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Diagnostic studies on the relationship between Drosophila willistoni and Nosema kingi revealed that adult D. willistoni can acquire the microsporidian via the oral route from fecal contamination and transovarially. At least 50% of the flies exposed to fecal material from infected flies contained the parasite when examined two or three weeks later. 50–81% of the offspring which emerged from eggs laid by parasitized females were infected with N. kingi 14–21 days later. Infectivity experiments using other drosophilids indicated N. kingi infections can be transmitted to D. sturtevanti Duda, D. melanogaster Meigen, D. persimilis Dobzhansky and Epling, D. pseudoobscura Frolowa, D. virilis Sturtevant, D. simulans Sturtevant, and D. ananassae Doleshall without morphological changes occurring in the stages of the parasite and without changes of behavioural patterns in these experimental hosts.