Longitudinal Antimicrobial Susceptibility Shifts in Environmental Legionella pneumophila from a Drinking Water Distribution System

Legionella pneumophila (L. pneumophila) persists in engineered water systems and is the causative agent of Legionnaires’ disease. Despite its clinical importance, little is known about whether environmental populations are undergoing gradual changes in their antimicrobial susceptibility. Here, we compared the antimicrobial susceptibility of 95 environmental L. pneumophila isolates collected in 2013–2014 with those obtained in 2024–2025 from the same sites in a drinking water distribution system (DWDS) using Epsilometer (E-test). The analyses showed both temporal and spatial variation: Ciprofloxacin activity remained relatively stable, whereas levofloxacin exhibited a broader minimum inhibitory concentration (MIC) range among contemporary isolates, with the minimum inhibitory concentration required to inhibit the growth of 90% of isolates (MIC90) doubling from 0.5 to 1.0 mg/L, suggesting increased variability and the emergence of less susceptible subpopulations. Tigecycline showed a marked site-specific increase in MIC values, with the mean MIC at site D increasing from 0.26 mg/L in the 2013–2014 isolates to 1.86 mg/L in the 2024–2025 isolates, indicating reduced susceptibility at this site. Among macrolides, erythromycin exhibited the lowest MIC values across both isolate collections, whereas azithromycin showed higher MIC values. Moreover, isolates recovered from adjacent sites within the same DWDS exhibited significantly different susceptibility profiles, possibly reflecting the influence of local environmental conditions. These findings emphasize that L. pneumophila susceptibility is not static but is likely shaped by ecological and temporal factors. The study underscores the importance of long-term environmental surveillance to track the evolution of this pathogen and provides new insights into how micro-scale dynamics within water systems can influence antimicrobial susceptibility patterns.