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Molecular characterization of Portuguese populations of the pinewood nematode Bursaphelenchus xylophilus using cytochrome b and cellulase genes

  • V. Valadas (a1), M. Laranjo (a2) (a3), M. Mota (a1) and S. Oliveira (a2)

Abstract

Bursaphelenchus xylophilus is the causal agent of pine wilt disease and a worldwide pest with high economic impact. Since its first diagnosis in Portugal in 1999, it has been subjected to quarantine measures with impact on forest health and ecosystem stability, significantly affecting international trade of wood products. The disease was detected in the north and centre of continental Portugal and, since 2008, the whole country has been considered an affected area. Recently, it was detected in Madeira Island. In order to avoid new outbreaks, it has become of major importance to understand the patterns of spread, introduction points and to characterize the new populations from continental Portugal and Madeira Island. Mitochondrial cytochrome b (cytb) and parasitic cellulase gene sequences were used to evaluate the genetic relationships among isolates that could indicate possible origins of the new outbreaks. Portuguese isolates were compared with isolates from USA, China, Japan and South Korea, in order to investigate possible infection pathways and disease spread patterns in Portugal. Phylogenetic trees based on both genes show that Portuguese isolates group with Asian isolates. Isolates from USA are in a separate position in both gene trees. However, the phylogenetic tree based on the cellulase gene sequences shows higher differentiation among Portuguese isolates than that of cytb. These results agree with those previously obtained using inter-simple sequence repeats (ISSR). This was the first study to use cytb and cellulase genes to characterize pinewood nematode (PWN) populations. This study suggests that cellulase is a better marker than cytb to study genetic diversity in B. xylophilus.

