TABLE CONTENTS

Molecular Characterization of Bursaphelenchus (Nematoda:
Parasitaphelenchidae) spp. Distributed in Korea Based on ITS and D2D3
in rDNA Sequence Analysis
Han, H., B-Y. Han, Y-J. Chung & S-C. Shin
Division of Forest Insect Pests and Diseases, Department of Environmental Forest Korea Forest Research
Institute, Seoul, Korea (ROK), 130-712.
Pine wood nematode (PWN), Bursaphelenchus xylophilus, is a causal organism in inducing
pine wilt disease (PWD) in many varieties of pine trees. PWD was first introduced to Korea
in 1988, but the damage increased dramatically since 2000. Recently PWD was newly
reported in Korean pine trees (Pinus koraiensis) and is considered one of the most important
forest diseases in Korea. Fifteen isolates of B. xylophilus, 3 isolates of B. mucronatus, and 2
unidentified Bursaphelenchus spp. were collected from different geographical locations and
hosts in Korea, and were characterized by ITS and D2D3 rDNA sequence analysis. Template
DNA was prepared by DNA extraction from a single female nematode. ITS and D2D3
regions were amplified by PCR and followed by cloning and sequence. As a result, all the
sequences of ITS and D2D3 from B. xyophilus isolates were identical and there is no
intraspecific variation. However, 2 genotypes of B. mucronatus were found, one from P.
thunbergii was East Asia type and the other from P. koraiensis was European type. The data
of two unknown species of Busaphelenchus sp.(A) and Bursaphelenchus sp.(B) were closely
related to B. tusciae and B. lini, respectively, which was also supported by morphological
characterization. The amplified size of ITS and D2D3 for all isolates were 950bp and 750bp,
respectively. The only exception was for Busrsaphelenchus sp.(B). because it has 1.2 kb in
ITS size. ITS-RFLP data also discriminated between different species and genotypes by using
5 enzymes of Hinf I, Alu I, Msp I, Hae III, Rsa I. Bursaphelenchus conicaudatus was used as
the control species for this experiment, and informative nucleotide sequences of
Bursaphelenchus were downloaded from GenBank in NCBI. All DNA sequence data were
aligned by using Clustal W program and molecular phylogenetic analysis was performed by
PAUP*4.0.
5 th International Congress of Nematology, 2008 58