Hemostasis of tiger prawn Penaeus monodon affected by Vibrio harveyi, extracellular products and a toxic cysteine protease

The effects of bacterial cells, extracellular products (ECP) and a purified cysteine protease of Vibrio harveyi on hemostasis and plasma components of tiger prawn (Penaeus monodon) were studied. The clotting ability of the he molymph withdrawn from moribund prawns pre-injected with the bacteria, ECP, cysteine protease or PBS (control) was observed for 2 h at 25 C. Of these, only the control group was clottable while all the other groups were unclo ttable. A component of the plasma, previously identified as coagulogen-like protein, was further confirmed to be a coagulogen by the comparison of pla sma with serum on non-reduced SDS-PAGE or using rabbit antiserum to the coa gulogen-like protein (R alpha coagulogen) to neutralize the clotting abilit y of normal prawn hemolymph. The coagulogen was reduced in amount in plasma of moribund prawns after injection with the bacteria, ECP or cysteine prot ease while it apparently disappeared after pre-incubation with the ECP or c ysteine protease for 2 h at 25 C compared with normal prawn plasma as obser ved in crossed immunoelectrophoresis (CIE) gels. The reduction of the amoun t of coagulogen in plasma of moribund prawns was also evident in CIE gels u sing R alpha coagulogen. In addition, the apparent disappearance of the coa gulogen mentioned above was eventually proven to be due to the change of it s migration rate in CIE gels after pre-incubation with ECP or cysteine prot ease, since the disappeared coagulogen are tare 2) (migrated into arc 1) co uld be visualized by using R alpha coagulogen or by reducing the time for p re-incubation from 2 h to 30 min. Thus, the effects of cysteine protease on plasma coagulogen observed in vitro and in vivo may markedly interfere wit h hemostasis leading to the occurrence of unclottable hemolymph. These comp lex events may significantly contribute to the pathogenicity of V. harveyi in the prawn.