Using the entomopathogenic fungus, Metarhizium anisopliae, to control the rhinoceros beetle, Oryctes rhinoceros, was first attempted in 1976. Early screening showed M. anisopliae variety major to be highly pathogenic to O. rhinoceros. Further work concentrated on using the variety to control O. rhinoceros as the pest infestation increased in oil palm plantations. Assessment of isolates from different localities found little variation in pathogenicity between them. Dipping the larvae in a spore solution of 10 8 spores ml- 1 caused 100% mortality after 12 and 14 days. Analysis of the fungal DNA by RAPD-PCR showed a high similarity in the genetic base among isolates. Two primers can be used for fingerprinting and as a post-release monitoring tool as they were able to differentiate the isolates SE and BP from the others. An initial field study applying the fungus as spore solutions and sporulated substrates infected all stages of the pest, causing up to 84% reduction in its overall population.
The effects of the fungus on the oil palm pollinating weevil and non-target organisms were studied. The fungus did not affect development of the weevil. Toxicity tests showed the fungus to be harmless to rats. Fish exposed to very high spore concentrations of 1000 mg ml- 1 (eight times higher than the highest rate applied in the field) only caused 25% mortality and at 2000 mg ml- 1 only 40% mortality. The larvae of the stag beetle, Aegus chelifer, were susceptible to M. anisopliae, although less so than the larvae of O. rhinoceros. At 12 days after treatment, all the tested isolates caused 33.3% – 83.3% mortality to the stag beetle larvae but killed all (100%) of the O. rhinoceros larvae.
The spores of M. anisopliae were successfully mass produced using solid state fermentation. Fungal mycelia were first produced in liquid medium and then sporulated on a solid medium of maize. Harvesting was done by separating out the spores from the maize by washing in water, collecting them by vacuum filtration and drying at low temperature before finally grinding to powder. The yield of spores was 9.2-10.5 g per 200 g maize bag with a viability of > 80%. The powder formulation was tested in the field, pre-mixed with water and applied to rotting oil palm debris by spraying. M. anisopliae infected O. rhinoceros in all stages of its life cycle. Application to rotting debris reduced the O. rhinoceros population by up to 80%. The field application of M. anisopliae did not affect the populations of oil palm pollinating weevil and stag beetle.