Paravahlkampfia francinae n. sp Masquerading as an Agent of Primary Amoebic Meningoencephalitis

Paravahlkampfia francinae n. sp., a new species of the free-living amoeba genus Paravahlkampfia, designated as CDC:V595, was isolated from the cerebrospinal fluid of a patient with headache, sore throat, and vomiting, typical symptoms of primary amoebic meningoencephalitis (PAM) caused by Naegleria fowleri. The isolate grew at 33 degrees C, 37 degrees C, 40 degrees C, and 42 degrees C and destroyed mammalian cell cultures. However, it did not kill young mice upon intranasal inoculation. P. francinae does not produce flagellates and does not grow on agar plates coated with Gram- negative bacteria such as Escherichia coli, the usual food source of Paravahlkampfia ustiana, the type species of the genus. The trophozoite at light microscopy exhibited eruptive locomotion and possessed a single vesicular nucleus. Ultrastructurally, the trophozoites had numerous mitochondria with discoidal cristae but did not have a Golgi apparatus. The trophozoites differentiated into cysts after consuming most of the monolayer. The cyst had an inner well-differentiated endocyst and an outer thin, wrinkled, and wavy ectocyst with no pores. During excystation trophozoites ruptured the cyst wall and emerged from the cysts. A unique feature seen in the cysts was the presence of bacterial endosymbionts, both in the endoplasm and within the cyst wall. Full-length sequencing analysis of the 18S and 5.8S RNA genes of P. francinae showed that they were distinct from those of other Paravahlkampfia species. The patient recovered within a few days indicating that some of the previously reported cases of PAM that survived may have been due to P. francinae.