The RFLP technique was used to establish a reliable diagnostic method for 18 Pratylenchus species: Pratylenchus agilis, P. bolivianus, P. brachyurus, P. coffeae, P. crenatus, P. fallax, P. goodeyi, P. loosi, P. mediterraneus, P. neglectus, P. penetrans, P. pratensis, P. pseudocoffeae, P. scribneri, P. subranjani, P. thornei, P. vulnus and P. zeae. The polymerase chain reaction (PCR) amplified the ITS regions from all species and populations examined and revealed large differences in length, ranging in size from approximately 900 to 1250 bp. The rDNA fragments were digested with five restriction enzymes (CfoI, DdeI, HindIII, HpaII, and PstI). All Pratylenchus species can be differentiated from each other by a combination of at least two enzymes. CfoI differentiated all nematode species with the exception of P. fallax, P. penetrans and P. pseudocoffeae. P. fallax was further separated by a DdeI restriction, and P. pseudocoffeae by a PstI digestion. Intraspecific RFLP were observed. Upon CfoI, DdeI, HindIII, or HpaII digestion, it was possible to separate the three P. coffeae populations studied from each other. La technique RFLP a été utilisée pour créer une méthode fiable de diagnostic pour 18 espèces de Pratylenchus: Pratylenchus agilis, P. bolivianus, P. brachyurus, P. coffeae, P. crenatus, P. fallax, P. goodeyi, P. loosi, P. mediterraneus, P. neglectus, P. penetrans, P. pratensis, P. pseudocoffeae, P. scribneri, P. subranjani, P. thornei, P. vulnus et P. zeae. La réaction de polymérisation en chaîne (PCR) a amplifié les régions de l’ITS pour toutes les espèces et populations étudiées et a mis en évidence de grandes différences dans la taille des gammes de longueur, de 900 à 1250 bp approximativement. Les fragments de rDNA ont été digérés à l’aide de cinq enzymes de restriction (CfoI, DdeI, HindIII, HpaII, and PstI). Toutes les espèces de Pratylenchus ont pu être différenciées les unes des autres par une combinaison d’au moins deux enzymes. CfoI a différencié toutes les espèces à l’exception de P. fallax, P. penetrans et P. pseudocoffeae. P. fallax a été ultérieurement séparé par une restriction DdeI, et P. pseudocoffeae par une digestion PstI. Des RFLP intraspécifiques ont été observés. Par les digestions CfoI, DdeI, HindIII, ou HpaII, il s’est révélé possible de séparer les unes des autres les trois populations étudiées de P. coffeae.
Summary -The 75 valid species of the genus Bursaphelenchus are listed together with their synonyms. Diagnostic characters and their states are discussed and illustrated. Tabular and traditional text keys are provided for the genus. Two new subspecies are proposed to distinguish populations of B. piniperdae and B. poligraphi, as described by , from the original descriptions of these species published by . Known records of Bursaphelenchus species with their associated natural vectors, plants and plant families are given. Dendrograms of species relationships (UPGMA, standard distance: mean character difference) based on combined taxonomic characters and also on spicule characters only, are provided. Discussion as to whether the species groups are natural or artificial (and therefore purely diagnostic) is based on their relationships in the dendrogram and the vector and associated plant ranges of the species. Of the six species groups distinguished, two appear to represent natural assemblages, these being the xylophilus-group (with ten species) and the hunti-group (seven species), of which two, B. cocophilus and B. dongguanensis, form the cocophilus-cluster which is separated on the dendrogram from the main clusters. The remaining four species groups appear to be artificial and purely diagnostic in function, namely the aberrans-group (four species); the eidmanni-group (six species); the borealis-group (five species), and the piniperdae-group (43 species). Two new subspecies, both in the piniperdae-group, viz. B. piniperdae ruehmpiniperdae n. subsp. and B. poligraphi ruehmpoligraphi n. subsp., are proposed and diagnosed from B. piniperdae piniperdae and B. poligraphi poligraphi the respective type subspecies. Bursaphelenchus dongguanensis is regarded as being a valid member of the genus and its transfer to Parasitaphelenchus is rejected.Keywords -associated plants, dendrogram, key, morphology, new subspecies, taxonomy, vectors. The genus Bursaphelenchus Fuchs, 1937 was established by and includes nematodes that are associated with insects and dead or dying, mainly coniferous, trees and which have an ectophoretic stage. The type species is B. piniperdae . Most species are fungal feeders and are either transmitted to dead or dying trees during oviposition by insect vectors, or to healthy trees during maturation feeding of their insect vectors. The majority of vectors are beetles, mostly from the Scolytidae, Cerambycidae, Curculionidae and Buprestidae (see Appendix). Until recently, only one species of the genus, Bursaphelenchus cocophilus (Cobb, 1919) Baujard, 1989, was recorded outside of the northern hemisphere. However, with the record of B. leoni Baujard, 1980 in South Africa (Braasch et al., 1998, and more recently a Bursaphelenchus sp. from dying pine (Pinus * Corresponding author, e-mail: pvieira@uevora.pt halepensis Miller) in Australia (Ridley et al., 2001), the known range of the genus has significantly increased. Of the total number of known species, approximately 70% are associated with conifers, mainly...
