DETECTION OF XANTHOMONAS-CAMPESTRIS PV VESICATORIA ASSOCIATED WITH PEPPER AND TOMATO SEED BY DNA AMPLIFICATION

Detection of Xanthomonas campestris pv. vesicatoria associated with pe pper and tomato seed was achieved by amplification of a DNA fragment o f the bacterium by the polymerase chain reaction. Oligonucleotide prim ers specific for the hypersensitive reaction and pathogenicity (hrp) g ene duster were used in the reaction. The method includes extraction o f total DNA from buffered seed washings to which sodium ascorbate and insoluble polyvinylpolypyrrolidone were added. The hrp fragments were amplified from the DNA preparations if cells of X. c. pv. vesicatoria were added to seed washes. Identification of an hrp fragment as that f rom X. c. pv. vesicatoria was attained by restriction enzyme analyses of the amplified fragment. The minimum number of cells that could be d etected in washes from pepper or tomato seed was from 10(2) to 10(3) c fu per ml. This was about 1,000 times fewer than the minimum number of cells detected with an enzyme-linked immunosorbent assay. The pathoge n was also detected in washes obtained from several lots of naturally contaminated pepper and tomato seed, one of which contained background bacterial microflora greater than 10(7) cfu per g of seed.