The Pinguiophyceae classis nova, a new class of photosynthetic stramenopiles whose members produce large amounts of omega-3 fatty acids

The Pinguiophyceae class. nov., a new class of photo‐synthetic stramenopiles (chromophytes), is described. The class includes five monotypic genera, Glossomastix, Phaeomonas, Pinguiochrysis (type genus), Pinguio‐coccus and Polypodochrysis. These algae have an unusually high percentage of polyunsaturated fatty acids, especially 20:5 (n‐3)(EPA, eicosapentaenoic acid). These fatty acids are the basis for choosing the Latin noun ‘Pingue’ (= fat, grease) as the root for the class name. Analyses of nuclear‐encoded 18S rRNA and chloroplast‐encoded rbcL gene sequence data showed that these algae formed a monophyletic group that could not be placed in any other class. Morphologically, the species are all single‐celled microalgae from picoplanktonic size to over 40 urn in length. Each cell has one (or two) typical chloroplast(s) with a girdle lamella and a surrounding chloroplast endoplasmic reticulum. Pyrenoids occur within the chloroplast, varying from embedded to stalked, and membranes penetrate into the pyrenoid in all five genera. Phaeomonas has motile cells with two flagella, and the forward‐directed flagellum bears mastigonemes (tripartite flagellar hairs). Two other genera (Glossomastix, Polypodochrysis) produce zoospores that possess only one smooth flagellum (no mastigonemes), and this flagellum apparently is the mature flagellum, a feature previously unknown in the photosynthetic stramenopiles. The major carotenoid pigments in the pinguiophytes are fucoxanthin, violaxanthin, zeaxanthin and P‐carotene, as well as chlorophyll a and chlorophyll c‐related pigment(s). These features support recognition of the Pinguiophyceae class. nov. as a unique group of algae.     

Studies on Woloszynskioid Dinoflagellates X: Ultrastructure,Phylogeny and Colour Variation in Tovellia rubescens n. sp. (Dinophyceae)

The external morphology and internal cell fine structure of a new species of Tovelliaceae, Tovellia rubescens n. sp., is described. Phylogenetic analyses based on partial LSU rDNA sequences place the new species in a clade containing Tovellia species that accumulate red pigments and identify T. aveirensis as its closest known relative. Cells of T. rubescens n. sp. were mostly round and had the cingulum located near the middle, with its ends displaced about one cingular width. Small numbers of distinctly flat cells appeared in culture batches; their significance could not be determined. Cells of the new species in culture batches progressively changed from a yellowish‐green, mainly due to chloroplast colour, to a reddish‐brown colour that appeared associated with lipid bodies. The switch to a reddish colour happened earlier in batches grown in medium lacking sources of N or P. Pigment analyses by HPLC‐MS/MS revealed the presence of astaxanthin and astaxanthin‐related metabolites in the new species, but also in T. aveirensis, in which a reddish colour was never observed. The chloroplast arrangement of T. rubescens n. sp. resembled that of T. aveirensis, with lobes radiating from a central pyrenoid complex. The flagellar apparatus and pusular system fell within the general features described from other Tovelliaceae. A row of microtubules interpretable as a microtubular strand of the peduncle was present. Spiny resting cysts with red contents and an ITS sequence identical to that of cultured material of the new species were found in the original locality.     

TAXONOMIC STUDY OF TWO NEW GENERA OF FUSIFORM GREEN FLAGELLATES,TABRIS GEN. NOV. AND HAMAKKO GEN. NOV. (VOLVOCALES,CHLOROPHYCEAE)1

On the basis of LM, we isolated strains of two species of fusiform green flagellates that could be assigned to former Chlorogonium (Cg.) Ehrenb. One species, “Cg.”heimii Bourr., lacked a pyrenoid in its vegetative cells and required organic compounds for growth. The other was similar to Cg. elongatum (P. A. Dang.) Francé and “Cg.”acus Nayal, but with slightly smaller vegetative cells. Their molecular phylogeny was also studied based on combined 18S rRNA, RUBISCO LSU (rbcL), and P700 chl a‐apoprotein A2 (psaB) gene sequences. Both species were separated from Chlorogonium emend., Gungnir Nakada and Rusalka Nakada, which were formerly assigned to Chlorogonium. They were accordingly assigned to new genera, Tabris Nakada gen. nov. and Hamakko (Hk.) Nakada gen. nov. as T. heimii (Bourr.) Nakada comb. nov. and Hk. caudatus Nakada sp. nov., respectively. Tabris is differentiated from other genera of fusiform green flagellates by its vegetative cells, which only have two apical contractile vacuoles and lack a pyrenoid in the chloroplast. Hamakko, on the other hand, is distinguishable by the fact that its pyrenoids in vegetative cells are penetrated by flattened thylakoid lamellae.     

