Science and environmental educators implicitly presuppose that the ecosystem per se is clearly defined and consider learners' misunderstandings to be incorrect scientific interpretations of uncontested knowledge content. The present study, however, explores how in-service secondary education Greek biology teachers address the topic of 'ecosystem delimitation' under the assumption that the ecosystem is an ill-defined concept. If it is to be considered a scientific (holistic) entity, the ecosystem finds its definitional and accompanying features within the ecological field of systems ecology, which declined after the 1970s under pressures of persisting unsolved puzzles and problems. Our research focuses on the background assumptions fuelling the difficulties that system ecologists encounter in sufficiently determining ecosystem's borders and anchoring 'ecosystem' holism in the empirical world. The results indicate that biology teachers address the topic of ecosystem delimitation from a perspective that apparently differs from the realistic/empirical and instrumental perspectives of system ecologists. Moreover, biology teachers cannot engage with how system ecologists delimit ecosystems and instead resort to their personal experience (e.g. individualist insights) and naïve relational ideas, which along with deficiencies in systems thinking and in Nature of science (NOS) reasoning result in the development of an 'ecosystem' version that appears inconsistent with (ecosystem) holism.