Differential growth rates and in vitro drug susceptibility to currently used drugs for multiple isolates of Naegleria fowleri

The free-living amoeba, Naegleria fowleri , which typically dwells within warm, freshwater environments, can opportunistically cause Primary Amoebic Meningoencephalitis (PAM), a disease with a mortality rate of >98%, even with the administration of the best available drug regimens. The lack of positive outcomes for PAM has prompted a push for the discovery and development of more effective therapeutics, but most studies only utilize one or two clinical isolates in their drug discovery assays. The inability to assess possible heterogenic responses to drugs among isolates from varying geographical regions hinders progress in the field due to a lack of proven universal efficacy for novel therapeutics. Herein we conducted drug efficacy and growth rate determinations for 11 different clinical isolates, including one obtained from a successful treatment outcome, by applying a previously developed CellTiter-Glo 2.0 screening technique and flow cytometry. We found some significant differences in the susceptibility of these isolates to 7 of 8 different drugs tested, all of which comprise the cocktail that is recommended to physicians by the Centers for Disease Control. We also discovered significant variances in growth rates among isolates which draws attention to the dissidence among the amoebae populations collected from different patients. The findings of this study reiterate the need for inclusion of additional clinical isolates of varying genotypes in drug assays and highlight the necessity for more targeted therapeutics with universal efficacy across N. fowleri isolates. Our data establishes a needed baseline for drug susceptibility among clinical isolates and provides a segue for future combination therapy studies as well as research related to phenotypic or genetic differences that could shed light on mechanisms of action or predispositions to specific drugs. ### Competing Interest Statement The authors have declared no competing interest.