The purL gene of Bacillus subtilis is associated with nematicidal activity

Abstract

Parasitic nematodes of plants are important plant pathogens that represent a significant financial burden on agriculture. This study evaluated the efficacy of Bacillus spp. as nematode biocontrol agents and identified Bacillus genes associated with nematicidal activity. Culture by products of Bacillus subtilis strains OKB105 and 69 and Bacillus amyloliquefaciens strains FZB42 and B3 were used to treat Aphelenchoides besseyi, Ditylenchus destructor, Bursaphelenchus xylophilus and Meloidogyne javanica, respectively. The highest mortality rates were observed at 12 h when combinations of either A. besseyi/B3, D. destructor/OKB105, B. xylophilus/69 or M. javanica/OKB105 resulted in 10.6%, 27.6%, 35.6% and 100% mortality rates, respectively. Supernatant analysis demonstrated that the nematicidal active ingredients of strain OKB105, with a molecular weight of <1000 Da, were nonproteinaceous, heat and cold resistant, highly polar and could be evaporated but not extracted by some organic solvents. To identify nematicidal-related genes, 2000 OKB105 mutants were generated using the TnYLB-1 transposon. Mutant M1 lost nematicidal activity by 72 h and inverse PCR results demonstrated disruption of the purL gene. Nematicidal activity was restored when M1 mutant was complemented with either plasmid pMA5-purL or pUC18-purL, demonstrating a role for purL in mediating nematicidal activity.