Characterization of the amebae isolated from the first confirmed case of primary amebic meningoencephalitis in Oklahoma

Scope and method of study. Naegleria fowleri is an opportunistic pathogen and the cause of a rapidly fatal brain disease known as primary amebic meningoencephalitis (PAM). In August, 1998 a three year old girl died after she became infected while swimming at a local lake. Prior to her death, the cause was unknown. After autopsy a diagnosis of PAM was made. Amebae were isolated and cultivated from a small amount of brain tissue provided by the Medical Examiners Office. The isolate was designated HBT1-1998 and tentatively identified as N. fowleri. Standard laboratory tests were performed to positively identify the species of ameba. These tests included concanavalin A (Con A) agglutination, flagellate formation, growth studies at multiple temperatures in different growth media, ameba and cyst measurement, drug studies with azithromycin and amphotericin B, and indirect immunofluorescence (IIF). In vivo studies included histological examination of brain tissue sections obtained from infected mice. Percent mortality as well as mean time to death was determined using a murine model.; Findings and conclusions. The amebae of HBT1-1998were limax in appearance, elongate and slug-like, and moved with directional motility by explosive blunt pseudopodia. The amebae did not agglutinate in the presence of Con A. They were determined to be ameboflagellates. These preliminary tests were compatible with typical characteristics for N. fowleri. Growth conditions were also typical of N. fowleri, and growth curves were similar. Both HBT1-1998 and the control N. fowleri culture (Lee strain) had virtually identical results when cultured with azithromycin and amphotericin B. IIF positively confirmed the amebae as N. fowleri . The in vivo studies also matched positively with N. fowleri . The LD50 was 1.32 x 102 amebae per mouse, an extremely low dose, and percent mortality reached 100% with a dose of 1 x 105 amebae/mouse. Mean time to death averaged approximately 6 days post infection. After examining all of the data, HBT1-1998 was confirmed to be Naegleria fowleri.