MORPHOLOGY,MOLECULAR PHYLOGENY AND TAXONOMY OF TWO NEW SPECIES OF PLEODORINA (VOLVOCEAE,CHLOROPHYCEAE)1

The volvocacean genus Pleodorina has been morphologically characterized as having small somatic cells in spheroidal colonies and anisogamous sexual reproduction with sperm packets. In this study we examined two new species that can be assigned to the genus Pleodorina based on morphology: P. starrii H. Nozaki et al. sp. nov. and P. thompsonii F. D. Ott et al. sp. nov. P. starrii was collected from Japan and had 32‐ or 64‐celled colonies with anterior somatic cells and spheroidal individual cellular sheaths that were weakly attached to each other within the colonial envelope. P. thompsonii from Texas (USA) exhibited four or 12 somatic cells in the anterior pole of 16‐ or 32‐celled colonies, respectively, and had a single large pyrenoid in the chloroplast of mature reproductive cells. The chloroplast multigene phylogeny placed P. starrii and P. indica (Iyenger) H. Nozaki in a clade that was robustly separated from the type species P. californica Shaw and P. japonica H. Nozaki. Pleodorina thompsonii was resolved as a basal branch within a large monophyletic group (Eudorina group) composed of Eudorina, Pleodorina and Volvox (excluding section Volvox). Thus, Pleodorina was found among three separate lineages within the Eudorina group in which Eudorina and Volvox were also resolved as nonmonophyletic. The DNA sequences from additional species/strains as well as recognition of morphological attributes that characterize the monophyletic groups within the Eudorina group are needed to construct a natural generic classification within these members of the Volvocaceae.     

LAURENCIA MARILZAE SP. NOV. (CERAMIALES,RHODOPHYTA) FROM THE CANARY ISLANDS,SPAIN, BASED ON MORPHOLOGICAL AND MOLECULAR EVIDENCE1

Laurencia marilzae Gil‐Rodríguez, Sentíes et M.T. Fujii sp. nov. is described based on specimens that have been collected from the Canary Islands. This new species is characterized by distinctive yellow–orange as its natural habitat color, a terete thallus, four pericentral cells per vegetative axial segment, presence of secondary pit‐connections between adjacent cortical cells, markedly projecting cortical cells, and also by the presence of corps en cerise (one per cell) present in all cells of the thallus (cortical, medullary, including pericentral and axial cells, and trichoblasts). It also has a procarp‐bearing segment with five pericentral cells and tetrasporangia that are produced from the third and fourth pericentral cells, which are arranged in a parallel manner in relation to fertile branchlets. The phylogenetic position of this taxon was inferred based on chloroplast‐encoded rbcL gene sequence analyses. Within the Laurencia assemblage, L. marilzae formed a distinctive lineage sister to all other Laurencia species analyzed. Previously, a large number of unique diterpenes dactylomelane derivatives were isolated and identified from this taxon. L. marilzae is morphologically, genetically, and chemically distinct from all other related species of the Laurencia complex described.     

PHYLOGENETIC POSITION OF CRUSTOMASTIX STIGMATICA SP. NOV. AND DOLICHOMASTIX TENUILEPIS IN RELATION TO THE MAMIELLALES (PRASINOPHYCEAE,CHLOROPHYTA)1

