The history of dual therapy with ceftriaxone and azithromycin illustrates how quickly the situation can change. The combination was introduced as a strategic response to concerns about ceftriaxone monotherapy failure, and it carried the added benefit of activity against Chlamydia trachomatis and Mycoplasma genitalium, which frequently co-occur with gonorrhea. In its early years the strategy worked in many settings, because most ceftriaxone-resistant gonococcal strains were still susceptible to azithromycin, and the dual approach helped slow the international spread of ceftriaxone resistance. Over the past five years, however, azithromycin resistance has risen disturbingly across the globe, undermining the drug’s value within dual therapy and threatening the efficacy of current treatment protocols. Contributing factors include extensive and often inappropriate antibiotic use, suboptimal diagnostic practices, limited surveillance systems, and delayed updates to clinical guidelines.

The future therapeutic landscape appears increasingly precarious. Two novel antimicrobials, zoliflodacin and gepotidacin, have recently been approved for uncomplicated gonorrhea, yet few additional agents are in clinical development. At the same time, resistance to ceftriaxone itself continues to climb in many regions, and strains with concurrent resistance to both ceftriaxone and azithromycin have now been reported in several WHO regions. High treatment costs further restrict access in resource-limited settings, where surveillance and control measures are often weakest, compounding the risk that resistance emerges and spreads undetected. Without new therapeutic strategies, enhanced global surveillance and robust research investment, gonorrhea could become progressively harder to treat, with rising morbidity, complications and treatment failures.

Azithromycin is an oral macrolide long used against chlamydial and mycoplasmal infections, and it once demonstrated good efficacy against gonococci as well. Its widespread availability, particularly where antibiotics can be obtained without prescription or regulation, has contributed significantly to the development of resistance. Over the past decade, azithromycin resistance has steadily increased in both high- and low-income countries, with marked geographic variability shaped by local treatment practices, surveillance infrastructure and antibiotic consumption patterns. Although azithromycin is no longer routinely recommended for gonorrhea in many settings, rising resistance limits its remaining utility and underscores why continued antimicrobial resistance surveillance matters.

The numbers tell a sobering story. A retrospective genomic surveillance study by Daniel Golparian and colleagues, published in The Lancet Microbe, examined N. gonorrhoeae isolates collected across 21 European Economic Area countries in 2020 through the European Gonococcal Antimicrobial Surveillance Programme. It found that azithromycin resistance, defined by the EUCAST epidemiological cut-off of MIC greater than 1 mg/L, had risen to 9.4 percent of isolates, up from 8.0 percent in 2018. In the United States, resistance climbed from 0.01 percent in 1992 to 0.4 percent by 2012, then surged fourfold from 0.6 percent in 2013 to 2.5 percent in 2014, reaching 4.6 percent in 2018 and 5.9 percent by 2019 among isolates with elevated MICs. Globally, the WHO Gonococcal Antimicrobial Surveillance Programme reported that 46 of 53 countries, or 87 percent, detected azithromycin-resistant isolates in 2022, and 36 countries reported resistance in at least 5 percent of isolates.

Regional heterogeneity is striking. In the WHO Western Pacific region, seven of eight reporting countries detected resistant isolates in 2022, with Brunei, Cambodia, China, Japan and Singapore each reporting resistance in at least 5 percent of isolates, while Australia and New Zealand recorded lower levels. China’s Gonococcal Resistance Surveillance Program documented azithromycin resistance reaching 16.9 percent in 2022, a substantial increase over the preceding five years, whereas all isolates reported from the Philippines remained susceptible. Surveillance itself faces structural challenges: while high-income countries maintain robust systems, reporting is inconsistent in low- and middle-income countries due to constraints in laboratory capacity, funding and infrastructure. Only 77 countries and territories reported gonococcal resistance data to WHO GASP during 2019 to 2022, and the absence of data from parts of Africa and Southeast Asia means the true global extent of resistance, and of dual-therapy treatment failures, may be significantly underreported.

