3.3.1S
IMPACTS OF MOLECULAR DIAGNOSTIC TECHNOLOGIES ON PLANT DISEASE MANAGEMENT -- PAST, PRESENT AND FUTURE

SA MILLER

Department of Plant Pathology, The Ohio State University, OARDC, Wooster, OH 44691, USA

The era of modern diagnostic technology in plant disease management began in 1976 with the first application of the enzyme-linked immunosorbent assay (ELISA) to plant virus detection. This was soon followed by the development of highly specific monoclonal antibodies (MAbs) against most classes of plant pathogenic microorganisms. The speed and sensitivity of ELISA combined with the specificity of MAbs made possible the rapid and accurate diagnosis of plant diseases, and detection of pathogens in a variety of substrates. Kits were introduced commercially in the mid-1980s that could be completed in 10 minutes, involving only a few simple steps. A decade later, development of more advanced technology resulted in assays that could be completed ten times faster. These assays were targeted to the detection of plant pathogenic fungi, including species of Phytophthora , Pythium , Rhizoctonia and Septoria , as a diagnostic method and a tool in decision making. Information provided by the kits is used to select the appropriate fungicide and also indicate the appropriate time of application. Thus the use of such kits contributes to reducing the environmental impact of fungicides and can be an important component of precision agriculture. However, the costs of development of such tests are high and they are now available for a relatively small number of applications.

ELISA kits in a laboratory format and reagents are now commercially available from several firms worldwide. The markets for these kits are 1) public laboratories operated by universities or government organizations, 2) private, for-profit testing laboratories and corporations, 3) private, non profit organizations, e.g. various crop improvement associations and 4) individual researchers. ELISA and other related antibody-based technologies such as immunofluorescence and dot immunobinding, developed commercially or in the public domain, are widely used in seed and plant indexing and certification programs, as well as in laboratories and clinics for routine diagnosis. Phytophthora assays have been particularly useful in studies of distribution of this pathogen in soils, water and plant root systems.

Practical use of nucleic acid-based detection technologies in plant pathology was limited until the introduction of the polymerase chain reaction (PCR) assay in the late 1980s. Primers with the required degree of specificity, from the strain to the genus level can be designed relatively easily, even with little knowledge of the genome of the target microorganism. Primers may be developed from previously identified nucleic acid probes, from arbitrarily primed DNA markers, and from ribosomal RNA gene sequences, particularly the internal transcribed spacer regions, among others. PCR or reverse transcription (RT) PCR assays have been developed for a wide range of plant pathogens. Multiplex PCR and the reverse dot blot assay, a combination of PCR and DNA hybridization, allow detection of multiple pathogens in a single test. Innovations in detection of amplified products, from the use of fluorescent dyes to solid state DNA microarrays are being introduced. However, the widespread application of PCR-based technologies in plant health management remains at this time constrained by cost, including prohibitive royalty obligations, and relatively complicated assay technology. The latter is likely to be solved in the future but the former remains in question, at least in the near term.