Objectives: To assess and compare the implementation of antimicrobial stewardship (AMS) interventions recommended within the national AMS toolkits, TARGET and Start Smart Then Focus, in English primary and secondary healthcare settings in 2014, to determine the prevalence of cross-sector engagement to drive AMS interventions and to propose next steps to improve implementation of AMS. Methods: Electronic surveys were circulated to all 211 clinical commissioning groups (CCGs; primary sector) and to 146 (out of the 159) acute trusts (secondary sector) in England. Response rates were 39% and 63% for the primary and secondary sectors, respectively. Results: The majority of CCGs and acute trusts reported reviewing national AMS toolkits formally or informally (60% and 87%, respectively). However, only 13% of CCGs and 46% of acute trusts had developed an action plan for the implementation of these toolkits. Only 5% of CCGs had antimicrobial pharmacists in post; however, the role of specialist antimicrobial pharmacists continued to remain embedded within acute trusts, with 83% of responding trusts having an antimicrobial pharmacist at a senior grade. Conclusions: The majority of healthcare organizations review national AMS toolkits; however, implementation of the toolkits, through the development of action plans to deliver AMS interventions, requires improvement. For the first time, we report the extent of cross-sector and multidisciplinary collaboration to deliver AMS interventions in both primary and secondary care sectors in England. Results highlight that further qualitative and quantitative work is required to explore mutual benefits and promote best practice. Antimicrobial pharmacists remain leaders for implementing AMS interventions across both primary and secondary healthcare sectors.