Microhabitat specificity of Paulisentis missouriensis (Acanthocephala) in the intestine of creek chub (Semotilus atromaculatus) was studied. Data from 64 naturally infected fish harboring only P. missouriensis were utilized to determine if site specificity of worms varied by sex, size, and reproductive maturity, and to test the hypothesis that the first flexure of the intestine of creek chub is the preferred microhabitat, as reported in the literature. Location of worms within the intestine did not differ among size classes of either female or male worms. However, gravid female worms occupied intestinal segments around the first flexure of the intestine, whereas nongravid females were concentrated in the posterior part of the intestine around the second flexure. The distribution of males coincided with that of gravid females. These data suggest that P. missouriensis initially establishes in the posterior regions of the intestine of creek chub, after which they move anteriorly, mate, and come to occupy positions centered around the first flexure of the intestine.