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Applied and Environmental Microbiology, December 1999, p. 5378-5385, Vol. 65, No. 12
0099-2240/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.

Influence of Different Functional Elements of Plasmid pGT232 on Maintenance of Recombinant Plasmids in Lactobacillus reuteri Populations In Vitro and In Vivo

Nicholas C. K. Heng,1 Judith M. Bateup,1 Diane M. Loach,1 Xiyang Wu,1 Howard F. Jenkinson,2 Mark Morrison,3 and Gerald W. Tannock1,*

Department of Microbiology, University of Otago, Dunedin, New Zealand1; Department of Oral Science, University of Bristol Dental School, Bristol, United Kingdom2; and Department of Animal Science, University of Nebraska---Lincoln, Lincoln, Nebraska 68583-09083

Received 2 August 1999/Accepted 8 October 1999

Plasmid pGT232 (5.1 kb), an indigenous plasmid of Lactobacillus reuteri 100-23, was determined, on the basis of nucleotide and deduced protein sequence data, to belong to the pC194-pUB110 family of plasmids that replicate via the rolling-circle mechanism. The minimal replicon of pGT232 was located on a 1.7-kb sequence consisting of a double-strand origin of replication and a gene encoding the replication initiation protein, repA. An erythromycin-selectable recombinant plasmid containing this minimal replicon was stably maintained (>97% erythromycin-resistant cells) without antibiotic selection in an L. reuteri population under laboratory growth conditions but was poorly maintained (<33% resistant cells) in the L. reuteri population inhabiting the murine gastrointestinal tract. Stable maintenance (>90% resistant cells) of pGT232-derived plasmids in the lactobacillus population in vivo required an additional 1.0-kb sequence which contained a putative single-strand replication origin (SSO). The SSO of pGT232 is believed to be novel and functions in an orientation-specific manner.

* Corresponding author. Mailing address: Department of Microbiology, University of Otago, P.O. Box 56, Dunedin, New Zealand. Phone: 64-3-479-7734. Fax: 64-3-479-8540. E-mail: gerald.tannock{at}stonebow.otago.ac.nz.

Applied and Environmental Microbiology, December 1999, p. 5378-5385, Vol. 65, No. 12
0099-2240/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.


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