Ultrastructure of Diplophrys parva, a New Small Freshwater Species, and a Revised Analysis of Labyrinthulea (Heterokonta)

We describe Diplophrys parva n. sp., a freshwater heterotroph, using fine structural and sequence evidence. Cells are small (L = 6.5 +/- 0.08 mu m, W = 5.5 +/- 0.06 mu m; mean +/- SE) enclosed by an envelope/theca of overlapping scales, slightly oval to elongated-oval with rounded ends, (1.0 x 0.5-0.7 mu m), one to several intracellular refractive granules (similar to 1.0-2.0 mu m), smaller hyaline peripheral vacuoles, a nucleus with central nucleolus, tubulo-cristate mitochondria, and a prominent Golgi apparatus with multiple stacked saccules (= 10). It is smaller than published sizes of Diplophrys archeri (similar to 10-20 mu m), modestly less than Diplophrys marina (similar to 5-9 mu m), and differs in scale size and morphology from D. marina. No cysts were observed. We transfer D. marina to a new genus Amphifila as it falls within a molecular phylogenetic clade extremely distant from that including D. parva. Based on morphological and molecular phylogenetic evidence, Labyrinthulea are revised to include six new families, including Diplophryidae for Diplophrys and Amphifilidae containing Amphifila. The other new families have distinctive morphology: Oblongichytriidae and Aplanochytriidae are distinct clades on the rDNA tree, but Sorodiplophryidae and Althorniidae lack sequence data. Aplanochytriidae is in Labyrinthulida; the rest are in Thraustochytrida; Labyrinthomyxa is excluded.