Oral Presentation Australian Society for Microbiology Annual Scientific Meeting 2019

Induction of Erythromycin resistance in Bordetella holmesii confirmed by whole genome sequencing (#52)

Winkie Fong 1 2 , Verlaine Timms 1 , Vitali Sintchenko 1 2
  1. Centre of Infectious Diseases and Microbiology - Public Health, Westmead, NSW, Australia
  2. Marie Bashir Institute, University of Sydney, Sydney, NSW, Australia

Bordetella pertussis, the etiological agent of whooping cough, is highly infectious and re-merging globally despite widespread vaccination. The recommended antibiotics for the treatment of B. pertussis infections and post-exposure prophylaxis are Macrolides, primarily Erythromycin and Azithromycin. However, mutations have occurred in the 23S rRNA gene resulting in macrolide resistance. While still uncommon, macrolide-resistant B. pertussis have been reported in Europe and Asia. The mechanism of resistance was identified as an A-to-G transition SNP mutation  at position 2058 of the 23S rRNA gene.

The target sequence IS481 for routine clinical diagnosis has been found in both Bordetella holmesii and B. pertussis genomes. B. holmesii, is most likely the main cause of false-positive results when screening symptomatic patients for pertussis with routine PCR methods. Previous studies have shown that B. holmesii infections range from 0-32% of nasopharygeal isolates from patients with pertussis-like illnesses. We aimed to examine the development of macrolide resistance in Bordetella holmesii

To generate resistance in three B. holmesii strains, comprising of 2 clinical isolates and a type strain, isolates were grown on HBA media with an Erthryomycin E-test or disc for 15 weeks and subcultured every 3-4 days.

B. holmesii isolates took six to twelve weeks (13-25 passages) to develop resistance (>256 µg/mL), from an MIC ranging from 0.047-0.25 µg/mL. Passaged isolates were sequenced on an Illumina NextSeq500, every month (4 weeks/8 passages), to observe any intermediate genomic changes in the isolate that may have contributed to resistance. Average coverage of the isolates ranged from 60-90x, and was able to determine mutations in the 23S rRNA gene were present in B. holmesii. Thus, this study was able to demonstrate a potential resistance mechanisms of B. holmesii.