The role of wood-inhabiting bacteria in pine wilt disease.

The pathogenicity of the pine wood nematode (PWN), Bursaphelenchus xylophilus together with the bacteria isolated from black pine (Pinus thunbergii) bark inoculated to axenic black pine seedlings, significantly exceeded that of the axenic PWNs alone, demonstrating that the bacteria play an important role in pine wilt disease. Inoculation of seedlings with bacteria-free culture filtrates of the seven isolates from the dead seedlings from the above experiment showed that all isolate filtrates killed the seedlings within 8 days. Identification of the bacteria using 16S rDNA sequencing showed that the isolates belonged to strains By253Ydz-fq, S209, 210-50 and 210-50 in Bacillus and the DN1.1 strain of Stenotrophomonas maltophilia, respectively. Completing Koch's postulates using the seven bacterial isolates to inoculate pine seedlings showed that all the seedlings that received aseptic PWNs mixed with the seven bacterial isolates died within 18 days post inoculation, while those inoculated with 'wild' PWNs died 16 days post inoculation. No disease symptoms developed on seedlings that received sterile water or aseptic PWNs. The horizontal transfer of the pathogenic bacteria may explain differences in bacterial species carried by PWN in different geographic areas.