Optimization of the isolation of entomopathogenic fungi Metarhizium spp. associated with the rhizosphere and insects in western Burkina Faso in the context of malaria control

Abstract

Malaria is the leading cause of consultations, hospitalizations, and deaths in Africa. Given the decreasing effectiveness of current control tools, the World Health Organization (WHO) encourages the exploration of alternative methods, such as biological control using microorganisms, parasites, and natural predators of mosquitoes. Entomopathogenic fungi of the genus Metarhizium spp. stand out for their promising laboratory results (BILGO et al., 2018). Metarhizium spp. exhibit a dual lifestyle: as insect pathogens and as plant endophytes. However, isolation techniques for these fungi pose challenges, particularly in selecting optimal culture media. This study aims to optimize the isolation techniques for Metarhizium fungi associated with the rhizosphere and insects in western Burkina Faso. A total of 565 rhizosphere and insect samples were collected in Bama (11°23'N, 4°24'W), a ricegrowing area, and Soumousso (11°04'N, 4°03'W), a savanna and maize cultivation area, both located in western Burkina Faso. These samples were cultured on two selective media enriched with antibiotics based on Potato Dextrose Agar (PDA): “PDA + Penicillin + Ampicillin” and “PDA + Gentamicin + Ampicillin.” The cultures were incubated at 27°C ± 2°C with 70% humidity for 21 days. After macroscopic and microscopic identification, suspect colonies underwent amplification of the Internal Transcribed Spacer (ITS) region by PCR for confirmation. Among the media tested,

PDA + Gentamicin + Ampicillin proved most effective for isolating Metarhizium. Based on macroscopic and microscopic characteristics, 48 and 36 Metarhizium spp. strains were identified from the rhizosphere and insects, respectively. A subsample of 50 strains, for which the ITS region was amplified, was used to identify 15 Metarhizium strains associated with insects and 21 strains associated with the rhizosphere.