2.2.19
PTHA HOMOLOGUE-MEDIATED DNA ARRANGEMENTS IN XANTHOMONAS CITRI

S TSUYUMU, H KANAMORI and H NAGAI

Faculty of Agriculture, Shizuoka University, 836 Ohya, Shizuoka, Japan 422

Background and objectives
In several microorganisms, dynamic arrangements of DNA have been reported to be involved in the rapid adaptation at population level upon exposure to abrupt change in environmental conditions. It is important to determine the presence of such arrangements of DNA in plant pathogens for the study of the evolution and life cycle of plant pathogens.

Materials and methods
In Xanthomonas citri strain NA-1, three regions showed homology with pthA [1], a gene reported to be responsible for the formation of canker symptom on Citrus plants. All of these three regions (APL1, APL2 and APL3) were shown to exist on the plasmids in strain NA-1. These regions were cloned and their DNA sequences determined. Using this information, the arrangements of DNA at these regions were studied.

Results and conclusions
DNA sequences of APL1, APL2 and APL3 were found out to be identical up to 240nt upstream and 111nt downstream of their open reading frames (ORF), except for the 102 bp tandem repeating sections within their ORF. In the repeating sections, the number of repeating units were distinguishable and minor differences were found at the variable regions within the repeating units. When the transformants of each APL in a Tn5-induced nonpathogenic mutants were inoculated into the leaves of C. natsudaidai, only the APL1 transformant formed canker symptoms to the level of the wild type. The APL2 transformant also formed canker symptoms but it took twice as long compared with the wild type. The APL3 transformant did not cause canker symptoms 1 month after the inoculation. Thus, the importance of the specific arrangement of the repeating units for the formation of canker symptom was confirmed. Arrangements of DNA at these APLs were shown to occur at relatively high frequency during transfer on YP medium, especially when the culture was briefly exposed to high temperature immediately after transferring. Fifty per cent of the strains that showed alteration in the sizes of the plasmids resulted in the alteration in restriction enzyme cleavage patterns of APLs. From the strain which showed fused plasmid by apparent recombination between APL regions, several strains, which reverted to the original sizes of the plasmids, were isolated. In such revertant strains the restriction enzyme cleavage pattern of APLs were same as that of the wild type. Thus, arrangement by recombination at these homologous regions was found to occur. Furthermore, APLs commonly posses identical inverted repeats at their ends. Transposition of DNA segment with these inverted repeats at its ends was shown to occur experimentally. Thus, arrangement by transposition using the inverted repeats was also found to occur.

References
1. Swarup S, Feyer R, Brlansky RH, Gabriel DW, 1991 Phytopathology 81, 802-809.