Bacteriological agar

Conidia removal by the insect was determined after exposing Xyleborus affinis females to B. bassiana CHE-CNRCB 44 strain at a concentration of 1 × 10⁹ conidia mL⁻¹. After exposure, beetles were transferred to 2 mL Eppendorf tubes with diet for 12 h. For the evaluation at time 0, insects were immediately placed in tubes without diet. Insects were removed from tubes with or without diet and individually transferred to 1.5 mL Eppendorf tubes containing 150 μL of 0.05% Tween 80. Tubes were then gently shaken for one minute and 10 μL was diluted to 1:1000, after which 30 µL was placed in a selective medium for B. bassiana (SMBb) (39 g L⁻¹ potato dextrose agar; BD Bioxon, Guadalajara, Mexico), supplemented with 2.5 g L⁻¹ bacteriological agar (BD Bioxon Mexico), 80 mg L⁻¹ tetracycline (Sigma-Aldrich, St. Louis, MO, USA), 25 mg L⁻¹ cycloheximide (Sigma-Aldrich), 80 mg L⁻¹ streptomycin (Sigma-Aldrich), 80 mg L⁻¹ penicillin (Sigma-Aldrich), and 1 mg L⁻¹ benomyl (Robust R; Promotora Técnica Industrial, Morelos, Mexico) [26 (link),27 (link),28 (link)]. The inoculum was distributed using Drigalski rods, and Petri dishes were incubated at 25 °C ± 2 °C for 72 h, after which colony-forming units (CFU) were determined. A control group was kept for 12 h in a humid chamber after exposure. All bioassays were performed in triplicate.