Foggy D-shaped zone of inhibition in Staphylococcus aureus owing to a dual character of both inducible and constitutive resistance to macrolide-lincosamide-streptogramin B

Objectives: We identified Staphylococcus aureus strains having blunt inhibitory zones around the clindamycin or josamycin discs proximal to the erythromycin disc filled with slight bacterial growth. This ambiguous phenotype was termed as 'macrolide-lincosamide-streptogramin B antimicrobial (MLSB) resistance having a foggy D-shaped inhibitory zone' (fMLS(B)), and we tried to analyse its molecular mechanisms. Methods: Forty-one clinically isolated strains of fMLS(B) S. aureus were studied. The regulatory region of the erm(A) gene, which was found to be the only molecular mechanism of fMLS(B), was sequenced. Then, beta-galactosidase assays were performed to observe their expression patterns through the nucleotide sequential alteration. Results: According to the sequencing electropherogram, a grouping was made of a homogeneous nucleotide sequence group (73.2%) and a heterogeneous nucleotide sequence group (26.8%). The former group was composed of a 25 bp tandem duplication type and a 25 bp tandem triplication type. Their beta-galactosidase activity was similar to that of constitutive MLSB resistance due to its high basal level. Nevertheless, the predicted mRNA secondary structure of the regulatory region maintains the stem-loops of inducible wild-type erm(A), and thereby its inducible character might be expected in vivo. Strains in the latter group were proven to have two different erm(A) genes, and then dual effect of expression was observed. Conclusions: The ambiguous phenotype of fMLS(B) is due to its possessing the dual character of inducible and constitutive expression of erm(A). The dual character is due to having one erm(A) gene of dual character or coexistence of two characterized erm(A) genes simultaneously.