STUDY OF MICROBIAL BIODIVERSITY OF WATERS AND SOILS CONTAMINATED BY ORGANIC AND INORGANIC POLLUTANTS, PRESENCE AND DIFFUSION OF ANTIBIOTIC-RESISTANT BACTERIA (ARB) AND ANTIBIOTIC-RESISTANCE GENES (ARGS)

ANTIBIOTIC RESISTANCE (AR) IS INCREASING AND RECOGNIZED AS A CRITICAL ENVIRONMENTAL AND PUBLIC HEALTH CHALLENGE, PARTICULARLY IN URBANIZED SYSTEMS WHERE MULTIPLE ANTHROPOGENIC PRESSURES CONVERGE. WITHIN A ONE HEALTH FRAMEWORK, THIS DOCTORAL THESIS INVESTIGATES THE OCCURRENCE, DISTRIBUTION, AND MITIGATION OF ANTIBIOTIC-RESISTANT BACTERIA AND GENES (ARB/ARGS) ALONG THE URBAN WATER-SOIL-PLANT CONTINUUM. AN URBAN SOIL FROM THE METROPOLITAN AREA OF MILAN WAS CHARACTERIZED COMBINING CULTURE-DEPENDENT ANALYSES WITH SHOTGUN METAGENOMIC SEQUENCING TO CHARACTERIZE MICROBIAL COMMUNITIES AND RESISTANCE PATTERNS IN BULK SOIL AND PLANT RHIZOSPHERE. RESULTS INDICATE THAT URBAN SOILS HOST DIVERSE MICROBIAL ASSEMBLAGES WITH HETEROGENEOUS RESISTANCE PROFILES, SHAPED BY LONG-TERM ANTHROPOGENIC STRESSORS. CLEAR COMPARTMENT-SPECIFIC DIFFERENCES EMERGED, WITH THE RHIZOSPHERE ACTING AS A DYNAMIC HOTSPOT FOR ARGS ENRICHMENT AND DIVERSIFICATION, LIKELY DRIVEN BY INTENSIFIED MICROBIAL COMPETITIONS, PLANT-INTERACTIONS AND HORIZONTAL GENE TRANSFER. THEREFORE, GIVEN THAT AR IS NOT CURRENTLY SUBJECT TO SPECIFIC REGULATORY FRAMEWORKS, TOGETHER WITH INCREASING EVIDENCE OF ITS WIDESPREAD PRESENCE IN THE ENVIRONMENT, CONSTRUCTED WETLANDS (CWS) WERE EVALUATED AS NATURE-BASED SOLUTIONS FOR CIVIL WASTEWATER RECLAMATION DUE TO THEIR RELEVANCE AS SOURCES OF AR. TWO PILOT-SCALE CONFIGURATIONS (VERTICAL-VCW AND HYBRID-HCW) WERE ASSESSED FOR THEIR ABILITY TO REMOVE CONVENTIONAL CONTAMINANTS, ARB AND ARGS IN COMPLIANCE WITH ITALIAN REGULATIONS FOR WATER REUSE (D.M. 185/03). BOTH SYSTEMS ACHIEVED HIGH TREATMENT EFFICIENCIES FOR ALL THE CHEMICO-PHYSICAL AND BIOLOGICAL PARAMETERS MEETING ITALIAN WATER REUSE STANDARDS, WITH THE HYBRID CONFIGURATION SHOWING ENHANCED PERFORMANCE, PARTICULARLY FOR NITROGEN REMOVAL AND MICROBIAL ABATEMENT. FURTHERMORE, ARB AND ARGS WERE EFFECTIVELY REDUCED IN BOTH SYSTEMS, EMPHASIZING THE POTENTIAL OF THE CWS IN MITIGATING AR. FINALLY, THE REUSE OF CW-TREATED WASTEWATER FOR LETTUCE SEEDLINGS IRRIGATION WAS INVESTIGATED TO ASSESS THE POTENTIAL TRANSFER OF AR DETERMINANTS TO AGRICULTURAL SOILS AND EDIBLE PLANT TISSUES. WHILE LIMITED ACCUMULATION OF ARB WAS DETECTED IN SOIL AND RHIZOSPHERE, NO TRANSLOCATION OF ARB OR ARGS TO PLANT LEAVES WAS OBSERVED, INDICATING A LOW RISK TO FOOD SAFETY UNDER THE EXPERIMENTAL CONDITIONS. OVERALL, THIS DOCTORAL WORK DEMONSTRATES THAT URBAN SOILS PLAY AN ACTIVE ROLE AS RESISTOME RESERVOIRS IN THE ENVIRONMENT AND THAT CWS CAN EFFECTIVELY REDUCE AR DISSEMINATION IN WATER REUSE SCENARIOS. THE FINDINGS SUPPORT THE IMPLEMENTATION OF INTEGRATED ONE HEALTH STRATEGIES AND HIGHLIGHT THE POTENTIAL OF NATURE-BASED SOLUTIONS TO ENSURE SAFE AND SUSTAINABLE WASTEWATER REUSE IN URBAN AND PERI-URBAN AGRICULTURE.