Quantification of antibiotic resistance genes (ARGs) in clouds at a mountain site (puy de Dome, central France)

Antibiotic resistance in bacteria is becoming a major sanitary concern worldwide. The extensive use of large quantities of antibiotics to sustain human activity has led to the rapid acquisition and maintenance of antibiotic resistant genes (ARGs) in bacteria and to their spread into the environment. Eventually, these can be disseminated over long distances by atmospheric transport. Here, we assessed the presence of ARGs in clouds as an indicator of long-distance travel po-tential of antibiotic resistance in the atmosphere. We hypothesized that a variety of ARGs can reach the altitude of clouds mainly located within the free troposphere. Once incorporated in the atmosphere, they are efficiently transported and their respective concentrations should differ depending on the sources and the geographical origin of the air masses. We deployed high-flow rate impingers and collected twelve clouds between September 2019 and Oc-tober 2021 at the meteorological station of the puy de Dome summit (1465 m a.s.l., France). Total airborne bacteria concentration was assessed by flow cytometry, and ARGs subtypes of the main families of antibiotic resistance (quin-olone, sulfonamide, tetracycline; glycopeptide, aminoglycoside, beta-lactamase, macrolide) including one mobile genetic element (transposase) were quantified by qPCR. Our results indicate the presence of 29 different ARGs' subtypes at concentrations ranging from 1.01 x 103 to 1.61 x 104 copies m-3 of air. Clear distinctions could be observed be-tween clouds in air masses transported over marine areas (Atlantic Ocean) and clouds influenced by continental sur-faces. Specifically, quinolones (mostly qepA) resistance genes were prevalent in marine clouds (54 % of the total ARGs on average), whereas higher contributions of sulfonamide, tetracycline; glycopeptide, beta-lactamase and macrolide were found in continental clouds. This study constitutes the first evidence for the presence of microbial ARGs in clouds at concentrations comparable to other natural environments. This highlights the atmosphere as routes for the dissemination of ARGs at large scale.