Trypophobia refers to the extreme negative reaction when viewing clusters of circular objects. Two major evolutionary frameworks have been proposed to account for trypophobic visual discomfort. The skin disease-avoidance (SD) framework proposes that trypophobia is an over-generalised response to stimuli resembling pathogen-related skin diseases. The dangerous animal (DA) framework posits that some dangerous organisms and trypophobic stimuli share similar visual characteristics. Here, we performed the first experimental manipulations which directly compare these two frameworks by superimposing trypophobic imagery onto multiple image (...) categories to evaluate changes in comfort. Participants from two countries (United States and Croatia) were evaluated on several measures, including general trypophobia levels, perceived vulnerability to disease, and generalised anxiety. Several analyses showed stronger changes in comfort in the human skin condition (hand, feet, and chest images) compared to the dangerous animal condition (snake and spider images). Furthermore, participants with higher levels of trypophobia showed significantly stronger changes in comfort in the skin condition than the dangerous animal condition, with comparable effects obtained across nationalities. Several variables entered as covariates failed to significantly account for this effect. The present work is the first to experimentally test both evolutionary frameworks of trypophobia, with results supporting the skin disease-avoidance framework. (shrink)