Abstract:

Objective: Monochamus alternates, causing great economic losses every year, is not only an important borer pest in pinewood, but also the main vector of pine nematode disease. To screen for more effective biocontrol agents against M. alternates, four Metarhizium strains were identified by molecular and morphological characteristics, and their virulence against M. alternates adults was analyzed. Method: In this study, cultural characteristics and conidia morphology of four Metarhizium strains were observed respectively. The EF-1α gene of each strain was cloned and sequenced, and a phylogenetic tree was constructed to comprehensively identify the taxonomic status of 4 strains to species level. The virulence of four Metarhizium strains against M. alternatus adult was tested with different conidia concentrations and inoculation methods(dipping, twig dipping, and nonwoven cloth), and the pathogenesis was evaluated. Result: 1) WP08 and WTKH were clusted into the clade of M. pingshaense, and LV2 was into M. anisopliae, and qc1401 was into M. pemphogum. 2) Among the four strains, WP08 and LV2 caused the highest corrected death rate (100%) in the 40-day experimental period, and the corrected death rate of qc1401 and WTKH was 91.9% and 87.9%, respectively. Lethal speed of each strain was in the order of WP08 > LV2 > qc1401 > WTKH with the LT₅₀ of 5.52, 10.38, 16.93 and 20.80 d, respectively. 3) Among the conidium concentration series of 1.0×10⁸, 1.0×10⁷, 1.0×10⁶ conidia·mL^-1 of WP08, the higher the conidia concentration, the higher the corrected death rate and the shorter the lethal time of M. alternatus adults. The corrected death rate was as high as 100% at conidium concentration of 1.0×10⁸ and 1.0×10⁷ conidia·mL^-1 of WP08, and was 83.9% at conidium concentration of 1.0×10⁶ conidia·mL^-1, and the LT₅₀ was 11.38, 16.44 and 24.22 d at the three concentration, separately. 4) For strain of WP08, all the three inoculation method could cause a death rate of 100% in the experimental period of 40 days, and the LT₅₀ of insect dipping, twig dipping, and nonwoven cloth were 5.52, 11.38, and 17.21 d, respectively. Conclusion: Four Metarhizium strains are identified as M. pingshaense, M. anisopliae, and M. pemphogum, and all the strains can infect M. alternatus adults and cause their death. M. pingshaense WP08 strain exhibits higher death rate and faster lethal speed than the other three stains, and shows great potential as biocontrol agent for M. alternatus.

Key words: entomopathogenic fungus, Metarhizium, Monochamus alteratus, virulence test, EF-1α, LT₅₀