5, 1, 1.5, 2, or 2.5 hours. For the dry-heat shock test, conidia were dried in a desiccator containing silica gel until the moisture content was less than 5%. Dried conidia were maintained in an incubator oven at 65°C for 1, 2, 3, 4, or 5 hours, and then suspended in sterilized water (1 × 107 conidia·mL-1). The conidial suspensions maintained at 28°C were used as a control. Germinations were measured by plating 50 μL on 1/4SDA plates. After 24 hours incubation in the dark at 28°C, the find more germination rate

was checked with a microscope (Motic, china) CP-690550 in vitro at 400× magnification. About 300 conidia were evaluated for germination from different areas in each plate. Inhibition time values for 50% germination (IT50) were used to estimate the conidiospore thermotolerance

of M. acridum using DPS software [49]. Bioassays Locusta migratoria were reared in our lab under crowded conditions as previously described by He et al. [50]. Male and female insects were separated after adult emergence. Male adult locusts (2-3 days after eclosion) were used in the bioassay tests. A 5-μL solution of 2 × 106 conidia/mL of either wild-type M. acridum or transformants in cottonseed oil (Sigma) was applied to the locusts’ head-thorax junctions. Treated locusts were separately confined in cages (20 × 20 × 20 cm) by 40 mesh, and kept at a temperature of 28°C RG7112 chemical structure with a 16:8 h (light:day) photoperiod. Mannose-binding protein-associated serine protease There were four replications of n = 30 locusts in each treatment. Mortality was recorded daily and lethal time values for 50% mortality (LT50) values were used to estimate the infectivity of M. acridum by DPS software [49]. Statistical analysis All samples and treatments were carried out in triplicate unless stated otherwise. Data were square root arcsine transformed before being subjected to analysis of variance (ANOVA) for a completely randomized design. The means were separated

using Tukey’s multiple range test, carried out using DPS software [47]. Statistical significance was established at p < 0.05. Acknowledgements The research was supported by grants from the Natural Science Foundation of China (No. 30170630), and the Natural Science Foundation of Chongqing Sci-Tech Commission, P. R. China (No. 2008BB1178). References 1. Charnley AK, Collins SA: Entomopathogenic fungi and their role in pest control. Mycota: Environmental and Microbial Relationships 2007, 4:159–187.CrossRef 2. Lomer C, Bateman R, Johnson D, Langewald J, Thomas M: Biological control of locusts and grasshoppers. Annu Rev Entomol 2001, 46:667–702.PubMedCrossRef 3. Peng G, Wang Z, Yin Y, Zeng D, Xia Y: Field trials of Metarhizium anisopliae var. acridum (Ascomycota: Hypocreales) against oriental migratory locusts, Locusta migratoria manilensis (Meyen) in Northern China. Crop Prot 2008, 27:1244–1250.CrossRef 4.