Distribution and molecular analysis of mef(A)-containing elements in tetracycline-susceptible and -resistant Streptococcus pyogenes clinical isolates with efflux-mediated erythromycin resistance

Objectives: To analyse the distribution and molecular features of mef(A)-containing elements in a large collection of different Streptococcus pyogenes clinical isolates with efflux-mediated erythromycin resistance. To further characterize a tet(O)-mef(A) element. Methods: Gene detection was carried out by PCR using primers designed from established sequences or from sequences in this study. From a tet(O)-mef(A) element (similar to60 kb), an 11 972 bp region including the tet(O) and mef(A) genes was sequenced. Results: In the tetracycline-susceptible isolates (n =28), the mef(A) gene was contained in a regular Tn1207.1 transposon (7.2 kb), which was inserted into one of two previously described elements, Tn1207.3 (similar to52 kb) or a 58.8 kb chimeric element, both flanked by the comEC gene. In the tetracycline-resistant isolates (n =61), all of which carried the tet(O) gene, the mef(A) gene was part of a variable Tn1207.1-related transposon inserted into unique elements which contained the tet(O) gene similar to2.3 to 5.5 kb upstream of the mef(A) gene and were not flanked by the comEC gene. In the Tn1207.1-like transposon of these tet(O)-mef(A) elements, only msr(D) (orf5) and a modified orf6, in addition to mef(A), were detected by PCR in all isolates tested; while orf1 and orf2 were always undetectable, orf3, orf7 and orf8 were found in variable percentages. In an orf3-positive element, sequencing identified four new open reading frames downstream of the tet(O) gene, followed by three short sequences with homology to sequences of the pneumococcal mega element. Conclusions: The mef(A) gene is carried on different chromosomal genetic elements depending on whether the isolates are susceptible or resistant to tetracycline.