FruitDisease - Root rot, diagnostics

Root rot diagnostics


PCR based detection

The diagnostic polymerase chain reaction (PCR) test and associated protocols for P. idaei and other Phytophthoras were designed and optimised in a research environment at SCRI and were the subject of a European patent application (No. 96303105.9) held by Mylnefield Research Services. The test has been used for many years and is based on PCR primers designed to species-specific variation in the sequences of the internal transcribed spacer (ITS) sequences of rDNA (e.g. Cooke et al 2000; Bonants et al 1997). There are very few sequence differences between P. idaei and P. cactorum (Cooke et al 1996) thus the same primers are currently used to detect both agents and digestion with a restriction enzyme used to discriminate the two organisms if needed. The test has been optimised as a nested assay with the first round PCR primers based in the flanking 18S and 28S rDNA regions. However, SCRI is continuing to seek improvements in the test, specifically relating to throughput and validation of the test in raspberry. Such improvements will benefit the application in the testing of high health status plants (see below).

Detection of Phytophthoras in high health status material

SCRI is the sole source of raspberry nuclear stocks in the UK and tests for a range of pathogens in this material, which is thus the key stage for the whole industry in their use of clean propagation material. New cultivars entering the certification system, by necessity, originate from breeding material in the field and the ongoing risk of contamination from such sources must be managed. Effective, validated diagnostic assays for Phytophthoras are critical to ensuring this. More details of this work and the raspberry root rot diagnostic service we provide can be found at our FruitHealth.co.uk web site.

References

Bonants PJM, Hagenaar De Veerdt M, Van Gent-Pelzer MP, Lacourt I, Cooke DEL & Duncan JM (1997) Detection and identification of Phytophthora fragariae Hickman by the polymerase chain reaction. European Journal of Plant Pathology 103:345-355.
Cooke DEL, Kennedy DM, Guy DC, Russel J, Unkles SE & Duncan JM (1996) Relatedness of Group I species of Phytophthora as assessed by RAPDs and sequences of ribosomal DNA. Mycological Research 100:297-303.
Cooke DEL, Drenth A, Duncan JM, Wagels G & Brasier CM (2000) A molecular phylogeny of Phytophthora and related Oomycetes. Fungal Genetics and Biology 30:17-32.