Taxonomic relationships of Bursaphelenchus xylophilus and B. mucronatus based on interspecific and intraspecific cross-hybridization and DNA analysis

Summary - Cross-hybridization between five geographic isolates from North America of Bursaphelenchus xylophilus (St. John, St. William, MSP4, Q1426) and Japan (Ibaraki) and two isolates Uapanese and French) of B. mucronalUS, members of the pinewood nematode species complex (PWNSC), was done under controlled laboratory conditions. Intraspecific hybridization among isolates of B. xylophilus from Japan and North America produced fertile offspring. Cross-hybridization among isolates of B. mucronalUs from France (female) and Japan (male) produced fertile Fn generations, however the reciprocal cross died out. Interspecific hybridization among isolates of B. mucronalUS and B. xylophilus produced an FI that died out in most of the crosses in which the female was from B. mucronalUS and the male was from B. xylophilus. Genomic DNA from B. xylophilus, B. mucronalUs and their hybrid-crosses were Southern blotted and probed with a ribosomal gene clone which produced band pattern differences between the isolates of B. xylophilusand of B. mucronalUsas weil as between the French and Japanese isolates of B. mucronalus. The results support the distinct species status of B. xylophilus and B. mucronalUs and suggest also that B. xylophilus, and B. mucronalUS from France and Japan come from a common ancestor. In addition, they provide evidence for separate species status for the French and Japanese isolates of B. mucronalUs. Resume - Relations taxonomiquesentl'e Bursaphelenchus xylophilus et B. mucronatus fondees SUl' des hybridations intel'specifiques et intraspecifiques cl'oisees et l'analyse de l'ADN - Des hybridations croisees entre cinq isolats geographi­ ques de Bursaphelenchusxylophilus provenant d'Amerique du Nord (St. Jolm, St. William, Q1426 et MSP4) et du Japon (Ibaraki), et deux isolats Gaponais et francais) de B. mucronalUs, membres du complexe d'especes du nematode du bois de pin (PWNSC), Ont ete realisees en conditions controlees au laboratoire. L'hybridation intraspecifique entre isolats de B. xylophilusvenant du Japon et d'Amerique du Nord produit des descendants fertiles. L'hybridation entre isolats de B. muCronalUSvenant de France (femelle) et du Japon (mâle) produit des generations fertiles Fo> mais la descendance de l'hybridation reciproque s'est eteinte. L'hybridation interspecifique entre isolats de B. mucronalUs et de B. xylophilus produit une FI qui meurt dans la plupart des cas ou la femelle appartient a B. mucronalUs et le mâle a B. xylophilus. L'hybridation d'une sonde d'ADN ribosomique avec les ADN genomiques de B. xylophilus, B. mucronalUs et de leurs hybrides montre des differences dans les profils d'hybridation entre les isolats de B. xylophilus et ceux de B. mucronatus, ainsi qu'entre les isolats francais et japonais de B. muCronalUs. Ces resultats confirment le statut d'especes distinctes de B. xylophilus et B. mucronalus et suggerent egalement que B. xylophilus et B. mucronatusde France et du Japon proviennent d'un ancetre commun. De plus, ils fournissent des arguments en faveur d'un statut d'especes distinctes pour les isolats francais et japonais de B. mucronatus. Bursaphelenchusxylophilus(Steiner & Buhrer) NickIe, the pinewood nematode, and its close relative B. mucro­ nalUs Mamiya & Enda, both members of the pinewood nematode species complex (PWNSC) (Webster el al., 1990), are important parasites of pines and sorne related species. Both species have attracted interest because of their economic importance to lumber trading nations, their relative pathogenicity and uncertainty regarding their respective taxonomie status. Differentiating be­ tween sorne PWNSC isolates, based solely on morpho­ logical characters, can sometimes be inconclusive be­ cause the characters of sorne isolates are similar and at the same time variable. Cross-hybridization experiments