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Streptococcus pneumoniae is a leading cause of pneumonia,
bacterial meningitis, bronchitis, sinusitis, and acute otitis media
in humans. The rapid emergence of macrolide resistance has complicated
treatment for these infections. The two major mechanisms of pneumococcal
macrolide resistance are target modification by erythromycin-resistant
methylase (ermAM), and a macrolide efflux mechanism, the mefE gene
product. Another gene, mel, is located downstream of mefE and the
two genes are cotranscribed as an operon. Population-based surveillance
of all invasive pneumococcal isolates in metropolitan Atlanta indicates
that this efflux mechanism accounts for a vast majority of erythromycin
resistance in S. pneumoniae. Characterization of the genetic locus
containing mefE and mel revealed a 5.5 or 5.4 kb transposon-derived
chromosomal insert indicated as mega (macrolide efflux genetic assembly.)
Two types of mega inserts were found in S. pneumoniae, designated
Type I and Type II, with the latter having a 99 bp deletion in the
intergenic region between mefE and mel. In S. pneumoniae serotype
14, 95% of the mega elements are type II. The specific contributions
of mefE and mel in the macrolide efflux mechanism of resistance
are not known. To analyze the specific role of mefE in macrolide
resistance, deletion mutants of S. pneumoniae have been generated
in GA16638, a serotype 14 strain that has an erythromycin minimal
inhibitory concentration (MIC) of 16 µg/ml. These mutants
were characterized by determining their loss of resistance to erythromycin
or their MIC levels. Additionally, constructs for insertion-duplication
mutagenesis in mel have been generated. These constructs will be
transformed into GA16638 and mutants will be similarly analyzed
for loss of resistance by determining MIC levels.
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