Enzyme production by the entomopathogenic fungi Beauveria bassiana and Metarhizium anisopliae and their application in the control of nematodes (Haemonchus spp. and Meloidogyne incognita) in vitro

Brazil is affected by the infection of gastrointestinal nematodes (Haemonchus spp.) because it causes subclinical diseases in small ruminants that directly affect weight and milk production and, in turn, generate a health risk for the animals. In the same way root-knot nematodes (Meloidogyne incognita) is a serious disease, which parasitize the roots of tomato plants causing damage such as poor nutrient absorption leading to significant yield losses. In order to minimize the economic impact of these nematodes, it is important to establish new control strategies. Beauveria bassiana and Metarhizium anisopliae are two main mycoinsecticides used to control many orders of insects, such as Lepidoptera and Hemiptera. These fungi have the ability to produce extracellular enzymes, which play an important role to control in the pest infection process. The aim of this study was to evaluate the production of proteases and chitinases by the entomopathogenic fungi B. bassiana and M. anisopliae on different solid and liquid culture media and their application in vitro to the control of Haemonchus spp and Meloidogyne incognita. To achieve this goal, enzymes were produced under solid and liquid fermentation conditions to determine the highest chitinolytic and proteolytic activity of commercial two strain of B. bassiana (IBCB 66 and ESALQ PL63) and M. anisopliae (IBCB 425 and ESALQ E9) in liquid media: SDY broth, YPG culture medium, synthetic, and soluble starch with yeast extract. The solid media tested were: rice supplemented with whey and chrysalis flour mixed with rice. In addition, the nematicidal action of their extracts (containing concentrated enzymes and without fungal cells) was measured. The results showed that the best culture medium (p< 0.01) for protease was solid serum rice medium, with a value for M. anisopliae IBCB 425 of 52 U/mg, and for B. bassiana ESALQ PL63 the value was 36 U/mg, compared to all the tested media. On the other hand, for chitinase of M. anisopliae strain IBCB 425, the activity value was close to 0.60 U/mg. In contrast, for B. bassiana isolate ESALQ PL63, SDY medium was the best inducer for chitinase production, with a value of 0.90 U/mg. Regarding the nematicidal activity of the crude extracts, the reduction percentages were 58% for ESALQ PL63 and 100% for IBCB 425 in the case of animal parasites. For plant parasites, the reduction percentages were 19% for ESALQ PL63 and 71% for IBCB 425. Thus, the use of enzyme-rich crude extracts presents promising control options for pest control.