The pine wood nematode, Bursaphelenchus xylophilus, in Portugal: possible introductions and spread routes of a serious biological invasion revealed by molecular methods

Abstract

The pine wood nematode (PWN), Bursaphelenchus xylophilus, the causal agent of pine wilt disease (PWD), is a major world-wide pathogen and pest of pine, with impacts on forest health, natural ecosystem stability and international trade. In Portugal, PWN was first diagnosed in 1999, the first occurrence also for Europe. The disease was recently detected on the island of Madeira and in northern Spain. In an attempt to search for more reliable and robust molecular markers that enable the study of intraspecific variability of B. xylophilus from different geographic locations, the intergenic spacer (IGS) region of the 5S rRNA gene and inter-simple sequence repeats (ISSR) analysis were used to determine the genetic relationships among 43 B. xylophilus isolates from Portugal, China, Japan, South Korea and USA. IGS sequence analysis showed that this region can only be used to establish interspecific relationships, since no differences were detected among Portuguese isolates from different geographic locations. Fingerprints obtained with ISSR show high genetic variability among Portuguese isolates, except for the ones obtained prior to 2008. The ISSR dendrogram suggests the spread of the disease inside continental Portugal and to Madeira. Until 2008, B. xylophilus populations found in continental Portugal showed low genetic diversity, pointing to a single introduction, probably from Asia, whereas recent populations from continental Portugal (2009-2010) and Madeira show high genetic diversity, suggesting multiple introductions from different origins.

Molecular characterization of Portuguese populations of the pinewood nematodeBursaphelenchus xylophilususing cytochromeband cellulase genes

“…Our results agree with previous studies (Vieira et al, 2007;Valadas et al, 2012) in suggesting a possible Asian origin for Portuguese isolates. Due to the high copy number in individual cells, the lack of recombination and the strict maternal inheritance, mitochondrial DNA (mtDNA) seems to be a potential marker to study genetic diversity and characterize populations from the same organism (Madani et al, 2010).…”

Specifically Expressed Genes of the Nematode Bursaphelenchus Xylophilus Involved with Early Interactions with Pine Trees

“…Pine wilt disease (PWD) is one of the most serious diseases [1,2] that affects coniferous forests around the world and it is considered to be caused by pine wood nematodes (PWNs), Bursaphelenchus xylophilus (Steiner & Buhrer) Nickle [3,4]. In Asia, PWD was introduced into Japan during the early 20 th century [5] and it spread subsequently to other countries, including China [6] and Korea [7], where pine trees have no natural resistance to B. xylophilus .…”

Pine Wilt Disease: a threat to European forestry

“…However, in 2008 the PWD was detected in other areas in central Portugal, and also in the island of Madeira, 1000 km SW of mainland Portugal , and the entire territory is now being classified as "affected area" (Rodrigues 2008). An ongoing study shows that these recent outbreaks seem to point out possible multiple introductions, most likely from other origins and spreading from the initial affected area (Valadas et al 2011). The recent report of PWN in Spain Robertson et al 2011), demonstrates a tendency for expansion of the PWN into new areas, although the origin of these new introductions is still unknown.…”