Multigene identification of new subtypes in the Acanthamoeba T4 genotype and description of Acanthamoeba pussardi sp. nov.

Acanthamoeba is a widespread free-living amoeba that can cause rare but serious opportunistic infections in humans and animals. Various species have been described based on morphological criteria that have proven unreliable, and strain identification is currently based on genotype assignment using nuclear 18S rDNA sequences. Most Acanthamoeba strains belong to genotype T4, which is the most abundant in the environment as well as the most frequently reported in clinical samples. Due to its richness, the T4 genotype has been further subdivided into subtypes, which correspond almost perfectly to the subtypes of the mitochondrial small subunit (mtSSU) rDNA. The 18S/mtSSU subtype clusters provide a clearer picture of species groups, and other subtypes remain to be identified. In this study, we provide evidence, through 18S/mtSSU co-clustering, for the existence of additional subtypes of the Acanthamoeba T4 genotype, two nuclear and four mitochondrial, and describe a new species, Acanthamoeba pussardi sp. nov. The data are supported by the analysis of two mitochondrial protein-coding genes, cytochrome c oxidase subunit 1 (cox1) and nicotinamide adenine dinucleotide (NADH) dehydrogenase subunit 5 (nad5), as well as the mitochondrial rDNA spacer.