AAC Accepts, published online ahead of print on 26 October 2009

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Antimicrob. Agents Chemother. doi:10.1128/AAC.00499-09
Copyright (c) 2009, American Society for Microbiology and/or the Listed Authors/Institutions. All Rights Reserved.

Characterization of m46.1, the Main Streptococcus pyogenes Element Carrying mef(A) and tet(O) Genes

Andrea Brenciani, Alessandro Bacciaglia, Carla Vignaroli, Armanda Pugnaloni, Pietro E. Varaldo, and Eleonora Giovanetti^*

Departments of Biomedical Sciences, Section of Microbiology, and Molecular Pathology and Innovative Therapies, Section of Histology, Polytechnic University of Marche, 60020 Ancona, Italy

^* To whom correspondence should be addressed. Email: e.giovanetti{at}univpm.it.

m46.1, the recognized representative of the most common variant of mobile, prophage-associated genetic elements carrying resistance genes mef(A) (efflux-mediated erythromycin resistance) and tet(O) (tetracycline resistance) in Streptococcus pyogenes, was fully characterized. Sequencing of the m46.1 genome (55,172 bp) demonstrated a modular organization typical of tailed bacteriophages. Electron microscopic analysis of mitomycin-induced m46.1 revealed phage particles with the distinctive icosahedral head and tail morphology of Siphoviridae. The chromosome integration site was within a 23S rRNA uracil methyltransferase gene. BLASTP analysis revealed that the proteins of m46.1 had high amino acid sequence similarities to proteins from other prophages, especially 10394.4 of S. pyogenes and Sa04 of S. agalactiae. Phage DNA was present in the host cell both as a prophage and as free circular DNA. The lysogeny module appears to have been split due to the insertion of a segment containing tet(O) (from ICE 2096-RD.2) and mef(A) (from a Tn1207.1-like transposon) into the unintegrated phage DNA. The phage attachment sequence lies in the region between tet(O) and mef(A) in the unintegrated form. Thus, whereas in this form tet(O) is 5.5 kb upstream of mef(A), in the integrated form tet(O), lying close to the right end of the prophage, is 46.3 kb downstream of mef(A), which lies close to the left end of the prophage.