Moramonas marocensis gen. nov., sp. nov.: a jakobid flagellate isolated from desert soil with a bacteria-like, but bloated mitochondrial genome

A new jakobid genus has been isolated from Moroccan desert soil. The cyst-forming protist Moramonas marocensis gen. nov., sp. nov. has two anteriorly inserted flagella of which one points to the posterior cell pole accompanying the ventral feeding groove and is equipped with a dorsal vane—a feature typical for the Jakobida. It further shows a flagellar root system consisting of singlet microtubular root, left root (R1), right root (R2) and typical fibres associated with R1 and R2. The affiliation of M. marocensis to the Jakobida was confirmed by molecular phylogenetic analyses of the SSU rRNA gene, five nuclear genes and 66 mitochondrial protein-coding genes. The mitochondrial genome has the high number of genes typical for jakobids, and bacterial features, such as the four-subunit RNA polymerase and Shine–Dalgarno sequences upstream of the coding regions of several genes. The M. marocensis mitochondrial genome encodes a similar number of genes as other jakobids, but is unique in its very large genome size (greater than 264 kbp), which is three to four times higher than that of any other jakobid species investigated yet. This increase seems to be due to a massive expansion in non-coding DNA, creating a bloated genome like those of plant mitochondria.

Pyramimonas robusta sp.nov., a new punctate species, and a reappraisal of the subgenus Punctatae

The ultrastructure of a new punctate species, Pyramimonas robusta, is described and, by comparison with features of Pyramimonas olivacea N. Carter and Pyramimonas mucifera Sym et Pienaar, has allowed for a more defined delimitation to the subgenus Punctatae McFadden. Features, in addition to the obvious character of puncta, which are considered characteristic of this subgenus, include a benthic nature, a chloroplast with a ventral groove coincident with the left and right sinuses, box scales with spines on their side walls, crown scales with eccentric crosses on their base plates, segmented T hairs, limuloid scales with three subsidiary spines and five subsidiary radial ribs, a 3-1 type flagellar apparatus, a basal body with a long region of triplets with longitudinal fibrils separating proximal and distal cartwheel structures, and a 3(2/1):4 or 3:3(2/1):2 microtubular root system. Although as a suite these are characteristic, a great deal of the individual features are shared with other subgenera, specifically subgroup 2 members of the subgenus Trichocystis McFadden but especially members of the subgenus Pyramimonas McFadden. Features originally considered diagnostic that now are questionable include pyrenoid structure and starch investiture and the bilayered nature of the eyespot. Key words: Punctatae, Pyramimonas, Pyramimonas robusta sp. nov., ultrastructure.

Actin has multiple roles in the formation and architecture of zoospores of the oomycetes, Saprolegnia ferax and Achlya bisexualis

Very similar changing patterns of actin are described with rhodamine-phalloidin labelling during the zoosporic life cycle of the oomycetes, Saprolegnia ferax and Achlya bisexualis. By comparing the changes with previously described ultrastructural and functional changes, we show that actin functions in numerous previously unrecognized processes. Most spectacularly, the directed vesicle expansions of the cytokinetic system involve newly formed actin which outlines the developing zoospores. Disruption of this actin with cytochalasins leads to abnormal cleavage as witnessed by the formation of enlarged and irregular cysts. Prior to cytokinesis, two new types of organelle are synthesized and one, known as K bodies, clusters around the nuclei. These organdies are actin-rich during development and clustering, consistent with actin functioning in their positioning. In the zoospores, actin is concentrated around the water expulsion vacuoles, indicating that they are contractile, and permeates the cytoplasm, probably with a skeletal role. This concept is supported by the first demonstration of actin specifically associated with a microtubular root in the secondary zoospore. Upon encystment there is a dramatic increase in stained actin in the form of peripheral plaques associated with the newly synthesized cell wall. When the cysts germinate, a fibrillar actin cap, comparable to that previously described in hyphal tips, forms in the germ tube apex, but only after cell wall softening to permit germ tube protrusion. This sequence is consistent with the actin cap modulating turgor-driven expansion of the tip as previously discussed for hyphae. In addition to disrupting cleavage-associated actin, cytochalasins show developmental stage, dose and drug (CE≥CD≥CB) specific effects on zoosporulation-related actin, which indicates that, contrary to previous suggestions, rhodamine-phalloidin staining is a useful indicator of actin behaviour in response to cytochalasins. These responses include differential effects on adjoining actin arrays, some of which are transient in the continued presence of the drugs, indicating a mechanism of drug adaptation.

