Chloramphenicol acetyltransferase as a selection marker for chlamydial transformation

BACKGROUND: Chlamydia is a common bacterial pathogen responsible for many diseases. Methods for transforming this important organism using a β-lactamase as a selection marker have been developed very recently. However, the National Institutes of Health Guidelines for Research Involving Recombinant DNA Molecules do not permit transformation experiments with β-lactamase gene-containing vectors for certain human chlamydial pathogens. Therefore, a different selection marker is urgently needed for transformation of those chlamydiae. RESULTS: After transformation of plasmid-free Chlamydia trachomatis with pGFP:SW2, which carries a β-lactamase and a chloramphenicol acetyltransferase gene fused to a green fluorescence protein gene, transformants were obtained by selection with either ampicillin or chloramphenicol. Stable chloramphenicol-resistant, but ampicillin-sensitive, transformants were obtained using a pGFP:SW2 derivative without the β-lactamase. All transformants expressed green fluorescence protein and had glycogen synthesis activity restored. CONCLUSIONS: Chloramphenicol resistance may be used as a selection marker for genetic experiments in Chlamydia. This eliminates the requirement for the use of β-lactamase, of which dissemination to some C. trachomatis serovars may jeopardize clinical treatment of chlamydial infections in pregnant women. Chloramphenicol acetyltransferase may also serve as a useful secondary selection marker for genetic analyses in β-lactamase-transformed chlamydial strains.