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Abelleira, A., Picoaga, A., Mansilla, J.P. & Aguin, O. (2011) Detection of Bursaphelenchus xylophilus, causal agent of pine wilt disease on Pinus pinaster in North-western Spain. EPPO Bulletin 39, 344.
Akbult, S. & Stamps, W.T. (2011) Insect vectors of the pinewood nematode: a review of the biology and ecology of Monochamus species. Forest Pathology 42, 8999.
Braasch, H., Tomiczek, C., Metge, K., Hoyer, U., Burgermeister, W., Wulfert, I. & Schonfeld, U. (2001) Records of Bursaphelenchus spp. (Nematoda, Parasitaphelenchidae) in coniferous timber imported from the Asian part of Russia. Forest Pathology 31, 129140.
Cheng, X.Y., Cheng, F.X., Xu, R.M. & Xie, B.Y. (2008) Genetic variation in the invasive process of Bursaphelenchus xylophilus (Aphelenchida: Aphelenchoididae) and its possible spread routes in China. Heredity 100, 356365.
Cheng, X.Y., Dai, S.M., Xiao, L. & Xie, B.Y. (2010) Influence of cellulase gene knockdown by dsRNA interference on the development and reproduction of the pine wood nematode, Bursaphelenchus xylophilus. Nematology 12, 225233.
Dieterich, C. & Sommer, R.J. (2009) How to become a parasite – lessons from the genomes of nematodes. Trends in Genetics 25, 203209.
Felsenstein, J. (1985) Confidence limits on phylogenies: an approach using the bootstrap. Evolution 39, 783791.
Fonseca, L., Lopes, A., Cardoso, J., Pestana, M., Abreu, F., Nunes, N., Mota, M. & Abrantes, I. (2010). The pinewood nematode, Bursaphelenchus xylophilus, in Madeira Island. 30th International Symposium of European Society of Nematologists, 19–23 September, Vienna, Austria, p. 176 (abstracts).
Fuchs, A.G. (1937) Neue parasitische und halbparasitische Nematoden bei Borkenkafern und einige andere Nematoden. I. Teil. Zoologische Jahrbuecher Abteilung fuer Systematik Oekologie und Geographie der Tiere 70, 291380.
Goellner, M., Smant, G., De Boer, J.M., Baum, T. & Davis, E.L. (2000) Isolation of β-1,4-endoglucanase genes from Globodera tabacum and their expression during parasitism. Journal of Nematology 32, 154165.
Gu, J., Braasch, H., Burgermeister, W., Brandstetter, M. & Zhang, J. (2006) Description of Bursaphelenchus yongensis sp. n. (Nematoda: Parasitaphelenchidae) isolated from Pinus massoniana in China. Russian Journal of Nematology 14, 9199.
Hall, T.A. (1999) BioEdit: a user-friendly biological sequences alignment editor and analysis program for Windows 95/98/NT. Nucleic Acids Symposium 41, 9598.
Jones, J.T., Furlanetto, C. & Kikuchi, T. (2005) Horizontal gene transfer from bacteria and fungi as a driving force in the evolution of plant parasitism in nematodes. Nematology 7, 641646.
Kikuchi, T. (2008) Parasitism genes of pine wood nematode. pp. 6780in Zhao, B.G., Futai, K., Sutherland, J.R. & Takeuchi, Y. (Eds) Pine wilt disease. Japan, Springer Press.
Kikuchi, T., Jones, J.T., Aikawa, T., Kosaka, H. & Ogura, N. (2004) A family of glycosyl hydrolase family 45 cellulases from the pinewood nematode Bursaphelenchus xylophilus. FEBS Letters 572, 201205.
Kikuchi, T., Aikawa, T., Kosaka, H., Pritchard, L., Ogura, N. & Jones, J.T. (2007) Expressed sequence tag (EST) analysisof the pine wood nematode Bursaphelenchus xylophilus and B. mucronatus. Molecular and Biochemical Parasitology 155, 917.
Kikuchi, T., Cotton, J.A., Dalzell, J.J., Hasegawa, K., Kanzaki, N., McVeigh, P., Takanashi, T., Tsai, I.J., Assefa, S.A., Cock, P.J.A., Otto, T.D., Hunt, M., Reid, A.J., Sanchez-Flores, A., Tsuchihara, K., Yokoi, T., Larsson, M.C., Miwa, J., Maule, A.G., Sahashi, N., Jones, J.T. & Berriman, M. (2011) Genomic insights into the origin of parasitism in the emerging plant pathogen Bursaphelenchus xylophilus. PLOS Pathogens 7 (9), e10022. doi:10.1371/journal.ppat.1002219.
Kimura, M. (1980) A simple method for estimating the evolutionary rate of base substitutions through comparative studies of nucleotide sequences. Journal of Molecular Evolution 16, 111120.
Kiyohara, T. & Bolla, R.I. (1990) Pathogenic variability among populations of the pinewood nematode, Bursaphelenchus xylophilus. Forest Science 36, 10611076.
Ma, H.B., Lu, Q., Liang, J. & Zhang, X.Y. (2011) Functional analysis of the cellulose gene of the pine wood nematode, Bursaphelenchus xylophilus, using RNA interference. Genetics and Molecular Research 10, 19311941.
Madani, M., Subbotin, S.A., Ward, L.J., Li, X. & De Boer, S.H. (2010) Molecular characterization of Canadian populations of potato cyst nematodes, Globodera rostochiensis and G. pallida using ribosomal nuclear RNA and cytochrome b genes. Canadian Journal of Plant Pathology 32, 252263.
Mota, M., Braasch, H., Bravo, M.A., Penas, A.C., Burgermeister, W., Metge, K. & Sousa, E. (1999) First report of Bursaphelenchus xylophilus in Portugal and in Europe. Nematology 1, 727734.