A new marine microalga from the Mediterranean Sea, Crustomastix stigmatica Zingone, is investigated by means of LM, SEM, TEM, and pigment and molecular analyses (nuclear‐encoded small subunit [SSU] rDNA and plastid‐encoded rbcL). Pigment and molecular information is also provided for the related species Dolichomastix tenuilepis Throndsen et Zingone. Crustomastix stigmatica has a bean‐shaped cell body 3–5 μm long and 1.5–2.8 μm wide, with two flagella four to five times the body length. The single chloroplast is pale yellow‐green, cup‐shaped, and lacks a pyrenoid. A small bright yellow stigma is located in the mid‐dorsal part of the cell under the chloroplast membrane. An additional accumulation of osmiophilic globules is at times seen in a chloroplast lobe. Cells lack flat scales, whereas three different types of hair‐like scales are present on the flagella. The main pigments of C. stigmatica are those typical of Mamiellales, though siphonein/siphonaxanthin replaces prasinoxanthin and uriolide is absent. The pigment pool of D. tenuilepis is more similar to that of Micromonas pusilla (Butcher) Manton et Parke and of other Mamiellales. The nuclear SSU rDNA phylogeny shows that the inclusion of C. stigmatica and D. tenuilepis in the Mamiellales retains monophyly for the order. The two species form a distinct clade, which is sister to a clade including all the other Mamiellales. Results of rbcL analyses failed to provide phylogenetic information at both the order and species level. No unique morphological or pigment characteristics circumscribe the mamiellalean clade as a whole nor its two daughter clades.     

Observations on Purpureofilum apyrenoidigerum gen. et sp. nov. from Australia and Bangiopsis subsimplex from India (Stylonematales, Bangiophyceae, Rhodophyta)

Purpureofilum apyrenoidigerum gen et sp. nov. was obtained from a mangrove habitat in New South Wales, Australia. It had unbranched uniseriate to multiseriate filaments less than 1 mm tall, with a unicellular base. Each cell had a single multilobed parietal chloroplast without a pyrenoid. During reproduction vegetative cells were discharged directly as monospores that remained motile for several hours after release. Spores with long tails moved more slowly (0.053–0.195 μm sχ) than spores without tails (0.43–1.76 μm s′¹). Phylo‐genetic analysis of sequences of the small subunit of the nuclear‐encoded rRNA and plastid‐encoded ribu‐lose bisphosphate carboxylase/oxygenase genes revealed that Purpureofilum is a member of the Stylonematales and is most closely related to the filamentous genus Bangiopsis. Bangiopsis differs from Purpureofilum by having longer (to 5 mm) multiseriate filaments, cells containing a stellate chloroplast, a conspicuous central pyrenoid, and monospores often formed in packets. Monospores of Bangiopsis were also motile. Transmission electron microscopy investigation of Purpureofilum and Bangiopsis revealed that the Golgi complexes are associated only with rough endo‐plasmic reticulum and that the plastid contains a peripheral thylakoid; this combination of features being the same as in all other multicellular members of the Stylonematales. The low molecular weight carbohydrates of Purpureofilum and Bangiopsis were digenea‐side and sorbitol, which were present in most other Stylonematales.     

pEffects of UV radiation on the ultrastructure of several red algae

The effect of ultraviolet (UV) radiation on the ultrastructure of four red algae, the endemic Antarctic Palmaria decipiens (Reinsch) Ricker and Phycodrys austrogeorgica Skottsberg, the Arctic‐cold temperate Palmaria palmata (Linnaeus) O. Kuntze and the cosmopolitan Bangia atropurpurea (Roth) C. Agardh was studied. All four species showed a formation of ‘inside‐out’ vesicles from the chloroplast thylakoids upon exposure to artificial UV‐radiation. In P. decipiens, most vesicles were developed after 8 h and in P. palmata, after 48 h of UV exposure. In B. atropurpurea, vesi‐culation of thylakoids was observed after 72 h of UV irradiation. In Ph. austrogeorgica, the chloroplast envelope and thylakoid membranes were damaged and the phycobilisomes became detached from the thylakoids after 12 h of UV exposure. Ultraviolet‐induced changes in the membrane structure of mitochondria were observed in P. decipiens and P. palmata. However, in P. decipiens they were reversible as was the damage in chloroplast fine structure after 12 h of UV treatment. Protein crystals in Ph. austrogeorgica showed degradation after exposure to UV radiation. Different methods of fixation and embedding macroalgal material are discussed. These findings give insight into the fine structural changes which occur during and after UV exposure and indicate a relationship between the species dependent sensitivity to UV‐exposure and the depth distribution of the different species.     

