Spectrophotometer, paper chromatography, and liquid chromatography were used to isolate and characterize the substances associated with tissue browning of Japanese red pine, (Pinus densiflora) and Japanese black pine (P. thunbergii) inoculated withBursaphelenchus xylophilus or B. mucronatus. Discoloration, as determined by optical density, and numbers of B. xylophilus was highest near the inoculation site and decreased with increasing distance from that point. Absorption spectra of water extracts from pines infected with B. xylophilus had a distinct peak at 278 nm. The absorption at 278 nm decreased when the extract was oxidized and shifted to 287 nm when 1N NaOH was added, suggesting that the extract contained polyphenolic substances. The increase in browning with increase in nematode population indicates that these substances were related to the nematode infection. B. xylophilus-inoculated seedlings contained much greater amounts of phenolics than B. mucronatus-inoculated and control seedlings. A catechin-like substance was the most abundant polyphenolic associated with large numbers of B. xylophilus. Changes in the amount of condensed tannin, the polymer of catechin, were surveyed in relation to the numbers of nematodes in pine seedlings. Jpn. J. Nematol. 33 (2), 45-56 (2003).

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PCR-RFLP analyses, reproduction tests on sweet potato and taro, as well as inter-RFLP-phenotypic hybridization were performed using 20 isolates of Pratylenchus coffeaeand two isolates of P. penetrans to identify relationships between these nematodes. F194 and F195 primers yielded rDNA ITS region amplification products of ca. 1, 080 bp (Japanese and Indonesian isolates), and ca.1, 020 bp (Guatemalan isolate) that were digested using Hinf I, Alu I, Dde I, and Hha I restriction endonucleases. There were four distinct DNA fragment patterns for isolates of P. coffeae that were designated to be RFLP phenotypesA, B, C and D. All the Japanese isolates corresponded to RFLP-phenotypes A, B or C. The Indonesian isolate was phenotype A and the Guatemalan isolate was phenotype D. The Pf/Pi ratios 55 days or 90 days after inoculation to sweet potato in a greenhouse experiment differed between phenotypes. Phenotype A isolates had the greatest Pf/Pi of 1.1 - 17.3. Phenotype B and C had values of 0.05 - 0.40 and 0.12 - 0.36 respectively. Reproduction on taro in a greenhouse experiment differed among phenotypes. Phenotypes A and B had greater reproduction (Pf/Pi>1) than did phenotypes C (Pf/Pi =0). Laboratory hybridization tests between phenotypes A × B, A × C, and B × C generated abundant F1 hybrids, though reciprocal breedings of A × D and A × P. penetrans failed to reproduce. Inbreedings of Fis obtained from A × C were rarely successful, those from A × B generated a few F_2s in 1/3 of replications, and those from B × C generated F_2s in 2/3 of replications, suggesting incomplete reproductive isolation among the three rDNA RFLP phenotypes of P. coffeae. The allopatric distribution accompanied by distinct host adaptation, thermal adaptation and incomplete reproductive isolation exemplify microevolution in the P. coffeae species complex. Jpn. J. Nematol. 33 (2), 57-76 (2003).

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[in Japanese]

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