Ultrastructure of the flagellar roots in Chlamydomonas gametes

The cytoskeleton of Chlamydomonas reinhardtii gametes has been studied by electron microscopy. The microtubular system, consisting of four flagellar roots inserted into the basal apparatus, is shown to include two daughter basal bodies and two striated fibres, newly described in this report. One new fibre associates with the 3-over-1 root and is similar to its counterpart, the striated fibre of the 2-member root. These similar root fibres connect each daughter basal body to the V-shaped microtubular root pair. The other new striated fibre joins the daughter basal body to both flagellar roots and is similar to the proximal striated fibre. In mt+ gametes, the conventional root microtubules make direct contact with the doublet zone of the non-activated mating structure. During activation, doublet zone microfilaments associate with the daughter basal body and the finely striated fibre of the 3-over-1 root. These observations suggest that the cytoskeleton acts as a scaffolding for membrane extension by the mt+ mating structure microfilaments.

Flagellar roots, mating structure and gametic fusion in the green alga Ulva lactuca (Ulvales)

The slightly anisogamous gametes of Ulva lactuca exhibit a cruciate flagellar root system consisting of 4 microtubular roots (4-2-4-2 system) and an elaborate system of fibrous roots associated with the 2-stranded microtubular roots. Two fibres (32-nm striation periodicity; system I fibres) closely underlie each of the 2-stranded roots, while different fibres (150-nm striation periodicity; system II fibres) run parallel to the root microtubules, and are 150–200 nm more internally located. Female gametes have 4 system II fibres, 3 of which are combined into a compound fibre associated with one microtubular root, while the fourth fibre is associated with the opposite root. In male gametes only 2 system II fibres are present, each underlying one of the two 2-stranded roots. A special region of the plasmalemma of both gamete types about 0.5 mum away from the basal bodies and located between 2 adjacent microtubular roots is structurally specialized and acts as a mating structure in gametic fusion. The region is oval-shaped and up to I.I mum long with a maximum diameter of 0.7 mum. A continuous electron-dense boundary layer underlies the plasmalemma at the edges of the mating structure. In both gamete types the mating structure consists of a fuzzy layer of material underlying the plasmalemma and special granules (60 nm diameter) are associated with this layer on its cytoplasmic side. In addition diffuse material overlies the mating structure, especially in male gametes. The mating structure is connected to 3 different kinds of flagellar roots: the boundary layer is linked to a 2-stranded microtubular root and its associated system I fibre; the fuzzy layer of the mating structure is connected with a system II fibre; and in female gametes this is the compound system II fibre. The ultrastructural changes which occur after mixing the 2 gamete types have been followed. Mating structure activation involves contraction of system II fibres (change of striation periodicity to 100 nm), detachment of special granules from the fuzzy layer of the mating structure and their replacement by electron-transparent vesicles at the prospective cell fusion site. Furthermore, release of electron-dense contents from Golgi-derived vesicles in the anterior part of both gamete types precedes cell fusion. Cell fusion is exclusively initiated in a region delimited by the 2 mating structures. After partial dissolution the 2 plasma membranes unite within the mating structure regions. The ultrastructure of gametic fusion in Ulva lactuca is compared to that of other green algae and the significance of flagellar roots in the mating process of green algae is discussed.

Ultrastructural Features of the Oral Apparatus of Stentor

The ultrastructure of the normal oral apparatus of Stentor has not been extensively studied. I report here on the ultrastructure of the buccal cavity of Stentor coeruleus.Stentor coeruleus was fixed in either a buffered mixture of osmium tetroxide and glutaraldehyde, or in buffered glutaraldehyde alone. Cells were then dehydrated and embedded in a mixture of Epon and Araldite.An extensive adoral zone of membranelles surrounds the anterior of the cell, and each membranelle consists of 2 parallel rows of cilia. These extend down into the buccal cavity. Two microtubular root fibers, or nemadesmata (Figs. 2 and 5), extend deeply into the cytoplasm from the base of each ciliary kinetosome. Mitochondria are usually closely associated with the root fiber bundles, and small vesicles are present between the nemadesmata of adjacent kinetosomes (Fig. 5). In the cytopharyngeal, non-ciliated areas of the buccal cavity, microtubular ribbons which extend into the cytoplasm are aligned perpendicular to the plasma membrane of the buccal cavity (Figs. 1 and 2).