Assessing the function of the alternative electron transport chain in the <i>Cryptosporidium parvum</i> mitosome

Cryptosporidiosis is a leading cause of diarrhea in young children and immunocompromised individuals, particularly AIDS/HIV patients. The only FDA-approved drug against cryptosporidiosis, nitazoxanide, has limited effectiveness in immunocompromised patients and is not approved for use in children under 1 year. Genomic analysis and previous studies proposed an alternative respiration pathway involving alternative oxidase (AOX) and type II NAD(P)H dehydrogenase (NDH2), which are thought to generate the mitosome membrane potential in <i>Cryptosporidium parvum</i>. Additionally, AOX was nominated as potential drug targets, based on its absence in mammalian hosts and sensitivity of parasite growth to known inhibitors of AOX. However, our study demonstrated that NDH2 is not localized in the mitosome, AOX is non-essential for parasite growth, and knockout lines lacking this enzyme are equally sensitive to AOX inhibitors. These findings indicate that AOX and NDH2 are not ideal candidates for future drug development against cryptosporidiosis and force a re-evaluation of models of how the mitosome generates its membrane potential.