2015 Volume 45 Issue 2 Pages 101-114
We developed a very simple method for simultaneous detection and discrimination of the principal Pratylenchus and Meloidogyne species in nematode communities collected from Japanese fields. The method consists of a single polymerase chain reaction (PCR) and polyacrylamide gel electrophoresis (PAGE) process using group-specific primers (PNem-F and PNem-R1, 2) constructed from the rDNA-ITS region. DNA from the target species (nine Pratylenchus species and three Meloidogyne species) was amplified with sufficient specificity, and most PCR products could be clearly separated by PAGE using a commercial precast polyacrylamide gel. Although these primers also amplified the DNAs of cyst nematodes and Scutellonema brachyurum, as the sizes of these PCR products were larger than those of the target species, detection and discrimination of the target species bands was not hindered. Even when two or more target species coexisted in a community, these could easily be detected and distinguished, simultaneously and accurately. We confirmed that the combination of a group-specific primer with a community analysis technique was effective for nematode diagnosis. Thus, this method could be a useful tool for the diagnosis of Pratylenchus and Meloidogyne species in Japanese fields.
