Bursaphelenchus tiliae sp. n. (Nematoda: Parasitaphelenchidae), a nematode associate of the bark beetle Ernoporus tiliae (Panz.) (Coleoptera: Curculionidae, Scolytinae), in small-leaved lime, Tilia cordata Mill.

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Bursaphelenchus tiliae sp. n. is described from the bark of dead branches of the small-leaved lime, Tilia cordata, infested with the lime bark beetle, Ernoporus tiliae. The nematode adults and propagative juveniles were present in larval galleries, while dispersal dauer juveniles colonised the Malpighian tubules of adult beetles. Bursaphelenchus tiliae sp. n. is characterised by the body length of 664 (568-817) μm in female and 622 (515-754) μm in male, moderately slender body (a = 41.0 (34.7-47.9) and 43.1 (37.8-51.9) in female and male, respectively), and spicules 20.0 (18.1-21.8) μm long. The extended anterior vulval lip in female, lateral fields with four incisures, long post-uterine sac, and number (7) and arrangement of male caudal papillae, may indicate that B. tiliae sp. n. is closely related to the xylophilus group. It differs from this group by smaller, arcuate spicules with relatively short rostrum, narrow capitulum (5.2-6.9 μm), and indistinct condylus in an almost continuous line with dorsal lamina. Distal third of spicule dorsal contour is usually almost straight. The spicules have a distinct, disk-like cucullus at the terminus. The status of the new species is confirmed by the unique molecular profile of the ITS region (ITS-RFLP) and its close relation to the xylophilus group by DNA sequencing and phylogenetic analysis of the 28S rDNA region. The presence of B. tiliae sp. n. dauer juveniles in Malpighian tubules of their vector bark beetle, E. tiliae, caused limited pathogenicity expressed as a local expansion of the tubule basement membrane and partial degradation of the cellular epithelium.

Bursaphelenchus tiliae sp. n. (Nematoda: Parasitaphelenchidae), a nematode associate of the bark beetle Ernoporus tiliae (Panz.) (Coleoptera: Curculionidae, Scolytinae), in small-leaved lime, Tilia cordata Mill.

in Nematology

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References

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Figures

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    Bursaphelenchus tiliae sp. n. A: Female; B: Male; C: Anterior region (female); D: Vulva region (female); E, F: Male tail with spicules (range of forms), bursa and positions of caudal papillae (lateral view); G, H: Female tail (range of forms); I-K: Male tail showing shape of bursa (range of forms) and position of caudal papillae (partially ventral view, somewhat flattened); L: Dauer juvenile (dissected from a Malpighian tubule of vector bark beetle, Ernoporus tiliae).

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    Light (DIC) and SEM micrographs of Bursaphelenchus tiliae sp. n. A-L: Adults. A: Head with stylet, median bulb and nerve ring; B: Head and median bulb showing position of excretory pore; C: Vulva with extended anterior lip (flap) and spermatheca, crustaformeria (anterior) and post-uterine sac (posterior); D: Vulva with extended anterior lip; E: Lateral field with four incisures; F, G: Range of spicule forms; H: Positions of male caudal papillae (SEM); I, J: Modifications of otherwise ovoid bursa (partially ventral view – somewhat flattened); K, L: Female tail (range of forms); M: Dauer juvenile (entire); N: Dauer anterior region with median bulb; O: Dauer tail.

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    ITS-RFLP profile of Bursaphelenchus tiliae sp. n. Lanes 1 and 8 = DNA marker (100 bp, Invitrogen, with lowest band at 200 bp and prominent bands at 600, 1500, 2072 bp). Lane 2 = uncleaved PCR product; Lanes 3-7 = restriction fragments obtained with: Lane 3 = RsaI; Lane 4 = HaeIII; Lane 5 = MspI; Lane 6 = HinfI; Lane 7 = AluI.

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    Maximum likelihood (ML) analysis of Bursaphelenchus tiliae sp. n. and 39 other Bursaphelenchus species based on partial 28S rDNA. Aphelenchoides fragariae served as outgroup species.

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    Neighbour-joining (NJ) analysis of Bursaphelenchus tiliae sp. n. and 39 other Bursaphelenchus species based on partial 28S rDNA. Aphelenchoides fragariae served as outgroup species.

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    Maximum parsimony (MP) analysis of Bursaphelenchus tiliae sp. n. and 39 other Bursaphelenchus species based on partial 28S rDNA. Aphelenchoides fragariae served as outgroup species.

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    Light micrographs of Malpighian tubules of adult Ernoporus tiliae (DIC). A: Entire tubule with dauer juveniles of Bursaphelenchus tiliae sp. n. in distal part; B: Uninfested tubule with cellular epithelium and distinct nuclei; C-E: Tubules colonised by dauer juveniles of B. tiliae sp. n.; C, D: Overall view of tubule filled with nematodes showing local expansion of tubule width; E: Detailed view of tubule filled with nematodes showing epithelial tissue only partially damaged with cells and distinct nuclei still present.

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