Understanding the molecular machinery behind resistance clarifies why surveillance is difficult. Azithromycin inhibits bacterial protein synthesis by binding the peptidyltransferase region of domain V of the 23S rRNA in the 50S ribosomal subunit, blocking peptide exit. Resistance arises through several mechanisms: mutations in the 23S rRNA itself, particularly A2059G and C2611T, alter the drug-binding site; the A2059G transition in all four rRNA alleles produces extraordinarily high MICs of 256 to 4096 mg/L, while C2611T yields moderate resistance of 2 to 32 mg/L. Acquired erm genes encoding rRNA methylases can modify the target, though they have become rare in recent isolates. Mutations in rplD and rplV, encoding ribosomal proteins L4 and L22, moderately raise MICs, and efflux pump overexpression adds another layer of resistance.

Efflux is now central to the story. The MtrCDE pump, the best-characterized system, expels macrolides when its regulator MtrR is disabled by mutations such as G45D, or when promoter mutations, including a single-nucleotide deletion in a 13-base-pair inverted repeat, increase pump transcription. Mosaic mtr alleles, acquired through recombination with sequences from Neisseria meningitidis or Neisseria lactamica, substantially raise azithromycin MICs by altering both the pump and its regulation. The MacAB and mef-encoded efflux systems may also contribute, though they remain less studied in gonococci. Collectively, these mechanisms reveal a complex and geographically variable genetic architecture of resistance.

That variability is precisely what undermines molecular diagnostics. Historically, 23S rRNA mutations were the predominant, best-characterized determinants, and systematic reviews confirm that assays targeting A2059G accurately detect high-level resistance while C2611T assays identify moderate resistance. But the European genomic surveillance revealed a decisive shift: only 10.5 percent of azithromycin-resistant isolates in 2020 carried 23S rRNA mutations, down from 24.1 percent in 2018, while roughly 89 percent instead carried mtrD/mtrR promoter mosaic 2 or semi-mosaic mtrD variants. The authors attribute the resistance surge to the expansion of specific NG-STAR clonal complexes, notably CC63, CC168 and CC213 harboring the mosaic promoter, plus the emergence of novel CC1031 carrying a semi-mosaic mtrD variant. An assay built for the old targets would miss the dominant circulating resistance genotypes.

The implications for public health are clear. The WHO advocates molecular assays that detect resistance markers to guide therapy and support stewardship, but their validity, accuracy and specificity falter when marker distributions differ across regions; in areas dominated by mtrD mosaic variants, a 23S rRNA-focused assay would yield a very low positive predictive value. Tailoring molecular targets to local genetic epidemiology is therefore essential. In low- and middle-income countries where routine whole-genome sequencing is impractical, a pragmatic sentinel-site strategy, with representative laboratories performing susceptibility testing and periodically referring isolates to regional reference centers for genomic characterization, offers a feasible alternative. A standardized minimum dataset reported to WHO GASP, covering susceptibility results, specimen source, geography, collection year and resistance determinants, would enable continuous refinement of regionally calibrated assays. The shift from dual therapy toward ceftriaxone monotherapy may also reshape selection pressure on azithromycin resistance, though the magnitude and direction of that effect remain uncertain, since macrolide use for other infections, fitness effects and lineage dynamics all play a role. Sustained phenotypic and genomic surveillance, novel antimicrobials and combination therapies, and collaboration among researchers, clinicians and public health stakeholders will determine whether gonorrhea remains treatable in the decade ahead.

Subject of Research: Heterogeneity of azithromycin resistance markers in Neisseria gonorrhoeae and the challenges of implementing molecular assays for routine antimicrobial resistance surveillance

Article Title: Heterogeneity of azithromycin resistance markers in Neisseria gonorrhoeae and challenges in implementing molecular assays for routine surveillance

Article References: Heterogeneity of azithromycin resistance markers in Neisseria gonorrhoeae and challenges in implementing molecular assays for routine surveillance. (n.d.). https://doi.org/10.1016/j.nmni.2026.101838

Image Credits: AI Generated

DOI: 10.1016/j.nmni.2026.101838

Keywords: Neisseria gonorrhoeae, azithromycin resistance, antimicrobial resistance, molecular surveillance, 23S rRNA mutations, mtr mosaic variants, efflux pumps, WHO GASP, ceftriaxone, gonorrhea treatment, genomic epidemiology, molecular diagnostics