First chromosomal restriction map of Actinobacillus pleuropneumoniae and localization of putative virulence-associated genes

Combined physical and genetic maps of the genomes of Actinobacillus pleurop neumoniae AP76 (serotype 7 clinical isolate) and of A, pleuropneumoniae ATC C 27088 (serotype 1 reference strain) were constructed by using the restric tion endonucleases ApaI, AscI, NotI, and SalI, The chromosome sizes as dete rmined by the addition of estimated fragment sizes were 2.4 Mbp, and both m aps had a resolution of approximately 100 kbp. The linkages between the Apa I, AscI, NotI, and San fragments and their relative positions were determin ed by (i) fragment excision and redigestion and (ii) partial digests of def ined fragments and Southern blot using end-standing probes. The single SalI site within the chromosome of strain A. pleuropneumoniae AP76 was defined as position 1 of the map; for the map of A. pleuropneumoniae ATCC 27088, th e corresponding San site was chosen. Putative virulence-associated genes (a px, omlA, sodA, tbpBA, ureC, and a repeat element) and housekeeping genes ( glyA, metJ, recA, and rhoAP) were positioned on the physical maps and locat ed on the ApaI and NotI fragments of A. pleuropneumoniae serotype reference strains.