The complete mitochondrial genome of Paravannella minima (Amoebozoa, Discosea, Vannellida).

We present a complete sequence and describe the organization of the mitochondrial genome of the amoeba Paravannella minima (Amoebooza, Discosea, Vannellida). This tiny species represents a branch at the base of Vannellida tree, to the moment being its earliest-branching lineage. The circular mitochondrial DNA of this species has 53,464 bp in length and contains 30 protein-coding genes, 2 ribosomal RNAs, 23 transfer RNAs, and 15 open reading frames. This genome is significantly longer and contains more protein-coding genes than any yet sequenced mitochondrial genome of vannellid amoebae. Unlike the previously sequenced mitochondrial genomes of Vannellida, which should be translated using the "Table 4" (the mold, protozoan, and coelenterate mitochondrial code), that of P. minima can be properly translated using the universal genetic code.

Mitochondrial Genome of Vannella croatica (Amoebozoa, Discosea, Vannellida).

Mitochondrial genome sequence of Vannella croatica (Amoebozoa, Discosea, Vannellida) was obtained using pulse-field gel electrophoretic isolation of the circular mitochondrial DNA, followed by the next-generation sequencing. The mitochondrial DNA of this species has the length of 28,933 bp and contains 12 protein-coding genes, two ribosomal RNAs, and 16 transfer RNAs. Vannella croatica mitochondrial genome is relatively short compared to other known amoebozoan mitochondrial genomes but is rather gene-rich and contains significant number of open reading frames.

The complete mitochondrial genome of Vannella simplex (Amoebozoa, Discosea, Vannellida).

Vannella simplex (Amoebozoa, Discosea, Vannellida) is one of the commonest freshwater free-living lobose amoebae, known from many locations worldwide. In the present study, we describe the complete mitochondrial genome of this species. The circular mitochondrial DNA of V. simplex has 34,145öbp in length and contains 27 protein-coding genes, 2 ribosomal RNAs, 16 transfer RNAs and 4 open reading frames. Mitochondiral genome of V. simplex is one of the most gene compact due to overlapping genes and reduced intergenic space. It has much in common with its closest relative, mitochondrial genome of V. croatica GenBank number MF508648. In the same time, both of them show considerable differences in length and in gene order from the next close relative - that of Neoparamoeba pemaquidensis KX611830 (deposited as Paramoeba) and even more - from other sequenced amoebozoan mitochondrial genomes. The present study confirms the opinion that the level of synteny between the mitochondrial genomes across the entire Amoebozoa clade is low. More or less considerable similarity yet was found only between members of the same clade of the genera or family level, but hardly - among more distant lineages.

Phylogeny and Systematics of Leptomyxid Amoebae (Amoebozoa, Tubulinea, Leptomyxida).

We describe four new species of Flabellula, Leptomyxa and Rhizamoeba and publish new SSU rRNA gene and actin gene sequences of leptomyxids. Using these data we provide the most comprehensive SSU phylogeny of leptomyxids to date. Based on the analyses of morphological data and results of the SSU rRNA gene phylogeny we suggest changes in the systematics of the order Leptomyxida (Amoebozoa: Lobosa: Tubulinea). We propose to merge the genera Flabellula and Paraflabellula (the genus Flabellula remains valid by priority rule). The genus Rhizamoeba is evidently polyphyletic in all phylogenetic trees; we suggest retaining the generic name Rhizamoeba for the group unifying R. saxonica, R.matisi n. sp. and R. polyura, the latter remains the type species of the genus Rhizamoeba. Based on molecular and morphological evidence we move all remaining Rhizamoeba species to the genus Leptomyxa. New family Rhizamoebidae is established here in order to avoid paraphyly of the family Leptomyxidae. With the suggested changes both molecular and morphological systems of the order Leptomyxida are now fully congruent to each other.

Description of Flamella daurica n. sp., with notes on the phylogeny of the genus Flamella and related taxa.

A new species of naked amoebae, Flamella daurica (Amoebozoa, Conosa, Variosea), from the Russian Far East is studied and described. In addition, we isolated a strain, identified as Flamella aegyptia from Lake Ontario in Canada. Its SSU rRNA gene sequence is almost identical with that of the type strain, except for the presence of an intron in the SSU sequence. The phylogenetic analysis of a variosean alignment after inclusion of a number of environmental sequences found in GenBank confirmed that the species Talaepolella tubasferens Lahr 2012 is closely related to the Flamella clade but probably represents an independent genus and includes no less than two different species. This conclusion is supported by considerable differences between Talaepolella and Flamella in morphology and life history. Talaepolella tubasferens is an evolutionary important species demonstrating the transition from compact flamellian to expanded acramoebian morphology. The genus Flamella now includes twelve named species, which makes it one of the most species-rich genera of naked lobose amoebae.

Morphology and phylogeny of Vannella croatica n. sp. (Amoebozoa, Discosea, Vannellida).

We isolated and described a new species of freshwater vannellid amoeba from Krka natural reserve in Croatia--Vannella croatica n. sp. This species has certain morphological differences from all known vannellids and differs at the level of SSU sequence. It resembles in size and morphology Vannella lata; to facilitate direct comparison we publish images of V. lata CCAP 1589/12 strain (type strain, which is now lost) taken in 1999. Vannela croatica feeds on bacteria and can be easily grown in large amount in relatively pure culture and thus is suitable for molecular and biochemical studies requiring large amounts of material.

Two new species of the genus Stenamoeba (Discosea, Longamoebia): cytoplasmic MTOC is present in one more amoebae lineage.

Two new species of the recently described genus Stenamoeba, named S. berchidia and S. sardiniensis were isolated from a single soil sample on Sardinia, Italy. Both share morphological features characteristic to Stenamoeba and form in phylogenetic analyses together with other Stenamoeba spp. a highly supported clade within the family Thecamoebidae. The ultrastructural investigation of Stenamoeba sardiniensis revealed the presence of cytoplasmic microtubule-organizing centers (MTOCs), located close to one of several dictyosomes found inside the cell. This is the first report of cytoplasmic MTOCs among Thecamoebidae. The presence of MTOCs is now shown in five of nine orders comprising the class Discosea and potentially could be a phylogenetic marker in this group. We re-isolated Stenamoeba limacina from German soils. This strain shows a similar morphology and an almost complete SSU rDNA sequence identity with the type strain of S. limacina originating from gills of fishes, collected in Czech Republic.