Clinical parameters of African penguins Spheniscus demersus have been recorded mostly from birds admitted to rehabilitation centres and are typically presented as mean values across a region. It is uncertain whether these values are representative of wild penguins and whether they are influenced by parasite infestations. We assessed general clinical parameters from 793 African penguins (210 adults and 583 chicks) at five South African colonies, in 2016 and 2017, and then established the relationship between those values and parasite infestations. Penguins at Dassen Island had indications of a different health status: lower body condition and total plasma protein, and higher haematocrit and ectoparasite richness. Overall, haematocrit values were lower for the mainland colonies than for the island colonies. All clinical parameters were significantly lower in spring than in autumn/winter. Both ectoparasite and haemoparasite taxon richness were negatively related to haematocrit values, while helminth taxon richness was positively related to body mass and haematocrit. At the Stony Point colony, tick abundance and ectoparasite and haemoparasite taxon richness were associated with lower haematocrit in adults and chicks. This study highlights intercolony variations in the health status of African penguins and potential factors (such as parasite infestations and/or food-resource accessibility) that might affect specific colonies, thereby helping to focus conservation efforts.