Introduction: The rise of antimicrobial resistance (AMR) is one of the greatest global threats to human health. Intensive monitoring to identify and track critical AMR “hotspots” across the environment, humans, animals, and the pathways connecting them in a One Health setting is recommended. Improper use of antimicrobials contaminates soils with antimicrobials and forces an evolutionary shift in bacteria towards developing or selecting for antimicrobial resistance. Currently, there is no surveillance of AMR in global agricultural soils, leaving a major knowledge gap. This project aims to address that gap through a nationwide survey of AMR genes in 3,000 Irish agricultural soil samples collected as part of the Irish Department of Agriculture, Food and the Marine ‘Farm Environmental Study Programme’. Methods: A national soil biobank of 3,008 soil samples was created and DNA was extracted to create a corresponding soil DNA biobank. The DNA extracts were screened using quantitative PCR (qPCR) for 15 clinically relevant AMR genes. Results: Preliminary findings indicate widespread distribution of sul1 (91.47%), intI1 (95.75%), ermB (79.85%) and aadA1 (81.05%). Clinically significant β-lactamase genes were identified at lower frequencies, including blaCTX-M (4.17%) and blaTEM (1.9%). The vancomycin resistance gene vanA was detected in 11.37% of samples. blaKPC (0.27%) and blaVIM (0.1%) were rarely detected. Conclusion: Overall, clinically significant β-lactamase genes were detected at low prevalence, but their presence confirms that agricultural soils can harbour resistance factors associated with human and animal health. This study provides the largest nationwide molecular baseline of clinically relevant ARGs in agricultural soils globally.