Identification of a Mutation Associated with Erythromycin Resistance in Bordetella pertussis : Implications for Surveillance of Antimicrobial Resistance

Abstract

Erythromycin treatment failures and in vitro resistance of Bordetella pertussis have been reported on several occasions in the past few years, but the mechanism of resistance has not been described. One potential mechanism, genetic modification of the erythromycin-binding site on the 23S rRNA of the 50S ribosomal subunit, has been observed in other bacteria. To explore this possibility, we amplified the portion of the 23S rRNA gene encoding the central loop of domain V. DNA sequencing and restriction fragment length polymorphism of the PCR products showed that each of the four erythromycin-resistant B. pertussis strains tested contained an A-to-G transition mutation at position 2058 (Escherichia coli numbering) of the 23S rRNA gene. The mutation was not found in seven erythromycin-susceptible isolates tested. Two of the resistant isolates were heterozygous, containing at least one mutant copy and one wild-type copy of the 23S rRNA gene. These results indicate that erythromycin resistance in these strains is likely due to a mutation of the erythromycinbinding site in the 23S rRNA gene. Identification of the resistance mechanism will facilitate development of molecular susceptibility testing methods that can be used directly on clinical specimens in the absence of an isolate.

Direct Detection of Erythromycin-Resistant Bordetella pertussis in Clinical Specimens by PCR

“…pertussis found can confer the heterogeneous susceptibility to erythromycin (3). However, we tested the 100 clinical samples using this allele-specific PCR and PCR-based sequencing and did not find any heterozygote variants, suggesting that the heterozygote variants of 23S rRNA in B. pertussis are rare in this study area.…”

“…It is known that B. pertussis carries three copies of the 23S rRNA gene. Heterozygous erythromycin-resistant strains have been reported (3). If the information on heterogeneous susceptibility to erythromycin or coinfection with erythromycin-susceptible B. pertussis is needed, the allele-specific PCR assay 2 should be performed.…”

“…The first erythromycin-resistant Bordetella pertussis was discovered in 1994 (2). Although resistant strains are still rare, they have been reported in several countries, including China (3)(4)(5)(6)(7), where macrolide-resistant B. pertussis has become prevalent (7).…”

“…Based on the molecular mechanism identified, methods have been developed for detection of the point mutation in 23S rRNA of B. pertussis in order to study the susceptibility to macrolides when B. pertussis isolates are available (3,5). In a previous study, we developed a PCR-based sequencing method for identification of the A2047G mutation in cultured B. pertussis isolates and clinical nasopharyngeal (NP) specimens (7).…”

“…Several studies have proven the association between macrolide resistance and the A2047G mutation in the 23S rRNA of B. pertussis (3)(4)(5)(6)(7). Based on the molecular mechanism identified, methods have been developed for detection of the point mutation in 23S rRNA of B. pertussis in order to study the susceptibility to macrolides when B. pertussis isolates are available (3,5).…”