Picard, D., Sempere, T. & Plantard, O. (2007) A northward colonisation of the Andes by the potato cyst nematode during geological times suggests multiple host-shifts from wild to cultivated potatoes. Molecular Phylogenetics and Evolution 42, 308316.
Plantard, O., Picard, D., Valette, S., Scurrah, M., Grenier, E. & Mugniéry, D. (2008) Origin and genetic diversity of Western European populations of the potato cyst nematode Globodera pallida inferred from mitochondrial sequences and microsatellite loci. Molecular Ecology 17, 22082218.
Robertson, L., Arcos, C.S., Escuer, M., Merino, S.R., Esparrago, G., Abelleira, A. & Navas, A. (2011) Incidence of the pinewood nematode Bursaphelenchus xylophlius Steiner & Buhrer, 1934 (Nickle, 1970) in Spain. Nematology 13, 755757.
Rodrigues, J. (2008) Eradication program for the pinewood nematode in Portugal. pp. 514in Mota, M. & Vieira, P. (Eds) Pine wilt disease: a worldwide threat to forest ecosystems. The Netherlands, Springer Press.
Rosso, M.N., Favery, B., Piotte, C., Arthaud, L., De Boer, J.M., Hussey, R.S., Bakker, J., Baum, T.J. & Abad, P. (1999) Isolation of a cDNA encoding a β-1,4-endoglucanase in the root-knot nematode Meloidogyne incognita and expression analysis during plant parasitism. Molecular Plant–Microbe Interactions 12, 585591.
Saitou, N. & Nei, M. (1987) The neighbor-joining method: a new method for reconstructing phylogenetic trees. Molecular Biological Evolution 4, 406425.
Smant, G., Stokkermans, J.P.W., Yan, Y., De Boer, J.M., Baum, T.J., Wang, X., Hussey, R.S., Gommers, F.J., Henrissat, B., Davis, E.L., Helder, J., Schots, A. & Bakker, J. (1998) Endogenous cellulases in animals: isolation of β-1,4-endoglucanase genes from two species of plant parasitic cyst nematodes. Proceedings of the National Academy of Sciences of the United States of America 95, 49064911.
Southey, J.F. (1986) Laboratory methods for work with plant and soil nematodes. 202 pp. Ministry of Agriculture Fisheries and Food. London, Her Majesty's Stationary Office.
Steiner, G. & Buhrer, E.M. (1934) Aphelenchoides xylophilus n. sp., a nematode associated with blue-stain and other fungi in timber. Journal of Agricultural Research 48, 949951.
Sutherland, J.R. & Peterson, M. (1999) The pinewood nematode in Canada: history, distribution, hosts, potential vectors and research. pp. 247253in Futai, K., Togashi, K. & Ikeda, T. (Eds) Sustainability of pine forests in relation to pine wilt and decline. Proceedings of the Symposium, 26–30 October 1998, Tokyo, Japan. Kyoto, Japan, Shokado Shoten.
Suzuki, K. (2002) Pine wilt disease – a threat to pine forest in Europe. Dendrobiology 48, 7174.
Tamura, K. & Nei, M. (1993) Estimation of the number of nucleotide substitutions in the control region of mitochondrial DNA in humans and chimpanzees. Molecular Biology and Evolution 10, 512526.
Tamura, K., Peterson, D., Peterson, N., Stecher, G., Nei, M. & Kumar, S. (2011) MEGA5: molecular evolutionary genetics analysis using maximum likelihood, evolutionary distance, and maximum parsimony methods. Molecular Biology and Evolution 28, 27312739.
Tomiczek, C., Braasch, H., Burgermeister, W., Metge, K., Hoyer, U. & Brandstetter, M. (2003) Identification of Bursaphelenchus spp. isolated from Chinese packaging wood imported to Austria. Nematology 5, 573581.
Tomminen, J., Halik, S. & Berglahl, D.R. (1991) Incubation temperature and time effects on life stages of Bursaphelenchus xylophilus in wood chips. Journal of Nematology 24, 477484.
Valadas, V., Laranjo, M., Barbosa, P., Espada, M., Mota, M. & Oliveira, S. (2012) The pinewood nematode, Bursaphelenchus xylophilus, in Portugal: possible introductions and spread routes of a serious biological invasion revealed by molecular methods. Nematology, in press. doi:10.1163/156854112X632673
Vicente, C., Espada, M., Vieira, P. & Mota, M. (2012) Pine wilt disease: a threat to European forestry. European Journal of Plant Pathology, in press. doi 10.1007/s10658-011–9924-x.
Vieira, P., Burgermeister, W., Mota, M., Metge, K. & Silva, G. (2007) Lack of genetic variation of Bursaphelenchus xylophilus in Portugal revealed by RAPD-PCR analyses. Journal of Nematology 39, 118126.
Xie, B.-Y., Cheng, X.-Y., Shi, J., Zhang, Q.-W., Dai, S.-M., Cheng, F.X. & Luo, Y.-Q. (2009) Mechanisms of invasive population establishment and spread of pinewood nematodes in China. Science in China Series C: Life Sciences 52, 587594.
Zhao, B. & Lin, F. (2005) Mutualistic symbiosis between Bursaphelenchus xylophilus and bacteria of the genus Pseudomonas. Forest Pathology 35, 339345.

Molecular characterization of Portuguese populations of the pinewood nematode Bursaphelenchus xylophilus using cytochrome b and cellulase genes

  • V. Valadas (a1), M. Laranjo (a2) (a3), M. Mota (a1) and S. Oliveira (a2)

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