Tn2009, a Tn916-like element containing mef(E) in Streptococcus pneumoniae

Abstract

The association between the macrolide efflux gene mef(E) and the tet(M) gene was studied in two clinical strains of Streptococcus pneumoniae that belonged to serotypes 19F and 6A, respectively, and that were resistant to both tetracycline and erythromycin. The mef(E)-carrying element mega (macrolide efflux genetic assembly; 5,511 bp) was found to be inserted into a Tn916-like genetic element present in the chromosomes of the two pneumococcal strains. In both strains, mega was integrated at the same site, an open reading frame identical to orf6 of Tn916. The new composite element, Tn2009, was about 23.5 kb and, with the exception of the tet(M)-coding sequence, appeared to be identical in both strains. By sequencing of the junction fragments of Tn2009 at the site of insertion into the chromosome, it was possible to show that (i) the insertion site was identical in the two clinical strains and (ii) the integration of Tn2009 caused a 9.5 kb-deletion in the pneumococcal chromosome. It was not possible to detect the conjugal transfer of Tn2009 to a recipient pneumococcal strain; however, transfer of the whole element by transformation was shown to occur. It is possible to hypothesize that Tn2009 relies on transformation for its spread among clinical strains of S. pneumoniae.

FIG. 1.

Schematic representation of Tn2009, composed of the mega element inserted into a Tn916-like transposon. The structure of mega is boxed, and ORFs are indicated by gray arrows. Tn916 is indicated by a black bar, and white arrows represent the relevant ORFs. The exogenous sequence at the right end of Tn2009 is in gray. The flanking chromosome of S. pneumoniae is represented by a black line; striped arrows indicate ORFs, designed according to the homologous ORFs of strain R6. Short black arrows indicate the positions of the oligonucleotide primers used for PCR amplification and sequencing.

FIG. 2.

Junctions of Tn2009 with the chromosome in clinical isolates PN150 and PN34 and transformant MF58. The sequences belonging to Tn2009 and the exogenous sequences are shown in boldface and boxed. The nucleotides at the left and right ends of Tn916 are italicized. At the left junction, the 6 nucleotides representing the coupling sequences are underlined. At the right junction, the 63- or 61-nucleotide exogenous sequences are indicated.