Antimicrobial resistance (AMR) is a threat to achieving the United Nation’s (UN) sustainable development goals (SDGs). The behavior of stakeholders has directly influenced the extent of AMR and understanding underpinning knowledge and attitudes is an important step towards understanding these behaviors. The aim of this study was to develop and validate a novel questionnaire, utilizing the theory of Appreciative Inquiry, to measure knowledge and attitudes around antibiotic resistance amongst community pharmacists throughout Thailand. A survey tool was developed using the Appreciative Inquiry theory, and was piloted in a non-probability sample of practicing community pharmacists. Descriptive and inferential statistics were applied and the tool validated, using a three-step psychometric validation process. A total of 373 community pharmacists participated in the study. The survey tool was found to be valid and reliable. The “Knowledge” domain of the survey tool showed an acceptable level of reliability (Cronbach’s alpha 0.64); while the “Attitude” domain showed an excellent reliability level (Cronbach’s alpha 0.84). This new survey tool has been designed to measure attitudes and knowledge of antibiotic resistance by utilizing the Discovery phase of Appreciative Inquiry theory amongst community pharmacists in Thailand. This survey tool has the potential to be used by other researchers across different settings.