Members of the genus Helicosporidium are the first described algal insect pathogens. They have a close affinity to the non-photosynthetic algae of the genus Prototheca, and have a wide host range, infecting many species of aquatic and terrestrial insects. In this study the infectivity of two Helicosporidium spp. isolates, originating from a black fly (SjHe) and an aquatic weevil (CsHe), was tested against a weevil Diaprepes abbreviatus (L.) and three mosquito species Anopheles quadrimaculatus Say, Culex quinquefasciatus Say, and Aedes aegypti (L.). The weevil constitutes a new experimental host record for helicosporidia. The CsHe isolate was more virulent than the SjHe isolate in D. abbreviatus. Anopheles quadrimaculatus was the most susceptible mosquito species measured by infection rate and mortality. The infectivity and virulence of SjHe and CsHe isolates did not differ in any of the mosquito species.