TWENTY-FIFTH ANNUAL EASTERN FISH HEALTH WORKSHOP MARCH 10-13, 2000     Biochemical Characterization Of Two Perkinsus Species Isolated From The Softshell Clam, Mya arenaria       Shawn M. McLaughlin1 and M. Faisal2     1NOAA, National Ocean Service, Center for Coastal Environmental Health and Biomolecular Research, Cooperative Oxford Laboratory, 904 S. Morris St., Oxford, MD  21654; 2Virginia Institute of Marine Science, School of Marine Science, The College of William and Mary, Gloucester Point, VA  23062       The ability of pathogenic protozoa to damage tissue depends, at least partially, upon the production of lytic enzymes and adhesion molecules. Previous studies demonstrated that Perkinsus marinus found in the eastern oyster, Crassostrea virginica, depends in its pathogenicity on a number of extracellular proteins (ECP) including highly potent serine proteases. Two species of Perkinsus were recently isolated and propagated in vitro from hemolymph (H-49) and gill tissue (G-117) of the softshell clam, Mya arenaria. Morphological, cultural, and genetic characteristics examined during previous investigations identified H-49 as P. marinus and G-117 as a new species, Perkinsus chesapeaki sp. nov. In this study, the biochemical characterization of ECP produced in vitro by the H-49 and G-117 isolates was performed and compared to that of the oyster-derived Perkinsus marinus isolate P-1.  The G-117 and H-49 isolates demonstrated distinct differences in enzyme activities; however, all three isolates shared common bands. Further examination using substrate-impregnated gels showed H-49 to possess proteolytic activities while G-117 did not. The lack of proteolytic activity in G-117 ECP provides additional evidence of differences between the G-117 and H-49 isolates, both isolated from softshell clams. Inhibition studies revealed that H-49 ECP contain serine proteases similar to those described for the P. marinus P-1 isolate. The G-117 ECP lacked proteolytic activity but showed a higher production of lipolytic enzymes than H-49 or P-1. Differences in the electrophoretic patterns of the three isolates are most likely to be associated with virulence factors and host specificity.  The effects of temperature and salinity on in vitro growth of G-117 and H-49 were also examined. Optimal growth temperatures for the two clam isolates were, in general, lower than P-1. G-117 showed faster growth at lower salinities than either H-49 or P-1. Clam isolates appear to be more adapted to lower salinities and temperatures than P. marinus of the eastern oyster. The reduced ability of G-117 to grow at higher salinities may impinge on its geographical distribution and host range.     Return to 25th Annual Eastern Fish Health Workshop Return to Leetown Science Center Home Page