PLASMID-HOMOLOGOUS SEQUENCES IN THE CHROMOSOME OF PLASMIDLESS COXIELLA-BURNETII SCURRY Q217

Chromosomal DNA from Coxiella burnetii Scurry Q217 was screened for th e presence of plasmid-homologous sequences, Total DNA from Scurry Q217 was digested with NotI, and the resulting DNA fragments were separate d by contour-clamped homogeneous electric field pulsed-field gel elect rophoresis (CHEF-PFGE). Following hybridization with biotin-labeled Qp H1 plasmid as a probe, two DNA fragments of 40 and 170 kb were identif ied as targets, These fragments were cloned, and subclones containing QpH1-homologous sequences were completely sequenced, The physical mapp ing of DNA fragments was achieved by PCR with primers derived from adj acent fragments, and a total of 18,360 bp was sequenced, Within the QD H1-homolooous region spanning 16,624 bp, homology was as high as 99%. Deletions were identified within EcoRI fragments A(H)-C-H-K-H-B-H (13, 490 bp) and J(H)-G(H)-E-H-L+-D-H (6,509 bp) and in fragment A(H) alone (619 bp). An insertion of 744 bp was identified within the JD(C) regi on of Scurry Q217. A search for putative coding regions identified a t otal of 17 open reading frames (ORFs). Compared to plasmid QpH1, 6 ORF s were identical, 5 ORFs were different in size, 6 ORFs were newly gen erated, and 25 ORFs were lost, It was found that plasmid-homologous se quences in Scurry Q217 were of chromosomal origin.