Intraspecific variability of Neoparamoeba pemaquidensis (Page, 1970)

Neoparamoeba pemaquidensis (Amoebozoa, Dactylopodida), initially described by F.C. Page in the genus Paramoeba, is mainly free-living and is one of the most sequenced species of the genus Neoparamoeba. However, the boundaries of this species are still unclear due to its poorly studied morphological features. In this study, we present an integrative analysis of molecular, morphological, and morphometric data to examine diversity within N. pemaquidensis and to clarify species boundaries between N. pemaquidensis and N. aestuarina. We studied nine newly isolated strains of N. pemaquidensis. Morphological and ultrastructural characteristics vary both within and between strains, without any regular patterns. Molecular phylogenetic analyses based on three different loci showed that only the COI gene, the ITS1-5.8S-ITS2 region, and a two-gene analysis (18S rDNA and COI gene) reliably separate N. pemaquidensis and N. aestuarina. All three markers reveal three clades of genetically diverse strains within N. pemaquidensis; however, the variability of nuclear markers within clades is comparable to putative intragenomic variability. These clades are stable in composition and congruent across different markers, although higher-level relationships between them are poorly resolved. The Perkinsela amoebae-like (PLO) endosymbionts of N. pemaquidensis also cluster into three main clades, mostly congruent with the corresponding clades of host amoebae.