Characterisation of Bordetella pertussis virulence and macrolide resistance in Australia by targeted culture-independent sequencing: a genomic epidemiology study
Authors:
- Fong, Winkie
- Rockett, Rebecca J
- Tam, Kingsley King-Gee
- Nguyen, Trang
- Sim, Eby M
- Tay, Enoch
- Suster, Carl J E
- Agius, Jessica E
- Chandra, Shona
- Watt, Anne E
- Speers, David
- Graham, Maryza
- Tran, Thomas
- Lim, Chuan Kok
- Wehrhahn, Michael C
- Ginn, Andrew N
- Gray, Darcy
- Robson, Jennifer
- Gardner, Indya
- McDougall, Rodney
- Papanicolas, Lito
- Howard-Jones, Annaleise R
- Outhred, Alexander C
- Kennedy, Karina
- Cooley, Louise
- Wang, Qinning
- Jeoffreys, Neisha
- Chen, Sharon C-A
- Basile, Kerri
- Golubchik, Tanya
- Kok, Jen
- Sintchenko, Vitali
Details:
The Lancet Microbe, Volume 7, Issue 3, 2026-03-31
Article Link: Click here
Background Bordetella pertussis continues to circulate globally despite widespread vaccination, with a notable epidemic in 2024. Its resurgence is confounded by the emergence of pertactin-deficient, macrolide-resistant B pertussis strains in Asia and Europe, which are under-recognised by conventional diagnostics. We aimed to apply targeted culture-independent next-generation sequencing (tNGS) of respiratory specimens to improve global B pertussis diagnostic capability and genomic surveillance. Methods We did a nationwide genomic epidemiology study of B pertussis RT-PCR-positive respiratory specimens that were retrospectively and prospectively collected by diagnostic and public health laboratories in six of seven states and territories of Australia. Specimens underwent tNGS and macrolide-resistant B pertussis-specific PCR, and an opportunistic subset from New South Wales and Queensland were cultured for confirmatory susceptibility testing and whole-genome sequencing. Sequencing data were analysed for genome recovery, virulence profiles, and macrolide resistance mutations, and were compared with international macrolide-resistant B pertussis genomes and ancestral Australian genomes. The performance of the tNGS approach was assessed with logistic regression relative to RT-PCR cycle threshold values, and sensitivity and specificity values were calculated. Findings 255 respiratory specimens positive for B pertussis were included in the study. 64 (25%) were retrospectively collected between Jan 12, 2012, and Dec 31, 2023, and 191 (75%) were prospectively collected between Jan 1 and Oct 28, 2024. Of these 255 specimens, 148 (58%) yielded near-complete B pertussis genomes through tNGS. Seven co-circulating lineages of B pertussis were documented, including two associated with macrolide-resistance. Eight epidemiologically unrelated and geographically dispersed cases of macrolide-resistant B pertussis with a 23S rRNA 2037A→G mutation were identified by tNGS and confirmed by whole-genome sequencing. Three of these were further validated by phenotypic testing. The estimated prevalence of macrolide resistance among Australian cases positive for B pertussis was 4% (eight of 188). Interpretation tNGS can recover near-complete B pertussis genomes directly from clinical specimens, enabling identification of macrolide resistance mutations and high-resolution phylogenetic analysis. These findings show that tNGS complements PCR-based surveillance by providing genome-wide assessment of resistance, virulence, and genomic diversity in a single workflow. Funding NSW Health Prevention Research Support Program.