Lee Robertson

Incidence of the pinewood nematode Bursaphelenchus xylophlius Steiner & Buhrer, 1934 (Nickle, 1970) in Spain

The pine wood nematode (PWN) Bursaphelenchus xylophilus Steiner & Buhrer, 1934 (Nickle, 1970), is a pathogenic species in the genus Bursaphelenchus Fuchs, 1937. The introduction and establishment of PWN in the Setubal Peninsula, Portugal (Mota et al., 1999), and the evidence that Monochamus galloprovincialis is its insect vector in this country (Sousa et al., 2001), has stimulated studies on its further distribution and its potential vector insects in Europe, including Spain. There is a clear risk of spread of pine wood nematode into Spanish territory from Portugal, both as a result of natural spread and due to the important timber trade between the two countries. The occurrence of Bursaphelenchus spp. in Spain has been previously studied (Abelleira et al., 2002, 2003; Escuer et al., 2003, 2004; Robertson et al., 2008). These studies have shown, in addition, the distribution of insects belonging to the Cerambycidae, Curculionidae and Scolytidae families (Cobos, 1986; Gil Sánchez & Pajares Alonso, 1986; Vives, 2000; Verdugo Paez, 2004), which are known to be vectors of Bursaphelenchus spp. other than B. xylophilus and are often found to be associated with stressed, dead or decaying conifers. Although it is generally accepted that B. xylophilus is the only truly pathogenic species (reviewed by Jones et al., 2008), other species may be involved in pine forest decline in some European regions or, alternatively, may simply be opportunistic infectors of stressed trees (Mamiya, 1999; Caroppo et al., 2000; Skarmoutsos et al., 2000). Pinus pinaster is the most representative

New host races of Meloidogyne arenaria, M. incognita, and M. javanica from horticultural regions of Spain.

In total, 140 populations of the genus Meloidogyne collected from representative horticultural regions of Spain and maintained at the Department of Agroecology, Institute of Agrarian Sciences, Madrid were characterized using perineal morphology, sequenced characterized amplified region polymerase chain reaction with species-specific primers, and bioassay. Using a modified North Carolina differential host test as a basis, 13 Meloidogyne populations did not fit into the published race scheme. These populations had very limited host ranges, reproducing only on susceptible tomato cv. Marmande and sometimes also on tobacco cv. NC95 and the pepper cv. Sonar in the case of Meloidogyne arenaria. They did not reproduce on cotton or peanut. The species and new races were identified as M. incognita races 5 (6 populations) and 6 (2 populations) and M. javanica race 5 (2 populations), with new records of M. arenaria race 3 (2 populations) and M. javanica race 1 (23 populations) for Spain.

Molecular Tools for Diagnostics

More than 4,000 nematode species are parasites of plants, some of which are responsible for considerable yield and quality losses in many important agronomical plants. Because of their small size, the homogeneity of their morphological characters, and their soil distribution, most plant-parasitic nematodes (PPNs) are difficult to detect and identify, even for well-qualified taxonomists. However, accurate identification to species level is central to the implementation of successful management strategies against PPNs. With the revolution of the polymerase chain reaction (PCR), DNA-based technology now provides numerous avenues to overcome the limitations intrinsic to the nature and biology of these parasites. The aim of this chapter is to present both the advances made in recent years and the current prospects for the development of practical, highly sensitive and specific molecular tools for the diagnostics of PPNs.

Protein markers of Bursaphelenchus xylophilus Steiner & Buhrer, 1934 (Nickle, 1970) populations using quantitative proteomics and character compatibility.

The Pine Wood Nematode (PWN) Bursaphelenchus xylophilus is a severe forest pathogen in countries where it has been introduced and is considered a worldwide quarantine organism. In this study, protein markers for differentiating populations of this nematode were identified by studying differences among four selected Iberian and one American population. These populations were compared by quantitative proteomics (iTRAQ). From a total of 2860 proteins identified using the public database from the B. xylophilus genome project, 216 were unambiguous and significantly differentially regulated in the studied populations. Comparisons of their pairwise ratio were statistically treated and supported in order to convert them into discrete character states, suggesting that 141 proteins were not informative as population specific markers. Application of the Character Compatibility methodology on the remaining 75 proteins (belonging to families with different biological functions) excludes 27 which are incompatible among them. Considering only the compatible proteins, the method selects a subset of 30 specific unique protein markers which allowed the compared classification of the Iberian isolates. This approach makes it easier search for diagnostic tools and phylogenetic inference within species and populations of a pathogen exhibiting a high level of genetic diversity.

Potential Insect Vectors of Bursaphelenchus spp. (Nematoda: Parasitaphelenchidae) in Spanish Pine Forests

Potential insect vectors of Bursaphelenchus xylophilus (PWN) were studied. Pathways of introduction of PWN from Portugal to Europe, through Spain, were determined and traps were located in pine stands sites along the pathways. 19 Cerambycidae, 12 Scolytidae, 12 Buprestidae and 10 Curculionidae species have been found. Trapped insects were examined for the presence of nematodes under their elytra. Nematodes were found on Arhopalus ferus, Spondylis buprestoides, Hylastes ater, Hylurgus lingniperda, Orthotomicus erosus, Pityogenes bidentatus, Tomicus piniperda, Hylobius abietis and Pissodes validirrostris specimens. Monochamus galloprovincialis was the most important insect species, representing a risk for the introduction of the PWN in Spanish pine forests; Cerambycidae and Curculionidae species, were taken into account because they have been reported as vectors of other Bursaphelenchus spp.