During parasitological surveys of freshwater fish from the Miranda River, Brazil, 199 Pygocentrus nattereri kner (Characidae) were caught. Two pentastomid families, Subtriquetridae Fain, 1961, represented by its single genus Subtriquetra Sambon, 1922, and Sebekidae Sambon, 1922, represented by three genera, were present. Free-living larvae of Subtriquetra subtriquetra (Diesing, 1835) were collected from the swim bladder. Encysted larvae of Alofia Giglioli, 1922 were found in the abdominal cavity, chambers of the heart, musculature, on the surface of the gonads and swim bladder. Some Alofia larvae were moving freely in the swim bladder. Larvae of Sebekia Sambon, 1922 were encysted in the musculature. Some larvae of Leiperia Sambon, 1922 were found encysted in the musculature and on the surface of the pyloric caeca, whereas others occurred free in the abdominal cavity. In some of the latter, the head was buried deep in the wall of the intestine, stomach or ovaries, whereas the rest of their body remained free. Infective pentastomid larvae were present throughout the year with an overall prevalence of 77%. Both prevalence and intensity were higher in members of the Sebekidae than in Su. subtriquetra, possibly due to the latter's mode of transmission and its high pathogenicity. No sex-related, statistically significant differences (p > 0.05) in prevalence or abundance were found. Fish weight and length had significant but weak positive correlations (r ≤ 0.27) with the abundance of pentastomid larvae, possibly reflecting an increased likelihood of prior exposure in older fish. Parasite abundance had no significant effect on host body condition (p ≥ 0.69). A higher prevalence and monthly mean abundance of pentastomids were seen in the dry season and might be due to increased host densities as habitats dry up. Pygocentrus nattereri represents a new intermediate host record for the genera Alofia, Leiperia and Subtriquetra.