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 The clotting ability of the hemolymph withdrawn from moribund prawns pre-injected with the bacteria, ECP,
cysteine protease of PBS (control) was observed for 2 h at 25 C. Of these, only the control group was clottable while all the other groups were unclottable. A component of the plasma, previously identified as
coagulogen-like
protein, was further confirmed to be a
coagulogen by the comparison of plasma with serum on non-reduced SDS-PAGE or using rabbit antiserum to the
coagulogen-like
protein (R alpha
coagulogen) to neutralize the clotting ability of normal prawn hemolymph. The
coagulogen was reduced in amount in plasma of moribund prawns after injection with the bacteria, ECP or
cysteine protease while it apparently disappeared after pre-incubation with the ECP or
cysteine protease for 2 h at 25 C compared with normal prawn plasma as observed in crossed immunoelectrophoresis (CIE)
gels. The reduction of the amount of
coagulogen in plasma of moribund prawns was also evident in CIE
gels using R alpha
coagulogen. In addition, the apparent disapperance of the
coagulogen mentioned above was eventually proven to be due to the change of its migration rate in CIE
gels after pre-incubation with ECP or
cysteine protease, since the disappeared
coagulogen arc (
arc 2) (migrated into
arc 1) could be visualized by using R alpha
coagulogen or by reducing the time for pre-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 with hemostasis leading to the occurrence of unclottable hemolymph. These complex events may significantly contribute to the pathogenicity of V. harveyi in the prawn.