Genomic dynamics and macrolide resistance in Bordetella pertussis: implications for surveillance and rapid visual...

The post-COVID-19 resurgence of pertussis has raised concerns regarding shifts in circulating Bordetella pertussis populations and increasing macrolide resistance. This study aims to characterize the population dynamics and clinically relevant genomic changes of B. pertussis in China and globally, and to develop a rapid visual assay for detecting the macrolide resistance-associated 23S rRNA A2047G mutation.

In this cross-sectional genomic epidemiology study, 22 recent clinical isolates were sequenced and analyzed with 7322 publicly available B. pertussis genomes. Global population structure was assessed using 7344 genomes, and Bayesian phylodynamic analysis was performed on 1054 temporally informative Chinese genomes selected from 1292 isolates collected during 1951-2024. We designed and optimized an allele-specific loop-mediated isothermal amplification (LAMP) assay incorporating a 3'-terminal artificial mismatch and a neutral red indicator for detection of the macrolide resistance-associated 23S rRNA A2047G mutation. Its analytical sensitivity and specificity were evaluated alongside quantitative PCR (qPCR) using serially diluted plasmids and 39 contrived DNA samples.

The macrolide resistance of B. pertussis in China, defined by the 23S rRNA A2047G mutation, reached 98.9% (88/89; 95% CI 93.9-100) among available genomes in 2024, concurrent with the rapid clonal expansion of the ptxP3-MT28 sub-lineage. Among the available Chinese isolates, the proportion of predicted Prn-deficient strains increased to 31.46% in 2024, with 12 distinct prn inactivation mechanisms, predominantly IS481-mediated insertions. The optimized LAMP assay differentiated mutant and wild-type alleles within 50 min at 65°C, with a neutral red visual limit of detection (LOD) of 10 copies/μL, compared with 100 copies/μL for qPCR. The results of LAMP-based differentiation were consistent with those obtained by qPCR for all 39 contrived samples.

B. pertussis populations have undergone marked genomic shifts characterized by antigenic variation and macrolide resistance, although the evolutionary drivers underlying these temporal and genomic patterns remain to be determined. The visual LAMP assay provides a proof-of-concept analytical evidence for rapid A2047G allele discrimination and may have future point-of-care potential following validation with prospective clinical respiratory specimens. These findings support continued genomic surveillance and further evaluation of vaccine antigen matching and antimicrobial resistance.