Serine proteases activity is important for the interaction of nematophagous fungus Duddingtonia flagrans with infective larvae of trichostrongylides and free-living nematodes Panagrellus spp.

The nematode-trapping fungus Duddingtonia flagrans has been studied as a possible control method for gastrointestinal nematodes of livestock animals. These fungi capture and infect the nematode by cuticle penetration, immobilization, and digestion of the internal contents. It has been suggested that this sequence of events occurs by a combination of physical and enzymatical activities. The aim of this study was to investigate the participation of proteolytic enzymatic activity during the interaction of the nematophagous fungus D. flagrans with infective larvae of trichostrongylides and the free-living nematode Panagrellus spp. Protease inhibitors used interfered in the predatory activity of D. flagrans. However, only PMSF significantly reduced the mean number of Panagrellus spp. captured by D. flagrans in comparison with the control. The experiment with fluorogenic substrate showed that maximum urokinase activity during the interaction of the fungus with the infective larvae of trichostrongylides or Panagrellus spp. occurred within 7 or 1 h of incubation, respectively. The protease activity, especially of the serine class, may be important during the interaction between the fungus and nematodes.

Optimizing protease production from an isolate of the nematophagous fungus Duddingtonia flagrans using response surface methodology and its larvicidal activity on horse cyathostomins.

Protease production from Duddingtonia flagrans (isolate AC001) was optimized and the larvicidal activity of the enzymatic extract was evaluated on infective horse cyathostomin larvae (L3). Duddingtonia flagrans was grown in liquid medium with eight different variables: glucose, casein, bibasic potassium phosphate (K2HPO4), magnesium sulphate (MgSO4), zinc sulphate (ZnSO4), ferrous sulphate (FeSO4), copper sulphate (CuSO4) and temperature. The Plackett-Burman analysis showed a significant influence of MgSO4, CuSO4 and casein (P < 0.05) on protease production by D. flagrans in liquid medium. Central composite design indicated that the highest proteolytic activity was 39.56 U/ml as a function of the concentrations of casein (18.409 g/l), MgSO4 (0.10 g/l) and CuSO4 (0.50 mg/l). A significant difference (P < 0.01) was found for the larval number between the treated and control groups at the end of the experiment. A reduction of 95.46% in the number of free-living larvae was found in the treated group compared with the control. The results of this study suggest that protease production by D. flagrans (AC001) in liquid medium was optimized by MgSO4, CuSO4 and casein, showing that the optimized enzymatic extract exerted larvicidal activity on cyathostomins and therefore may contribute to large-scale industrial production.