PCR detection and identification of Phymatotrichopsis omnivora
M. ARIF (1), S. M. Marek (1), F. M. Ochoa Corona (1), C. Young (2), C. D. Garzon (1)
(1) Department of Entomology and Plant Pathology, Oklahoma State University, Stillwater, OK, U.S.A.; (2) The Samuel Roberts Noble Foundation, Ardmore, OK, U.S.A.
Phytopathology 100:S7

The soilborne pathogen, Phymatotrichopsis omnivora is the causal fungus of cotton root rot of numerous dicots in the southwestern United States and Mexico. Disease diagnosis depends on the presence of characteristic mycelial cords on the roots of wilted plants, which are not always obvious. Early, accurate and sensitive detection of P. omnivora in affected plant tissues is needed by plant health officials for inspection of products from quarantined states, and locally, by extension specialists to predict disease outbreaks and identify reservoir hosts. Specific PCR primers recognizing conserved rDNA-ITS sequences were designed based on an alignment of 144 sequences from P. omnivora isolates collected throughout North America. Three primer pairs, PO1 (415 bp product), PO2 (499 bp product) and PO3 (146 bp product), were validated in silico against published sequences and in vivo against infected plant samples. PCR products were cloned and sequenced to confirm identity. All primer sets allowed early detection of infected, asymptomatic plants. PO1, PO2 and PO3 detected 5 × 10^–7, 5 × 10^–8 and 5 × 10^–8 ng per µl of the P. omnivora target DNA, respectively. PO3 also was compatible with qPCR and thermostable helicase dependent amplification (tHDA) assays. The described PCR assays should be useful for rapid, sensitive diagnosis, quantification of infection in resistance breeding programs and agricultural biosecurity.