Effects of the wet catching method on the detection of Bursaphelenchus xylophilus from trapped longhorn beetle vectors

in Nematology
Restricted Access
Get Access to Full Text
Rent on DeepDyve

Have an Access Token?

Enter your access token to activate and access content online.

Please login and go to your personal user account to enter your access token.


Have Institutional Access?

Access content through your institution. Any other coaching guidance?


The pine wood nematode (PWN), Bursaphelenchus xylophilus, causes pine wilt disease and is transmitted by Monochamus beetles. An efficient trapping system for these organisms is essential for their early detection in threatened regions. A wet catching method, using monoethylene glycol (MEG) in a collecting cup to preserve captured insects, was suitable for catching longhorn beetles. As a second step in developing this technique, we studied the influence of MEG on nematode detection. When M. galloprovincialis carrying PWN were submerged in MEG, nematodes died within 24 h. The preservative altered certain morphological features of nematodes immersed in solutions of 10, 30 or 60% MEG, impeding their morphological identification. However, molecular identification was possible as long as the MEG concentration was below 70%. At higher concentrations, a rinsing step with water before DNA extraction was enough to allow molecular detection. Wet trapping requires less frequent monitoring than dry trapping, thus reducing maintenance and related costs.


International Journal of Fundamental and Applied Nematological Research



AbelleiraA.PicoagaA.MansillaJ.AguinO. (2011). Detection of Bursaphelenchus xylophilus, causal agent of pine wilt disease on Pinus pinaster in northwestern Spain. Plant Disease 95, 776. DOI: 10.1094/PDIS-12-10-0902

AllisonJ.D.BhandariB.D.McKenneyJ.L.MillarJ.G. (2014). Design factors that influence the performance of light intercept traps for the capture of longhorned beetles (Coleoptera: Cerambycidae) from the subfamilies Lamiinae and Cerambycinae. PLoS ONE 9, e93203. DOI: 10.1371/journal.pone.0093203

ÁlvarezG.EtxebesteI.GallegoD.DavidG.BonifacioL.JactelH.SousaE.PajaresJ.A. (2015). Optimization of traps for live trapping of pine wood nematode vector Monochamus galloprovincialis. Journal of Applied Entomology 139, 618-626. DOI: 10.1111/jen.12186

BerkvensN.CasteelsH.DammeN.BighiuA.WittersJ.GregoireJ.C.BooneC.ViaeneN.MichelanteD. (2013). Bursaphelenchus xylophilus does not occur in Belgium, but what about its vectors, the Monochamus spp.? In: SchröderT. (Ed.). Pine wilt disease conference 2013, Braunschweig, Germany, pp.  116-117.

BerkvensN.ViaeneN.WayenbergeL.de SutterN.BooneC.GrégoireJ.-C.MichelanteD.CasteelsH. (2015). Vector required: the case of Bursaphelenchus xylophilus and Monochamus spp. in Belgium. Proceedings of the 67th ISCP symposium. Communications in agriculture and applied biological sciences 80, p. 15 [Abstr.].

DawsonM.N.RaskoK.A.JacobsD.K. (1998). Field preservation of marine invertebrate tissue for DNA analyses. Molecular Marine Biology and Biotechnology 7, 145-152.

De GrootP.NottR.W. (2003). Response of Monochamus (Col., Cerambycidae) and some Buprestidae to light intercept traps. Journal of Applied Entomology 127, 548-552. DOI: 10.1046/j.1439-0418.2003.00799.x

EPPO (2013). PM 7/4 (3) Bursaphelenchus xylophilus. EPPO Bulletin 43, 105-118. DOI: 10.1111/epp.12024

EPPO (2016). EPPO Global Database, Monochamus spp. Available online at https://gd.eppo.int/taxon/1MONCG (accessed 5 March 2017).

EvansH.F.McNamaraD.G.BraaschH.ChadoeufJ.MagnussonC. (1996). Pest Risk Analysis (PRA) for the territories of the European Union (as PRA area) on Bursaphelenchus xylophilus and its vectors in the genus Monochamus. EPPO Bulletin 26, 199-249. DOI: 10.1111/j.1365-2338.1996.tb00594.x

EvansS.EvansH.IkegamiM. (2008). Modeling PWN-induced wilt expression: a mechanistic approach. In: MotaM.VieiraP. (Eds). Pine wilt disease: a worldwide threat to forest ecosystems. Dordrecht, The Netherlands, Springer, pp.  259-278.

FutaiK. (2013). Pine wood nematode, Bursaphelenchus xylophilus. Annual Review of Phytopathology 51, 61-83. DOI: 10.1146/annurev-phyto-081211-172910

HoltermanM.van der WurffA.van den ElsenS.van MegenH.BongersT.HolovachovO.BakkerJ.HelderJ. (2006). Phylum-wide analysis of SSU rDNA reveals deep phylogenetic relationships among nematodes and accelerated evolution toward crown clades. Molecular Biology and Evolution 23, 1792-1800. DOI: 10.1093/molbev/msl044

InácioM.NóbregaF.VieiraP.BonifácioL.NavesP.SousaE.MotaM. (2015). First detection of Bursaphelenchus xylophilus associated with Pinus nigra in Portugal and in Europe. Forest Pathology 45, 235-238. DOI: 10.1111/efp.12162

KilpatrickC.W. (2002). Noncryogenic preservation of mammalian tissues for DNA extraction: an assessment of storage methods. Biochemical Genetics 40, 53-62. DOI: 10.1023/A:1014541222816

LinitM.AkbulutS. (2008). The insect vectors: biology and ecology. In: MotaM.VieiraP. (Eds). Pine wilt disease: a worldwide threat to forest ecosystems. Dordrecht, The Netherlands, Springer, pp.  211-213.

