Bursaphelenchus masseyi sp n. (Nematoda: Parasitaphelenchinae) - a nematode associate of the bark beetle, Trypophloeus populi Hopkins (Coleoptera: Curculionidae: Scolytinae), in aspen, Populus tremuloides Michx. affected by sudden aspen decline in Colorado

Bursaphelenchus masseyi sp. n. is described from trunks of unhealthy trembling aspen, Populus tremuloides, affected by sudden aspen decline (SAD) in Colorado, USA. All propagative stages of the nematode were present in larval galleries of a bark beetle, Trypophloeus populi. The dauer juveniles occupy the haemocoel of older larvae, pupae and adult beetles. The characteristic morphology of the male spicules with small but distinct cucullus, extended anterior vulval lip in female, lateral fields with four incisures, and number and arrangement of male caudal papillae, indicate that B. masseyi sp. n. is closely related to the xylophilus group. This relation has been confirmed by DNA sequencing and phylogenetic analysis of the 28S rDNA region. The new species is characterised by the body length of 958 (765-1203) mu m in female and 874 (691-1122) mu m in male, moderately slender body (a = 38.2 (33.5-41.1) and 38.0 (33.1-44.1) in female and male, respectively), and spicules 29.2 (26.2-34.7) mu m long with a small cucullus (1.3-1.5 mu m in diam.) at their tips. Bursaphelenchus masseyi sp. n. can be separated from other species in the xylophilus group by the morphology of spicules which have a short capitulum and unique rostrum that is pointed somewhat anteriorly, relatively thick vulval flap, which is straight, parallel to the body long axis and bent towards the body wall at its distal end, and other morphological and morphometric characters. The new species most closely resembles B. trypophloei, but differs by the morphology of spicules (short but distinct condylus vs condylus in a continuous line with dorsal lamina, and smaller cucullus). The taxonomic separation of the new species is also confirmed by the unique molecular profile of the ITS region (ITS-RFLP). Diallelic cross-breeding in vitro revealed also that B. masseyi sp. n. and B. trypophloei are reproductively incompatible. The new species showed poor ability to develop and reproduce on Botrytis cinerea cultures, although it grew vigorously on laboratory cultures of Polish and Colorado isolates of Cytospora chrysosperma, the fungus naturally associated with galleries of the nematode vector, T. populi.