Articles
PATHOVAR-SPECIFIC DETECTION OF XANTHOMONAS CAMPESTRIS IN HYACINTHS
This bacterial disease is easily spread in the field, or by handling the bulbs during sorting (Van Doorn et al., 1993). To make sure whether certain symptoms are caused by this yellow pigmented microbe quick, specific and sensitive detection methods are needed.
This problem was tackled by raising monoclonal antibodies against partly purified XCH fimbriae, filamenteous surface fibrils which may play a role in attaching XCH to hyacinth stomata (Van Doorn et al., 1994).
We obtained 20 different hybridoma cell-lines which produced monoclonal antibodies recognizing only XCH-isolates.
Two which performed best in DAS-ELISA were selected: LBO-1 recognizes a LPS-fraction of XCH whereas LBO -2 reacts with a part of the XCH fimbriae.
These monoclonals are currently used by the Bulb Inspection Service in Lisse (detection-limit 105 – 106 cells/ml).
In early symptoms of yellow disease on hyacinth leaves (necrotic spots or "spetters") even smaller amounts of XCH bacteria might be present.
More sensitive detection was possible by using the Polymerase Chain Reaction in amplifying the gene, coding for the fimbrial subunit protein of XCH. Primers flanking the central region of this structural gene were developed.
Only XCH-isolates reacted in PCR by showing a DNA-fragment of about 200 basepairs; under laboratory conditions 500 XCH cells could be easily detected.
Further research is directed at adapting and testing the PCR detection method not only for XCH but also for other X.campestris pathovars in (ornamental) plants.
Besides this we are studying the early events of XCH pathogenesis in hyacinths in order to develop means to hamper or even prevent infection.
We think that certain glycolipids are the receptors for the type 4 fimbriae of XCH; in vitro attachment studies in microtiterplates may present evidence for this theory.
Definitive evidence for the function of XCH type 4 fimbriae will be obtained by mutating the structural fim-gen and to test the resulting fimbriae-negative mutants for pathogenicity in planta.