This article presents the development and validation of a new instrument measuring social, cultural, and behavioral factors influencing exposure to occupational zoonoses in agricultural settings. The Social Ecological Model (SEM) and the Standards for Educational and Psychological Testing were used as guiding frameworks to ensure the instrument’s validity. The instrument’s content was compiled by combining the results of a scoping literature review and an expert qualitative study. The instrument items were drafted, organized, and underwent a meticulous process of revision and adjustment. It was translated into Spanish and tested in one-to-one cognitive interviews with five volunteer agricultural workers. Exploratory factor analysis was conducted for construct discrimination, and bivariate regression analyses were conducted to explore the association with exposure indicators. Evidence of validity was obtained from four out of five sources of validity evidence according to the AERA/APA’s Standards of Educational and Psychological Testing. The Social Ecology of Occupational Zoonoses (SEOZ) was successfully used to determine social and behavioral factors associated with a higher risk of exposure to occupational zoonoses. Further use of the SEOZ can provide valuable insights into developing effective interventions to improve the health and well-being of agricultural workers.