Despite the importance of the theory of evolution to scientific knowledge, a number of misconceptions continue to be found among teachers and undergraduate students. The aim of the present study was to describe and characterise knowledge about evolution among 120 freshman undergraduate students of two natural sciences programmes (environmental biology and veterinary medicine), 80 pre-service science teachers (elementary and biology) and 45 in-service teachers (elementary and biology high school) in Santiago, Chile. The research was carried out based on an eight-question questionnaire about evolution acceptance and understanding. The instrument included seven Likert-scale questions and one open-ended question. An analysis of the data revealed that more than 70% of teachers (pre-service and in-service) and undergraduate students recognised the theory of evolution as established scientific knowledge. When participants discussed the mechanism of evolution in the open-ended question, the most prevalent responses from students and teachers (33%) explained evolution as need-driven changes for survival purposes. Only 13% of the responses could be considered Darwinian, and 10% of responses included more than one view of evolution. The Darwinian responses generally included three important aspects: variation, inheritance and differential reproduction. The implications for biology teacher education are also discussed in this study.