A multilocus sequencing approach reveals the cryptic phylogeographical history of Phyllodoce nipponica Makino (Ericaceae)

Discordant phylogeographical patterns among species with similar distributions may not only denote specific biogeographical histories of different species, but also could represent stochastic variance of genealogies in applied genetic markers. A multilocus investigation representing different genomes can be used to address the latter concern, allowing robust inference to biogeographical history. In the present study, we conducted a multilocus phylogeographical analysis to re‐examine the genetic structuring of Phyllodoce nipponica, in which chloroplast (cp)DNA markers exhibited a discordant pattern compared to those of other alpine plants. The geographical structure of sequence variation at five nuclear loci was not consistent with that of cpDNA and showed differentiation between the northern and southern parts of the range of this species. Its demographic history inferred from the isolation‐with‐migration model suggests that the north–south divergence originated from Pleistocene vicariance. In addition, the demographic parameters showed a lack of chloroplast‐specific gene flow, suggesting that stochastic variance in genealogy resulted in the discordant geographical structure. Thus, P. nipponica probably experienced Pleistocene vicariance between its southern and northern range parts in concordance with other alpine plants in the Japanese archipelago. The findings of the present study demonstrates the importance of using a multilocus approach for inferring population dynamics, as well as for reconciling discordant phylogeographical patterns among species. © 2013 The Linnean Society of London, Biological Journal of the Linnean Society, 2013, 110 , 214–226.     

LIGHT MICROSCOPY AND ELECTRON MICROSCOPY OF NEPHROSELMIS SPINOSA SP. NOV. (PRASINOPHYCEAE,CHLOROPHYTA)1

Nephroselmis spinosa Suda sp. nov. is described based on LM and EM observations. Two strains of N. spinosa (S222 and SD959‐3) were isolated from sand samples collected from the northwest coast of western Australia. The cells were remarkably right–left flattened and appeared ellipse or bean‐shaped when viewed from their right or left side. A single, parietal, crescent chloroplast was pale green to yellowish green and contained one conspicuous eyespot in its anterior ventral edge near the base of the short flagellum. A pyrenoid with three starch plates was located at the dorsal of the chloroplast. The cells divided by transverse binary cell division, as is common in other species of this genus. This alga possessed four types of body scales, and three scale types were similar to N. olivacea Stein and N. astigmatica Inouye & Pienaar. However, the fourth and outermost scale type was distinctive because although it was a spiny stellate scale with nine spines, one of them extended about 1 μm and was slightly curved with a hook at the end. This scale morphology, an important taxonomic characteristic, has never been described for the genus Nephroselmis. The cell's morphology, pyrenoid structure, hair scales, and cell size were distinctive from previously described Nephroselmis species, and its unique scale characteristic led me to name this newly proposed species “spinosa,” meaning spiny.     

Comparative study of chloroplast morphology and ontogeny in <Emphasis Type="Italic">Asterochloris</Emphasis> (Trebouxiophyceae,Chlorophyta)

Confocal laser scanning microscopy was utilized to compare the chloroplast morphology and ontogeny among five strains of the green alga Asterochloris. Parsimony analysis inferred from the rDNA ITS sequences confirmed their placement in three distinct lineages: Asterochloris phycobiontica, Trebouxia pyriformis and Asterochloris sp. Examination by confocal microscopy revealed the existence of interspecific differences in the chloroplast ontogeny of Asterochloris; this was based upon either specific chloroplast structures observed in a single species, or on the differential timing of particular ontogenetic sequences. The occurrence of flat parietal chloroplasts prior to cell division, considered as a basic morphological discriminative character of Asterochloris, was clearly associated with the process of aplanosporogenesis. By contrast, chloroplast transformation prior to the formation of autospores proceeded simply by the multiple fission of the chloroplast matrix in the cell lumen. Presented at the International Symposium Biology and Taxonomy of Green Algae V, Smolenice, June 26–29, 2007, Slovakia.     

Brandtia ciliaticola gen. et sp. nov. (Chlorellaceae,Trebouxiophyceae) a common symbiotic green coccoid of various ciliate species

Many freshwater protists harbor unicellular green algae within their cells and these host‐symbiont relationships slowly are becoming better understood. Recently, we reported that several ciliate species shared a single species of symbiotic algae. Nonetheless, the algae from different host ciliates were each distinguishable by their different genotypes, and these host‐algal genotype combinations remained unchanged throughout a 15‐month period of sampling from natural populations. The same algal species had been reported as the shared symbiont of several ciliates from a remote lake. Consequently, this alga appears to play a key role in ciliate‐algae symbioses. In the present study, we successfully isolated the algae from ciliate cells and established unialgal cultures. This species is herein named Brandtia ciliaticola gen. et sp. nov. and has typical ‘Chlorella‐like’ morphology, being a spherical autosporic coccoid with a single chloroplast containing a pyrenoid. The alga belongs to the Chlorella‐clade in Chlorellaceae (Trebouxiophyceae), but it is not strongly connected to any of the other genera in this group. In addition to this phylogenetic distinctiveness, a unique compensatory base change in the SSU rRNA gene is decisive in distinguishing this genus. Sequences of SSU‐ITS (internal transcribed spacer) rDNA for each isolate were compared to those obtained previously from the same host ciliate. Consistent algal genotypes were recovered from each host, which strongly suggests that B. ciliaticola has established a persistent symbiosis in each ciliate species.     

