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Infection and Immunity, September 2002, p. 4798-4804, Vol. 70, No. 9
0019-9567/02/$04.00+0     DOI: 10.1128/IAI.70.9.4798-4804.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.

Decreased Electroporation Efficiency in Borrelia burgdorferi Containing Linear Plasmids lp25 and lp56: Impact on Transformation of Infectious B. burgdorferi

Matthew B. Lawrenz,1,2,3 Hiroki Kawabata,4 Joye E. Purser,1,2 and Steven J. Norris1,2,3*

Graduate School of Biomedical Sciences,1 Departments of Pathology and Laboratory Medicine,2 Microbiology and Molecular Genetics, Medical School, University of Texas Health Science Center, Houston, Texas 77225-0708,3 National Institute of Infectious Diseases, Tokyo, Japan4

Received 14 March 2002/ Returned for modification 11 April 2002/ Accepted 23 May 2002

The presence of the linear plasmids lp25 and lp56 of Borrelia burgdorferi B31 was found to dramatically decrease the rate of transformation by electroporation with the shuttle vector pBSV2, an autonomously replicating plasmid that confers kanamycin resistance (P. E. Stewart, R. Thalken, J. L. Bono, and P. Rosa, Mol. Microbiol. 39:714-721, 2001). B. burgdorferi B31 clones had transformation efficiencies that were either low, intermediate, or high, and this phenotype correlated with the presence or absence of lp25 and lp56. Under the conditions utilized in this study, no transformants were detected in clones that contained both lp25 and lp56; the few kanamycin-resistant colonies isolated did not contain pBSV2, indicating that the resistance was due to mutation. Intermediate electroporation rates (10 to 200 colonies per µg of DNA) were obtained with B31 clones that were either lp25- and lp56+ or lp25+ and lp56-. Clones in this group that initially contained lp25 lacked this plasmid in pBSV2 transformants, a finding consistent with selective transformation of lp25- variants. High transformation rates (>1,000 colonies per µg of DNA) occurred in clones that lacked both lp25 and lp56. Sequence analysis indicated that lp25 and lp56 contain genes that may encode restriction and/or modification systems that could result in the low transformation rates obtained with strains containing these plasmids. The previously reported correlation between lp25 and infectivity in mice, coupled with the barrier lp25 presents to transformation, may explain the difficulty in obtaining virulent transformants of B. burgdorferi.

* Corresponding author. Mailing address: Department of Pathology and Laboratory Medicine, University of Texas Medical School at Houston, P.O. Box 20708, Houston, TX 77225-0708. Phone: (713) 500-5338. Fax: (713) 500-0730. E-mail: Steven.J.Norris{at}uth.tmc.edu.

Editor: D. L. Burns

Infection and Immunity, September 2002, p. 4798-4804, Vol. 70, No. 9
0019-9567/02/$04.00+0     DOI: 10.1128/IAI.70.9.4798-4804.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.



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