Comparison of in vitro-cultured and wild-type Perkinsus marinus. I. Pathogen virulence.

Perkinsus marinus is a highly contagious pathogen of the eastern oyster Crassostrea virginica. Until recently, transmission studies have employed wild-type parasites isolated directly from infected oysters. Newly developed methods to propagate P. marinus in vitro have led to using cultured parasites for infection studies, but results suggest that cultured parasites are less virulent than wild-type parasites In this paper, we report results of experiments designed to quantify differences between wild-type and cultured P. marinus virulence and to test the following hypotheses: (1) in vitro-cultured parasites are less virulent than wild-type parasites; (2) virulence decreases gradually during in vitro culture; (3) virulence of in vitro cultures can be restored by in vivo passage; (4) virulence changes with culture phase. Our results demonstrate that parasites freshly isolated from infected hosts are much more virulent than those propagated in culture, indicating a potential deficiency in the culture medium used. Virulence was lost immediately in culture and, for that reason, the practice of repassing cultured cells through the host to restore virulence does not work for P. marinus. Virulence was also associated with culture phase: log-phase parasites were significantly more virulent than those obtained from lag- or stationary-phase cultures.