Cryptic Speciation in the Genus Cryptoglena (Euglenaceae) Revealed by Nuclear and Plastid SSU and LSU rRNA Gene

The photosynthetic euglenoid genus Cryptoglena is differentiated from other euglenoid genera by having a longitudinal sulcus, one chloroplast, two large trough‐shaped paramylon plates positioned between the chloroplast and pellicle, and lack of metaboly. The genus contains only two species. To understand genetic diversity and taxonomy of Cryptoglena species, we analyzed molecular and morphological data from 25 strains. A combined data set of nuclear SSU and LSU and plastid SSU and LSU rRNA genes was analyzed using Bayesian, maximum likelihood, maximum parsimony, and distance (neighbor joining) methods. Although morphological data of all strains showed no significant species‐specific pattern, molecular data segregated the taxa into five clades, two of which represented previously known species: C. skujae and C. pigra, and three of which were designated as the new species, C. soropigra, C. similis, and C. longisulca. Each species had unique molecular signatures that could be found in the plastid SSU rRNA Helix P23_1 and LSU rRNA H2 domain. The genetic similarity of intraspecies based on nr SSU rDNA ranged from 97.8% to 100% and interspecies ranged from 95.3% to 98.9%. Therefore, we propose three new species based on specific molecular signatures and gene divergence of the nr SSU rDNA sequences.     

ENDOBACTERIA IN THE DIATOM PINNULARIA (BACILLARIOPHYCEAE). I. “SCATTERED ct‐NUCLEOIDS” EXPLAINED: DAPI–DNA COMPLEXES STEM FROM EXOPLASTIDIAL BACTERIA BORING INTO THE CHLOROPLASTS1,2

A local strain of the pennate diatom Pinnularia cf. “nobilis” was investigated using cytochemistry and fluorescence and EM techniques. The regular perforation of the chloroplasts of P. “nobilis” and the lack of a typical diatom pyrenoid were confirmed at the ultrastructural level. Cavities and channels in the complex secondary plastid were found to harbor symbiotic bacteria, and their DNA elicited DAPI fluorescence. Wheat germ agglutinin, labeling bacteria walls, elicited a similar fluorescence pattern. Previous speculation that the apochlorotic DNA‐positive dots in the plastids of several Pinnularia species are “scattered ct‐nucleoids” is thus refuted. Bacteria were rod shaped and gram negative. They resided in the lumen of the endoplasmic reticulum (ER) during host interphase confined to the specialized ER compartment housing the secondary plastid, that is, to the space between the third and the fourth membrane profile, encircling the chloroplast. TEM images of chemically and cryofixed cells revealed that cavities resulted from the interaction of bacteria with the plastid according to the following sequence, alignment, attachment, deformation, and disintegration. This occurred without visible injury to the primary chloroplast envelope or the relict cell membrane of the reduced ancestral red alga that surrounds the chloroplast. The patterned arrangement of bacteria suggests recognition sites on the vestigial cell membrane, thought to interact with surface groups on the bacteria. The intimate association between bacteria and secondary plastid inside the common specialized ER cisterna suggests they form a functional unit. Comparison of thylakoid profiles, disrupted by bacteria in Pinnularia, with those disrupted by the pyrenoid in other pennate diatoms (e.g. Trachyneis) revealed a significant ultrastructural resemblance. No aposymbiotic Pinnularia cells were found at the sampling site.     

