Molecular and morphological characterisation of a new record of Bursaphelenchus arthuri (Nematoda: Aphelenchoididae) from a new host, Pinus pinea, in Europe

Nematodes with morphological characters of Bursaphelenchus arthuri, a species of the Fungivorus group, first described in 2005 in China from imported coniferous packaging wood, were extracted from a centennial Pinus pinea tree with wilting symptoms in Coimbra, Portugal. Nematodes were transferred to Botrytis cinerea cultures to establish a nematode isolate which was characterised using morphological, morphometrical and molecular methods. Restriction fragment length polymorphism (RFLP) analysis carried out with five endonucleases of amplified internal transcribed spacer (ITS) of the rDNA region, revealed the characteristic restriction pattern described for Bursaphelenchus arthuri. Sequencing of the D2-D3 expansion region of the large subunit (LSU) of rDNA confirmed this identification. Phylogenetic analysis based on multiple sequence alignment of selected D2-D3 sequences showed that the Portuguese B. arthuri isolate formed a separate group, along with the other B. arthuri isolate from Taiwan available in GenBank, from those of other isolates of closely related nematodes species belonging to the Fungivorus group. This study is the first report of B. arthuri in Europe and from P. pinea.