We have characterized a family of GHF45 cellulases from the pine wood nematode Bursaphelenchus xylophilus. The absence of such genes from other nematodes and their similarity to fungal genes suggests that they may have been acquired by horizontal gene transfer (HGT) from fungi. The cell wall degrading enzymes of other plant parasitic nematodes may have been acquired by HGT from bacteria. B. xylophilus is not directly related to other plant parasites and our data therefore suggest that horizontal transfer of cell wall degrading enzymes has played a key role in evolution of plant parasitism by nematodes on more than one occasion.

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Identifying the invasion routes and determining

The pinewood nematode, Bursaphelenchus xylophilus, is the causative agent of pine wilt disease. The virulence of nematodes significantly varies among pine trees, but does not vary within a single pine. To elucidate the reason for no variation in virulence within a single pine, a technique to investigate the population structure of nematodes of different virulence is needed. In this study, the demonstration of interbreeding between virulent and avirulent populations of B. xylophilus in vitro was attempted using PCR-RFLP (polymerase chain reaction-restriction fragment length polymorphism) method. Ribosomal DNA containing the 5.8S gene, the internal transcribed spacer regions 1 and 2, and partial regions of 18S and 28S genes were used for analysis. First, PCR-RFLP patterns of offspring produced by interbreeding between two individuals of virulent and avirulent isolates were analyzed. The offspring from a single-pair interbreeding showed 3-digested fragment patterns with the restriction enzyme HhaI, patterns the same as the virulent isolate, the avirulent isolate and a hybrid pattern containing both fragments of the virulent and avirulent isolates. The virulent population was mixed with the avirulent one in vitro and propagated nematodes were individually analyzed by PCR-RFLP method. The nematodes showed the same 3 PCR-RFLP patterns as the offspring from a single-pair interbreeding. The detection of nematodes with a hybrid pattern demonstrates the occurrence of interbreeding between virulent and avirulent populations of B. xylophilus.

Deladenus nitobein. sp., a parasite of a woodwasp species,Sirex nitobei, is described based on its typological characters and molecular profiles of part of the small subunit D2-D3 expansion segments of the large subunit and internal transcribed spacer region of the ribosomal RNA gene, as well as part of the mitochondrial cytochrome oxidase subunit I (mtCOI) gene. Adult host woodwasps emerging from dead Japanese red pine logs,Pinus densiflora, collected at Aomori, Japan, were dissected and examined for nematode association. The new species was isolated from the body cavity and reproductive system ofS. nitobeias large parasitic females and small parasitic juveniles. The nematodes were cultured successfully on 1.0% malt extract agar medium, inoculated withSirex-associated fungus,Amylostereumareolatum. The mycophagous adult nematodes were characterised by the relative position of the excretory pore, located at 25 (19-28) and 25 (18-30)μm anterior to the hemizonid in the male and female, respectively, and a broad female tail with a rounded distal end. Typologically, the new species forms a cryptic species complex withD. siricidicolaandD. canii. In addition, the new species andD. siricidicolashare the same host wasp, tree and fungal species in Japan. However, the cryptic species can be separated from each other based on the described morphological and molecular sequence differences in the mtCOI gene.