Phenoloxidase Activity in Hemolymph of Naive andBeauveria bassiana-InfectedSpodoptera exiguaLarvae

Abstract

The majority ofSpodoptera exiguahemolymph phenoloxidase (PO) activity is localized in the hemocytes of naive larvae. Mid sixth instar larvae possessed 0.2±0.5 and 3.4±2.6 units (1 unit = Δ0.001 Å/min at 490 nm) of PO activity/μl of hemolymph equivalent in the plasma and hemocyte lysate (HL) fractions, respectively. TheS. exiguaHL PO had a pH optimum of 7.0 when assayed againstdl-Dopa. This enzyme is present in hemocytes as a proenzyme: activation of PO could be inhibited with either 10 mmdiisopropyl fluorophosphate or soybean trypsin inhibitor (1 mg/ml). The relative activity of HL PO was not inhibited by the addition of EGTA or EDTA. The PO titers in HL and plasma fractions sampled from sixth instar larvae during the feeding period (0–60 hr) of the sixth larval instar were not significantly different. However, a 10 × increase in PO activity was observed in plasma sampled from the wandering and prepupal stages. A series of assays was conducted to monitor the PO activity inBeauveria bassiana-infectedS. exigualarvae. At 24 hr postchallenge the PO titers in plasma and HL fractions from infected larvae were comparable to PO titers in fractions from saline controls. Analysis of PO activities revealed that by 48–60 hr postchallenge the PO titers in the HL sampled from infected larvae decreased sevenfold, whereas plasma PO titers increased. The increased titer of PO in the plasma did not inhibit fungal development.