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Bursaphelenchus populi sp. n. is described from dying and dead aspen trees, Populus tremula, in Poland. The nematode was found in trees infested with a long-horn beetle, Saperda perforata, and is vectored in the insect haemocoel. The characteristic morphology of male spicules, extended anterior vulval lip, lateral fields with four incisures, and number and arrangement of male caudal papillae, place B. populi sp. n. in the xylophilus group. Bursaphelenchus populi sp. n. can be separated from all other species in that group by the distinctive vulval flap, which is always bent with its distal half sunken in a conspicuous, sharp depression posterior to the vulva, and other morphological and morphometric characters, i.e., female tail shape, excretory pore position, spicule length (32.1 (25.7-37.0) μm (as measured along arc) and shape, and a relatively long (i.e., L = 1020 (909-1111) μm in females), and L = 850 (756-1055) μm in males), and slender body (a = 45.4 (40.2-52.4) and 42.0 (36.1-49.3) in female and male, respectively). The status of the new species is confirmed by the unique molecular profile of the ITS region (ITS-RFLP). DNA sequencing and phylogenetic analysis of 28S rDNA region placed the new species close to B. fraudulentus in the xylophilus group. In vitro cross-breeding of the new nematode with B. xylophilus, B. mucronatus, B. fraudulentus and B. doui revealed full reproductive incompatibility between these species. In laboratory experiments on 2-year-old seedlings B. populi sp. n. was specific to aspen. It did not develop on pine or oak and reproduced only in weakened or dead aspen seedlings. No pathogenicity to the tree host or vector insect was observed. In Botrytis cinerea PDA cultures, body dimensions were significantly altered in both sexes and female tail morphology differed when compared to individuals extracted from aspen wood.

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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) μm in female and 874 (691-1122) μm 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) μm long with a small cucullus (1.3-1.5 μm 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.

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The control potential of a strain (ScP) of Steinernema feltiae genetically selected against the mushroom sciarid Lycoriella solani, was evaluated for the management of L. mali. Trials were conducted at two commercial mushroom farms with high and low levels of fly infestation. The efficacy of the ScP strain was compared with the SN strain of S. feltiae and the chitin synthesis inhibitor, diflubenzuron. At low densities of L. mali, the two strains did not differ in efficacy and caused 85-94% reduction in fly populations. At high fly densities with a mixed infestation of the phorid fly, Megaselia halterara, the ScP stain caused 5 6 8 3 % reduction in L. muli populations whereas the SN strain caused 51-73% reduction. Two doses of the improved strain (0.5 x 10' and 1.0 x 10' infective juveniles per m' cropping area) did not differ in efficacy from diflubenzuron which caused 80% reduction in L. mali populations. The lower dose (0.5 x 10') of the SN strain was less effective than diflubenzuron. The ScP strain had a major advantage over the SN strain in persistence in casing material. Nematodes had no significant effect on mushroom yields.

Bursaphelenchus xylophilus is an emerging pathogenic nematode that is responsible for a devastating epidemic of pine wilt disease worldwide, causing severe ecological damage and economic losses to forestry. Two forms of this nematode have been reported, i.e., with strong and weak virulence, commonly referred as virulent and avirulent strains. However, the pathogenicity-related genes of B. xylophilus are not sufficiently characterized. In this study, to find pathogenesis related genes we re-sequenced and compared genomes of two virulent and two avirulent populations. We identified genes affected by genomic variation, and functional annotation of those genes indicated that some of them might play potential roles in pathogenesis. The performed analysis showed that both avirulent populations differed from the virulent ones by 1576 genes with high impact variants. Demonstration of genetic differences between virulent and avirulent strains will provide effective methods to distinguish these two nematode virulence forms at the molecular level. The reported results provide basic information that can facilitate development of a better diagnosis for B. xylophilus isolates/strains which present different levels of virulence and better understanding of the molecular mechanism involved in the development of the PWD.

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