV entry might occur through a microfilament-dependent endocytic mechanism in which a low pH is necessary for proper virus uncoating.