Abstract
The pinewood nematode, Bursaphelenchus xylophilus, is native to North America but it only causes damaging pine wilt disease in those regions of the world where it has been introduced. The accurate detection of the species and its dispersal routes are thus essential to define effective control measures. The main goals of this study were to analyse the genetic diversity among B. xylophilus isolates from different geographic locations and identify single nucleotide polymorphism (SNPs) markers for geographic origin, through a comparative transcriptomic approach. The transcriptomes of seven B. xylophilus isolates, from Continental Portugal (4), China (1), Japan (1) and USA (1), were sequenced in the next generation platform Roche 454. Analysis of effector gene transcripts revealed inter-isolate nucleotide diversity that was validated by Sanger sequencing in the genomic DNA of the seven isolates and eight additional isolates from different geographic locations: Madeira Island (2), China (1), USA (1), Japan (2) and South Korea (2). The analysis identified 136 polymorphic positions in 10 effector transcripts. Pairwise comparison of the 136 SNPs through Neighbor-Joining and the Maximum Likelihood methods and 5-mer frequency analysis with the alignment-independent bilinear multivariate modelling approach correlated the SNPs with the isolates geographic origin. Furthermore, the SNP analysis indicated a closer proximity of the Portuguese isolates to the Korean and Chinese isolates than to the Japanese or American isolates. Each geographic cluster carried exclusive alleles that can be used as SNP markers for B. xylophilus isolate identification.
Highlights
Bursaphelenchus xylophilus, the pinewood nematode (PWN), is a migratory plant-parasite nematode and the causal agent of the pine wilt disease
This study provides the first single nucleotide polymorphisms (SNPs) screening of PWN collected from different geographic locations and demonstrates the power of high throughput transcriptome sequencing to uncover genetic diversity in non-model organisms
Additional Bursaphelenchus xylophilus isolates included in the validation of single nucleotide polymorphisms (SNPs) in genomic DNA
Summary
Bursaphelenchus xylophilus, the pinewood nematode (PWN), is a migratory plant-parasite nematode and the causal agent of the pine wilt disease. This nematode is transmitted from tree to tree by insect vectors, mainly belonging to the genus Monochamus [1,2,3]. The nematodes enter pines through the wounds caused by the insect feeding on the twig bark. First detected in North America (USA and Canada), the PWN spread to Japan in the early part of the twentieth century. In North America, the nematode caused limited damage, it became a severe conifer pest in Japan [7].
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