A survey was conducted in the northern conifer forests of Turkey in 2003 and 2004 for the pinewood nematode, Bursaphelenchus xylophilus. Wood samples were collected from declining pine trees, located in the following target areas: Düzce, Ankara, Bolu, and Artvin. Nematodes were extracted from over 310 samples and were observed and identified. B. xylophilus was not detected in any samples. Bursaphelenchus species were only found in 6% of the samples. From the B. xylophilus‐group, only the species B. mucronatus was reported. Species identification was performed from morphological characters, particularly male spicules, and by molecular analysis with ITS‐RFLP.
This article summarizes the results of the research papers presented at the International Symposium on pine wilt disease (IUFRO Working Party Meeting 4.04.03) held in July 2009, at Nanjing, China. The general topics covered were on pine wilt disease (PWD), its causal organism, the pinewood nematode (PWN) Bursaphelenchus xylophilus, plus other PWN-associated microorganisms that play a signifi cant role in PWD such as bacteria (e.g. Pseudomonas fl uorescens). Most of the papers that are reviewed are based on work on PWD-PWN in East Asia and Russia. Specifi c topics covered include: 1) the fundamental conceptions of PWD development, 2) pathogenicity, 3) host-parasite relationships including the histopathology of diseased conifers and the role of toxins from bacteria-nematode ecto-symbionts, 4) PWN life cycle and transmission, 5) B. xylophilus dissemination models, 6) associations (with other nematodes), 7) diagnostics, 8) quarantine and control of the PWN and 9) biocontrol of the PWN.
A new species, Bursaphelenchus ulmophilus sp. n., from the hofmanni group is described morphologically and molecularly. This nematode species was found associated with Dutch elm disease of Ulmus glabra in parks of St Petersburg, Russia, and is vectored by adults and larvae of the bark beetles Scolytus multistriatus and S. scolytus. Bursaphelenchus ulmophilus sp. n. is characterised by the following features: body length 600-850 μm, stylet 12-14 μm long with base slightly and smoothly expanded, but lacking knobs, median bulb almost spherical in female and slightly ovoid in male, pharyngeal gland lobe dorsal, 4-5 body diam. long. This species has an oval spermatheca filled with spherical nucleic sperm 4-5 μm diam. Female post-uterine sac ca 0.5 of the vulva-anus distance and ca 3 vulval body diam. long, female tail reflexed, strongly hooked ventrally with a digitate or conically rounded tip. The male has seven caudal papillae arranged as 1 + 2 + 2 + 2, P1 is unpaired, anterior to cloacal opening, paired P2 at cloacal aperture, paired P3 and paired pore-like 'gland papilla' P4 at the lateral edges of the bursa which has the posterior border rounded to truncate. Phylogenetic analyses of the D2-D3 of 28S rRNA, partial 18S rRNA and ITS rRNA gene sequences revealed that B. ulmophilus sp. n. formed a clade with species of the hofmanni group and shared close relationships with B. hofmanni and B. pinasteri.
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