Horizontal Transfer of Tetracycline Resistance among Chlamydia spp. In Vitro

Members of the chlamydiae are obligately intracellular bacteria that cause serious diseases in a wide variety of hosts (19). In humans, Chlamydia trachomatis causes trachoma and a variety of sexually transmitted conditions, diseases that affect millions of people around the world (25). Two related species, Chlamydia suis and Chlamydia muridarum, cause diseases of mucosal membranes in pigs and mice, respectively. A second genus within the chlamydiae includes the more distantly related guinea pig pathogen Chlamydophila caviae and the human pathogen Chlamydophila pneumoniae. While the members of the chlamydiae are generally similar in many aspects of basic biology, including genome order and gene content, there are differences in their intracellular survival strategies. One of these differences is the ability to form fusogenic inclusions. Wild-type strains of the Chlamydia spp. form intracellular vacuoles (termed inclusions) that undergo homotypic fusion (30), while no tested strains of these species form inclusions that fuse with inclusions of Chlamydophila caviae or Chlamydophila pneumoniae (23) (see Fig. ​Fig.1)1) (unpublished data). FIG. 1. Immunofluorescence images of McCoy cells coinfected with C. suis R19/tetR and C. muridarum MoPn/oflR (A) or with C. suis R19/tetR and Chlamydophila caviae GPIC/rifR (B). In both panels, C. suis R19/tetR is labeled red, while the alternate species is labeled ... The genome sequences of most chlamydial species do not contain examples of recent horizontal acquisition of DNA. To date, the only reported example of a genomic island within any species of Chlamydia is the tet(C) island of C. suis. This island contains a C. suis-specific insertion element (IScs605), plasmid sequence with roots in gram-negative bacteria, and a tet(C) resistance gene (10). The tetracycline-resistant strains are common in swine herds in the United States (1) and recently have been identified in pigs from Italy (8). The strains carrying this island are the only examples of naturally acquired antibiotic resistance in any chlamydial species. The C. suis tetracycline-resistant strains are also resistant to doxycycline, one of two contemporary front-line drugs of choice against chlamydial infection in humans. Because of its low cost, tetracycline is also used to treat millions of cases of trachoma in developing countries (20). Although there are reports of drug resistance in strains collected from patients suffering treatment failure, documented stable homotypic drug resistance to antibiotics used for treating acute human chlamydial infections is controversial (18, 24, 28). Transfer of a stable tetracycline-resistant phenotype to human clinical chlamydiae would represent a significant public health challenge. The very limited examples of horizontally acquired DNA in the chlamydiae support a hypothesis that recombination might be rare in this system. However, accumulating sequence data indicate that Chlamydia spp. are actively recombinogenic within a species. Early sequencing studies identified ompA variants that encode protein sequences from different classical serovars and showed that recombination may also occur at other locations in the chromosome (2, 11, 17, 21). Additionally, recent studies demonstrated conclusively that lateral gene transfer can be selected for in C. trachomatis in cell culture following coinfection with strains that have dissimilar drug resistance markers (6, 7). The mechanism of transfer in any of these recombination events remains to be elucidated. In order to explore possible tools for the development of a workable genetic system for directed mutagenesis of Chlamydia DNA, and to investigate the possibility of tetracycline resistance acquisition in human strains of C. trachomatis, we conducted in vitro recombination experiments using rifampin (rifampicin)-, ofloxacin-, and tetracycline-resistant strains as donors and recipients. Our results demonstrate that recombination can be readily demonstrated within and among species of the genus Chlamydia and that tetracycline resistance can be manifested by C. trachomatis following coculture with tetracycline-resistant C. suis strains. Tetracycline resistance was not successfully transferred to Chlamydophila caviae, suggesting a biological barrier that does not allow recombination across these genera.