Our findings could encourage further investigation and development of M. anisopliae isolate MAX-2, and attract research interest on the stress tolerance of biocontrol fungi. Methods Solid substrates
Wheat bran substrates with different moisture levels were used in this study. The substrates were sterilized at 121°C for 20 min. Sterile wheat bran without water was used selleck chemical as a dry substrate to test the efficacy of M. anisopliae under desiccation stress. The moisture contents of substrates were adjusted by adding a certain amount of water and heating 5 g of the sterilized substrate at 100°C for 4 h. Moisture content was then calculated using the dry and initial weights. Moisture content of the dry substrate was determined to be 8%. The gradient of the substrates from the initial moisture content was adjusted to 15%, 20%, 25%, 30%, and 35%. Sterile culture of host insects T. molitor larvae were selected as host insects because they can remain active under desiccation stress, and are easily reared under laboratory conditions. Such
conditions are convenient for testing the virulence of fungal pathogens under desiccation stress. To eliminate the effect of some possible microbes, we cultured the host insects under sterile conditions. T. molitor larvae were washed in sterile water, and the water on the surface was absorbed using sterile filter papers. The cuticles of the larvae were wiped carefully with 75% OSI-906 purchase alcohol cotton balls for seconds and transferred to sterile
filter Pexidartinib in vivo paper to dry in air for 5 min. Sterilized larvae were reared, incubated, and subcultured in sterile glass jars containing the wheat bran substrate with 15% moisture content. Screening of GNE-0877 MAX-2 with the capacity of infecting under desiccation stress M. anisopliae isolates in the experiment M. anisopliae isolates were collected from the arid regions of Yunnan Province in China during the dry season. The efficacy test was conducted in the wet substrate with 30% moisture content at 25°C. The isolates MAC-6, MAL-1, and MAQ-28, whose efficacies showed gradient descent, were chosen as controls to display the efficacy of MAX-2 under desiccation stress. The MAX-2 isolate was from Shangri-la, MAC-6 was from Chuxiong, MAL-1 was from Lanping, and MAQ-28 was from Qujing. Conidial production and inoculation The conidia of M. anisopliae isolates were produced by incubating the fungi on potato dextrose agar plates at 25°C for 14 d. Conidia powder of MAX-2 was obtained from the surface of fungal colonies using a sterile scoop and transferred to a sterile tube (20 mm?×?200 mm). Conidial powder was weighed and mixed with sterile wheat bran substrates. The conidial concentration was adjusted to 5?×?108 conidia/g, and the substrates were cultured at 25°C. The conidial concentration was controlled by adjusting the amount of conidial powder in the substrate, and determined by diluting 1 g of the mixture (conidial powder and substrate) with sterile water.