Identification of Anguina funesta from AnnualRyegrass Seed Lots in Oregon

摘要

In 2010, seed galls containing Anguina sp. were isolated from 14 annual ryegrass(Lolium multiflorum) seed lots submitted for phytosanitary testing. To identify thespecies present, the ITS1 region of the ribosomal DNA of the nematodes from theseed lots was analyzed using a PCR-RFLP method. All nematodes produced asingle 540-bp DNA amplicon, which was digested with three restriction enzymes,HaeIII, HinfI, and TaqI. The resulting RFLP patterns matched those of A. funesta.To confirm these results, 525 bp of the DNA amplicon was analyzed by DNAsequencing and BLAST analysis, which verified the sequence was identical to A.funesta (Genbank Accession nos. AF363095, AF363096, and AF363104). Becauseof the association of A. funesta with Rathayibacter toxicus, a second PCR test wasconducted to determine if the bacterium was present in the seed lots. A 200-bpDNA amplicon was amplified from two seed galls, sequenced, and subjected toBLAST analysis. Analysis of the entire DNA sequence failed to identify thebacterium present, although testing by USDA-APHIS verified the bacterium wasnot R. toxicus. This is the first report of A. funesta in the US; R. toxicus was notfound with this detection.