MotaM.BraaschH.BravoM.PenasA.BurgermeisterW.MetgeK.SousaE. (1999). First report of Bursaphelenchus xylophilus in Portugal and in Europe. Nematology 1, 727-734. DOI: 10.1163/156854199508757

NavesP.BonifácioL.SousaE. (2015). Nematode-insect vector. In: SousaE.ValeF.AbrantesI. (Eds). Pine wilt disease in Europe. Biological interactions and integrated management. Lisbon, Portugal, FNAPF, pp.  79-121.

NickleW.R.GoldenA.M.MamiyaY.WerginW.P. (1981). On the taxonomy and morphology of the pine wood nematode, Bursaphelenchus xylophilus (Steiner & Buhrer, 1934) Nickle 1970. Journal of Nematology 13, 385-392.

PenasA.C.BravoM.A.NavesP.BonifácioL.SousaE.MotaM. (2006). Species of Bursaphelenchus Fuchs, 1937 (Nematoda: Parasitaphelenchidae) and other nematode genera associated with insects from Pinus pinaster in Portugal. Annals of Applied Biology 148, 121-131. DOI: 10.1111/j.1744-7348.2006.00042.x

R Development Core Team (2014). R: a language and environment for statistical computing. Vienna, Austria, the R Foundation for Statistical Computing. Available online at http://www.R-project.org (accessed 5 March 2017).

RodriguesJ.SousaE.AbrantesI. (2015). Pine wilt disease. Historical overview. In: SousaE.ValeF.AbrantesI. (Eds). Pine wilt disease in Europe. Biological interactions and integrated management. Lisbon, Portugal, FNAPF, pp.  11-32.

SchröderT.McNamaraD.GaarV. (2009). Guidance on sampling to detect pine wood nematode Bursaphelenchus xylophilus in trees, wood and insects. EPPO Bulletin 39, 179-188. DOI: 10.1111/epp.2621

SousaE.BravoM.A.PiresJ.NavesP.PenasA.C.BonifacioL.MotaM. (2001). Bursaphelenchus xylophilus (Nematoda; Aphelenchoididae) associated with Monochamus galloprovincialis (Coleoptera; Cerambycidae) in Portugal. Nematology 3, 89-91. DOI: 10.1163/156854101300106937

SweeneyJ.GutowskiJ.M.PriceJ.De GrootP. (2006). Effect of semiochemical release rate, killing agent, and trap design on detection of Tetropium fuscum (F.) and other longhorn beetles (Coleoptera: Cerambycidae). Environmental Entomology 35, 645-654.

YoderM.De LeyI.T.KingI.W.Mundo-OcampoM.MannJ.BlaxterM.PoirasL.De LeyP. (2006). DESS: a versatile solution for preserving morphology and extractable DNA of nematodes. Nematology 8, 367-376. DOI: 10.1163/156854106778493448


  • Changes in visibility (1 = not visible, 5 = clearly visible) over time of three morphological features of the genus Bursaphelenchus when submerged in rainwater (0% monoethylene glycol (MEG)) or different solutions of MEG (10, 30, 60, 100%). A: Cephalic region high and offset by a constriction with six lips; B: Lateral field with four lines; C: Excretory pore at/or behind median bulb.

    View in gallery
  • Visibility scoring (1 = not visible, 5 = clearly visible) over time of four morphological features for females (A, B) or males (C, D) of Bursaphelenchus xylophilus, when submerged in 0, 10, 30, 60 or 100% monoethylene glycol (MEG) solution (mixed with rainwater). A: Female tail broadly sub-cylindrical with or without mucron; B: Female vulval flap straight, not ending in a deep depression; C: Male spicule with long and pointed rostrum; limbs of spicule with an angular curvature; D: Male spicule length < 30 μm.

    View in gallery
  • Electrophoresis of the 18S rRNA PCR product of Bursaphelenchus xylophilus adults submerged in pure rainwater (lanes 1-9), 30% monoethylene glycol (MEG) (lanes 10-18) or 100% MEG (lanes 19-27) for 28 days. In the lanes marked with ∗, nematodes were rinsed in 25 μl pure water before DNA extraction; DNA in lanes 3, 6, 9, 12, 15, 18, 21, 24 and 27 was extracted from five nematodes; in the other lanes DNA was extracted from a single nematode; (–): negative control; L: 100 bp DNA ladder (Thermo Scientific).

    View in gallery
  • Electrophoresis of the 18S rRNA PCR product of Bursaphelenchus xylophilus adults submerged in 20, 30, 40, 50, 60, 70, 80, 90 and 100% monoethylene glycol (lanes 1-9) for 28 days, DNA was extracted from one individual; (–): negative control; L: 100 bp DNA ladder (Thermo Scientific).

    View in gallery


Content Metrics

Content Metrics

All Time Past Year Past 30 Days
Abstract Views 22 22 5
Full Text Views 11 11 10
PDF Downloads 1 1 0
EPUB Downloads 0 0 0