Inter- and intraspecific polymorphism at chloroplast SSR loci and the inheritance of plastids in Pinus radiata D. Don

DNA sequence analysis of chloroplast genomes has revealed many short nucleotide repeats analogous to nuclear microsatellites, or simple sequence repeats (SSRs). We designed PCR primers flanking five of these regions identified in the chloroplast sequence from Pinus thunbergii and tested them for amplification in Pinus radiata, P. elliotii, P. taeda, P. strobus, Pseudotsuga menziesii, Cupressus macrocarpa, four New Zealand native conifer species (Podocarpus totara, Podocarpus hallii, Podocarpus nivalis, Agathis australis), and four angiosperms (Vitex lucens, Nestegis cunninghamii, Actinidia chinensis, and Arabidopsis thaliana). A PCR product in the expected size range was amplified from all species and interspecific polymorphism was detected at all five loci. Intraspecific polymorphism was detected in P. radiata with four of the five primer pairs. One of these polymorphic chloroplast SSR (cpSSR) was then used to determine the inheritance of chloroplasts in 206 progeny from four control-pollinated, full-sibling P. radiata families. Approximately 99% of the progeny had the cpSSR variant of the pollen parent indicating that in Pinus radiata, like most other conifers, chloroplasts are typically inherited from the paternal parent. These results suggest that polymorphic chloroplast SSRs will be a valuable tool for studying chloroplast diversity, cyto-nuclear disequilibrium, and plastid inheritance in a range of species, and for the analysis of gene flow via pollen and paternity in species with paternal transmission of chloroplasts.     

New unicellular red alga,Bulboplastis apyrenoidosa gen. et sp. nov. (Rhodellophyceae,Rhodophyta) from the mangroves of Japan: Phylogenetic and ultrastructural observations

A novel unicellular red alga collected from a mangrove area on Iriomote Island in southwest Japan is described as Bulboplastis apyrenoidosa gen. et sp. nov. The cells are spherical, mean 11.2 µm in diameter, and surrounded by a thick mucilaginous sheath. The grayish‐green chloroplast has many lobes extending throughout the cell and lacks a pyrenoid. This chloroplast type is similar to Glaucosphaera vacuolata, but differs from other unicellular red algae. Plastoglobuli clusters occur beneath the chloroplast envelope but only at the cell periphery. A peripheral encircling thylakoid is absent. Golgi bodies surround the central nucleus, which is an arrangement shared with all members of the Dixoniellales. The subcellular features of some mitotic phases are quite similar to those of other unicellular red algae. A pair of ring‐shaped structures located within electron‐dense material can be seen in cells undergoing telophase. The size of the polar rings ranged within those reported from the Dixoniellales. A phylogenetic analysis based on small subunit rDNA indicates that B. apyrenoidosa is a member of the Dixoniellales and a sister lineage to Neorhodellaand Dixoniella.     

EVOLUTIONARY ORIGIN OF GLOEOMONAS (VOLVOCALES,CHLOROPHYCEAE), BASED ON ULTRASTRUCTURE OF CHLOROPLASTS AND MOLECULAR PHYLOGENY1

Gloeomonas is a peculiar unicellular volvocalean genus because it lacks pyrenoids in the chloroplasts under the light microscope and has two flagellar bases that are remote from each other. However, ultrastructural features of chloroplasts are very limited, and no molecular phylogenetic analyses have been carried out in Gloeomonas. In this study, we observed ultrastructural features of chloroplasts of three species of Gloeomonas and Chloromonas rubrifilum (Korshikov ex Pascher) Pröschold, B. Marin, U. Schlösser et Melkonian SAG 3.85, and phylogenetic analyses were carried out based on the combined data set from 18S rRNA, ATP synthase beta‐subunit, and P700 chl a–apoprotein A2 gene sequences to deduce the natural phylogenetic positions of the genus Gloeomonas. The present EM demonstrated that the chloroplasts of the three Gloeomonas species and C. rubrifilum SAG 3.85 did not have typical pyrenoids with associated starch grains, but they possessed pyrenoid matrices that protruded interiorly within the stroma regions of the chloroplast. The pyrenoid matrices were large and broad in C. rubrifilum, whereas those of the three Gloeomonas species were recognized in only the small protruded regions of the chloroplast lobes. The present multigene phylogenetic analyses resolved that the three species of Gloeomonas belong to the Chloromonas lineage or Chloromonadinia of the Volvocales, and Chloromonas insignis (Anakhin) Gerloff et H. Ettl NIES‐447 and C. rubrifilum SAG 3.85, both of which have pyrenoids without associated starch grains, were positioned basally to the clade composed of the three species of Gloeomonas. Therefore, Gloeomonas might have evolved from such a Chloromonas species through reduction in pyrenoid matrix size within the chloroplast and by separating their two flagellar bases.