The present study aimed to evaluate the antimicrobial activity and the possible mechanisms of activity of polypeptide‑enriched Gastrodia elata extracts (GEP) against the gram‑negative bacteria Escherichia coli and Pseudomonas aeruginosa, the gram‑positive bacterium Staphylococcus aureus and the fungus Candida albicans. The antimicrobial activity of GEP was first confirmed by determining the minimum inhibitory concentration by growth curve analysis. GEP was found to damage the cell wall and membrane of the microorganisms tested, as revealed by the morphological changes visible through scanning electron microscopy, and by the observed leakage of alkaline phosphatase and β‑galactosidase from cells. GEP was demonstrated to perturb the metabolism of the microorganisms, especially the tricarboxylic acid cycle, as indicated by the reduced intracellular activity of succinate dehydrogenase, malate dehydrogenase and ATPases, including the Na+/K+‑ATPase and the Ca2+‑ATPase. In addition, GEP caused the leakage of the genetic material of the bacteria and the fungus, as indicated by the increased OD260. The results of the present study indicated that GEP may exert its antimicrobial activity by damaging cell walls and membranes, causing the leakage of genetic material, and by perturbing cellular metabolism.