Poster: Diseases of Plants: Disease Detection & Diagnosis
Real-time and conventional PCR assays for identifying the stubby root nematode Paratrichodorus allius
G. YAN (1), D. Huang (2), G. Yan (2) (1) North Dakota State University, U.S.A.; (2) North Dakota State University, Department of Plant Pathology,, U.S.A.
Stubby root nematode (SRN), Paratrichodorus allius, is an important pest on many crops, particularly, on potato due to its ability to transmit Tabacco rattle virus causing corky ringspot disease on tubers. Nematode surveys revealed that SRN is present in many fields on different crops in North Dakota and Minnesota. Detection and identification of SRN are important for effective disease management. Protocols of P. allius identification by real-time and conventional PCR with species-specific primers were developed. Primers were designed from internal transcribed spacer of rDNA based on sequences of P. allius and six closely related Paratrichodorus spp. A single amplicon (246 bp) was produced from DNA of P. allius only, but not from DNA of two other SRN species and 21 non-SRN species. Detection sensitivity analysis indicated that an equivalent of 1/20 of a nematode could be detected by conventional PCR and an equivalent of 1/1000 of a nematode by real-time PCR with Cq value at 32.6±0.5. The PCR assays were tested with SRN from 20 soil samples collected from various fields with five crops in ND and MN, and they successfully amplified DNA from all samples. These SRNs were confirmed to be P. allius by sequencing of three genomic regions. The PCR assays are rapid and reliable and can be used in P. allius identification using nematode individuals. This finding will help develop a real-time PCR assay for quantification of P. allius directly from DNA extracts of field soils.