TABLE CONTENTS

Revision of Quarantine Nematodes Reported in Spain
Escuer, M., S.C. Arcos, L. Robertson, M.A. Diez Rojo & A. Bello
Dpto Agroecología, Instituto de Ciencias Agrarias, CCMA, CSIC, C/ Serrano 115 dpdo, Madrid, 28006, Spain
In revising the quarantine status of nematodes in Spain, Ditylenchus dipsaci (Kühn, 1857)
Filipjev, 1936 is widespread on many different crops, including garlic, onion, strawberry,
sugarbeet, cereals and legumes. Reports from other crops and uncultivated areas should be
reviewed. Globodera pallida (Stone, 1973) Berenhs, 1975 is present, the main pathotypes
being Pa2/Pa3 and also Pa1 in Tenerife (Canary Islands). The pathotypes Ro1/Ro4 of G.
rostochiensis (Wollenweber, 1923) Skarbilovich, 1959 occur throughout Galicia, La Rioja,
Mallorca and Tenerife (Canary Islands). Xiphinema rivesi Dalmasso, 1969, a nematode vector
of the Tomato Ring Spot Virus (TomRSV) in grapes in the USA, is widely distributed in the
Spanish peninsula. Although cited, Ditylenchus destructor Thorne 1945; Nacobbus Thorne &
Allen, 1944 and Xiphinema americanum Cobb, 1913 sensu stricto have not been found in
Spain. However there are reports of X brevicollum Lordello & Dacosta, 1961, and X.
pachtaicum (Tulaganov, 1938) Kirjanova, 1951 which are species belonging to the X.
americanum-group. The other quarantine nematodes Aphelenchoides besseyi Christie, 1942;
Bursaphelenchus xylophilus (Steiner & Buhere, 1934) Nickel et al., 1970, Heterodera glycines
Ichinohe, 1952, Hirschmanniella Luc & Goodey, 1964, Longidorus diadecturus Eveliegh &
Allen, 1982, Meloidogyne chitwoodi Golden et al., 1980, M. fallax Karssen, 1996, Nacobbus
aberrans (Thorne, 1935) Thorne & Allen, 1944, Radopholus similis (Cobb,1893) Thorne 1949
(syn.: R. citrophillus Huettel et al., 1984), Xiphinema bricolense Ebarsy et al., 1989, and X.
californicum Lamberti & Bleve-Zacheo, 1979 have not been reported from Spain. Due to the
importance of quarantine measures, an in-depth study of the distribution and epidemiology must
be carried out to increase our knowledge on the existence of biotypes, their host range, and other
plant pathogen interactions as in the case of D. dipsaci and X. rivesi.
Detection and Quantification of Root-lesion Nematodes from Field Soil by
Conventional and Real Time PCR
Qiu, J., B B. Westerdahl & V.M. Williamson
Department of Nematology, University of California, One Shields Avenue, Davis, CA 95616.
It is challenging to detect and quantify nematodes from soil extracts by PCR due to PCR
inhibitors in soil. We have developed a protocol for detection of root lesion nematode
Pratylenchus spp. in soil extracts using conventional PCR with specific primers. Nematodes
are extracted from soil using Baermann funnels and centrifugal flotation in a sucrose gradient
that alleviates the inhibition and increases the sensitivity of PCR detection. The nematode
extract is then digested with proteinase K and the digestion used as a DNA template in the
PCR assay. With this protocol and species-specific primers designed from ITS sequences
obtained through cloning and sequencing technologies, we diagnosed P. penetrans, P.
vulnus, P. scribneri and P. thornei, four major species in California fields by multiplex PCR.
Analysis of unknown samples indicated that the detection was sensitive and specific. It
detected samples at levels as low as a single target nematode among hundreds of thousands of
other plant parasitic and free-living ones. We quantified P. vulnus from orchard soil by real
time PCR. Grinding prior to proteinase K digestion was used to prepare nematode DNA.
DNA quantity per female is about 1.5 times higher than for other stages, but quantities are
similar for males and juveniles. All stages are present in California field and females
5 th International Congress of Nematology, 2008 124