Objective: To investigate the factors influencing macrolide unresponsiveness in children with Chlamydia pneumoniae (C. pneumoniae) infection, amid its recent sharp increase as an emerging pathogen. Methods: A retrospective analysis of pediatric C. pneumoniae infection cases (2017–2025) was conducted. Logistic regression adjusted for confounding factors to identify independent predictors of macrolide unresponsiveness. Metagenomic next-generation sequencing (mNGS) and whole-genome sequencing (WGS) were used to analyze pathogen copy number, coinfections, resistance genes, virulence factors, and genetic characteristics. Results: Macrolide unresponsiveness primarily occurred in 2025 (95%, 57/60) with an incidence of 42.9% (57/133). Independent factors included age >6 years (OR 0.14, 95% CI 0.01–0.89), age >12 years (OR 0.08, 95% CI 0.01–0.54), summer/autumn (OR 3.65, 95% CI 1.04–14.7), and abnormal aspartate aminotransferase (OR 0.12, 95% CI 0.03–0.47). mNGS showed no association with C. pneumoniae copy number (P=0.9652) but a link with Streptococcus pneumoniae coinfection (P=0.017), with more resistance genes/virulence factors in unresponsive cases. WGS identified predominant ST16/ ST17 (not associated with unresponsiveness, P=0.851), conserved virulence factors, no known resistance genes, but multiple 23S rRNA point mutations. No clear phylogenetic clustering was observed in unresponsive cases. Conclusions: Macrolide unresponsiveness in pediatric C. pneumoniae infection is linked to age, season, abnormal liver function, and S. pneumoniae co-infection, with 23S rRNA mutations potentially contributing. Surveillance and optimized clinical management are urgently needed to address antimicrobial resistance.