The amoeba-to-flagellate transformation test is not reliable for the diagnosis of the genus Naegleria. Description of three new Naegleria spp.

Trophozoites of several isolates from one location in Australia have failed consistently to transform into flagellates, although they display all othe r characteristics of the genus Naegleria. When changing the standard transf ormation test, flagellates were produced. In phylogenetic trees derived fro m partial small subunit ribosomal DNA (SSUrDNA) sequences, one of these str ains branches close to a cluster comprising N. clarki, N. australiensis, N. italica and N. jadini. It is proposed that these Australian isolates repre sent a new species, named N. fultoni (strain NG885). Failing to form flagel lates since their isolation, even when different transformation procedures are used, are two Naegleria strains from Chile and Indonesia. In SSUrDNA-ba sed phylogenetic trees the Chilean strain clusters with N. pussardi and the Indonesian strain clusters with N. galeacystis, but the degree of sequence difference from these described species (3.5% and 2.2%, respectively) is s ufficient to propose that both of the strains represent new species, named N. chilensis (strain NG946) and N. indonesiensis (strain NG945), respective ly. The close relationships between each of the new species and the Naegler ia species with which they cluster in SSUrDNA-based trees were confirmed by ribosomal internal transcribed spacer region (ITS) sequence comparisons. I n France, several non-flagellating N. fowleri strains were isolated from on e location. ITS rDNA sequence comparisons indicated that they correspond to a 'type' of N. fowleri found in both Europe and the USA. A redefinition of the genus Naegleria is proposed as a consequence of these and previous fin dings.