A.1. Sampling and initial characterization of environments. Several biotopes will be targeted in order to obtain comprehensive information on the AMR phenomenon in the environment: WWTPs, hospital effluent, manure, surface waters, groundwater, soil.
A.2. Optimization of nucleic acids extraction protocols and pre-screening of ARGs using PCR.
A.3. Optimization of antibiotics detection methods in various environmental samples.
A.4. Determination of antibiotic residues in the environmental samples investigated. Determination of antibiotics from studied samples is important to improve their monitorization and preventing the dangerous effects on the environment.
A.5. Antimicrobial susceptibility testing – the phenotypic level. The antimicrobial susceptibility testing will be performed according to the EUCAST guidelines for several antibiotic classes. Isolates showing phenotypic resistance will be deposited in a biobank.
A.6. Molecular identification of antimicrobial resistant strains and ARGs. Quantification of ARGs in the environment using qPCR – the genotypic level. This activity will focus on two objectives regarding the genotypic level of AMR investigation: i) to classify the AMR strains deposited in the biobank using the SSU rRNA gene as marker and to determine the nucleotide sequence of the specific resistance gene; ii) to quantify, using qPCR, the most relevant ARGs as revealed by metagenomics.
A.7. Next Generation Sequencing analysis of ARGs and AMR mechanisms – the metagenomic level. All the environmental samples investigated will be analysed using a metagenomic approach in order to give insight in the complete and possibly new spectrum of resistance genes and mobile genetic elements present, together with microbes that carry them. This approach will allow to evaluate the prevalence of known AMR mechanisms, and might also reveal the existence of new or emerging types of resistance not currently identified.
A.8. Preliminary laboratory testing of wastewater treatment with carbon nanotubes for removal of antibiotics and AMR microorganisms. Nanotechnology and its applications is one of the most rapidly developing sciences with applications in many fields, including the development and improvement of water treatment process. The use of carbon nanoparticles has been shown to be successful in the removal of both antibiotic residues and pathogenic microorganisms.
A.9. Validation of results. Integrate the specific methods and protocols into the methodological guide. The individual protocols and methods developed in each technical activity have to be validated, interpreted and integrated in the final methodological guide. Based on the results obtained during the technical activities implemented, specific recommendations will be proposed for future studies aimed at increasing the monitorization quality of antibiotic residues and AMR phenomenon and at mitigation of antibiotics and antibiotic resistant genes and organisms in the environment with direct consequences on the quality of surface waters and groundwater.
Activity PM1. Project management – management structure and procedures. The main objective is to set up an effective management framework for the EnviroAMR consortium to ensure that the project is progressing towards its planned objectives and the collaboration agreement is respected.
Activity PM2. Project management – dissemination and training. Several dissemination schemes are proposed, together with hands-on trainings for the methods proposed.