Current Japanese clinical practice involves the usage of large amounts of new
macrolides such as
clarithromycin and
roxithromycin for the treatment of
diffuse panbronchiolitis, Helicobacter pylori and Mycobacterium avium complex
infections. In this study, the phenotypes, genotypes, and
macrolide resistance mechanisms of
macrolide-inactivating Escherichia coli recovered in Japan from 1996 to 1997, were investigated. The isolation rate of
erythromycin A highly-resistant E. coli (MIC > or = 1,600 microg/ml) in Japan slightly increased from 0.5% in 1986 to 1.2% in 1997. In six
macrolide-resistant strains, recovered from the strains collected for this study during 1996 to 1997, the inactivation of
macrolide could be detected with or without added
ATP in the assay system. The appearance of
erythromycin A-inactivating
enzyme independent of
ATP was novel from Japanese isolates, and the 1H NMR spectra of
oleandomycin hydrolyzed by the three
ATP-independent isolates were examined. It was clearly shown that the
lactone ring at the position of C-13 was cleaved as 13-H signal in aglycon of
oleandomycin upper shifted. These results suggested the first detection of
macrolide-
lactone ring-
hydrolase from clinical isolates in Japan. These results suggested the first detection of an
ATP-independent
macrolide-hydrolyzing
enzyme from Japanese clinical isolates. Substrate specificity of the
macrolide-hydrolyzing
enzyme was determined with twelve
macrolides including the newer members of this group and it was found that not only
erythromycin A but also the new
macrolides, such as
clarithromycin,
roxithromycin, and
azithromycin were inactivated. The NMR data, broad spectrum of activity, and independence of co-
enzyme supported our naming of the
enzyme as a
macrolide esterase. PCR methodology was employed to detect an ereB-like gene from the 3 isolates producing
macrolide esterase, and one of these was subsequently shown to contain both ereB-like and ermB-like genes. It was also clearly shown that the other three isolates, which inactivated
macrolide in the presence of
ATP, had an mphA-like gene.