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Journal of General Microbiology 139 (1993), 2485-2493; DOI  10.1099/00221287-139-10-2485
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Transformation of Helicobacter pylori by chromosomal metronidazole resistance and by a plasmid with a selectable chloramphenicol resistance marker

Ying Wang{dagger}, K. Pieter Roos and Diane E. Taylor*

Department of Medical Microbiology and Infectious Diseases. University of Alberta, Edmonton, Alberta, Canada T6G 2H7

*Author for correspondence. Tel. +1 403 492 4777; fax +1 403 492 7521; e-mail userdeht@mts.ucs.

ABSTRACT

SUMMARY: Most strains of Helicobacter pylori are naturally competent for uptake of chromosomal DNA. Transformation frequencies for streptomycin resistance or rifampicin resistance markers ranged from 1 x 10-4 to 1 x 10-3 per viable cell using a plate transformation procedure. Transformation of a metronidazole resistance marker (MtrR) was demonstrated when either a laboratory-derived mutant or a MtrR clinical isolate were used as the source of donor DNA. MtrR was transformed at a frequency of 3 x 10-5 per viable cell. All H. pylori strains tested produce large amounts of DNAase, which may reduce DNA available for transformation. Four H. pylori plasmids were isolated. DNA fragments from H. pylori plasmids were deleted or rearranged when cloned in pUC19 and propagated in Escherichia coli DH5{alpha}. An H. pylori plasmid, pUOA26 which contained a chloramphenicol resistance determinant from Campylobacter coli, was constructed in H. pylori. This plasmid could be successfully introduced by natural transformation only into H. pylori recipients which contained a homologous resident plasmid. Transformation of pUOA26 into plasmid-free cells of H. pylori was achieved by electroporation. Transformation frequencies were 1 x 10-4 transformants per viable cell when plasmid DNA was isolated from the same strain; however, introduction of pUOA26 DNA derived from H. pylori 8091 into a different H. pylori strain, NCTC 11639, resulted in transformation at much lower frequencies (less double equals 1 x 10-7 per viable cell). Changes in plasmid restriction patterns were also noted when pUOA26 was isolated from H. pylori NCTC 11639, suggesting the presence of at least two different DNA restriction and modification systems in H. pylori which may interfere with uptake of plasmid DNA.

{dagger} Present address: Biotechnology Research Institute, 6100 Royalmount Avenue, Montreal, Quebec, Canada H4P 2R2.




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