Arctic Sea Ice Microalga Chlamydomonas latifrons KNF0041: Identification and Statistical Optimization of Medium for Enhanced Biomass and Omega-3/Omega-6.

Polar microorganisms produce biologically active compounds that enable them to survive in harsh environments. These compounds have potential biomedical applications. The green microalga Chlamydomonas latifrons KNF0041, isolated from Arctic sea ice, has been found to produce polyunsaturated fatty acids (PUFAs), including omega-3 and omega-6, which have antioxidant properties. To improve the biomass production of strain KNF0041, statistical methods such as the Plackett-Burman design, Box-Behnken design, and response surface methodology (RSM) were utilized for medium optimization. The optimized medium was designed with increased potassium phosphate content and reduced acetic acid (AcOH) content. The use of the optimized medium resulted in an increase in the cell number as biomass of strain KNF0041 by 34.18% and the omega-3 and omega-6 fatty acid (FA) content by 10.04% and 58.29%, respectively, compared to that in normal TAP medium, which is known as the growth medium for Chlamydomonas culture. In this study, Chlamydomonas latifrons was discovered for the first time in the polar region and identified using morphology and molecular phylogenetic analyses, the secondary structures of the internal transcribed spacers, and optimized culture conditions. The results of this study provide an efficient method for the application of polar microalgae for the production of bioactive compounds.

The glacier melting process is an invisible barrier to the development of Antarctic subtidal macroalgal assemblages.

Ecological macroalgal succession in glacier-free areas has remained at the pioneer seral stage despite six decades of glacial retreat at Marian Cove, King George Island, Antarctica. With the rapid melting of glaciers in the West Antarctic Peninsula owing to global warming, a massive amount of meltwater is flowing into the coast, creating marine environmental gradients such as turbidity, water temperature, and salinity. This study examined the spatial and vertical distributions (up to a depth of 25 m) of macroalgal assemblages from nine sites in Maxwell Bay and Marian Cove. The macroalgal assemblages were analyzed for six sites located 0.2, 0.8, 1.2, 2.2, 3.6, and 4.1 km away from the glacier, including three sites where the glacial retreat history of Marian Cove could be estimated. To investigate the effects of meltwater, differences in the coastal environment were analyzed based on data collected from five stations located 0.4, 0.9, 3.0, 4.0, and 5.0 km away from the glacier. The macroalgal assemblages and marine environment were divided into two groups-inside and outside the cove-based on the region 2-3 km away from the glacier, which has been ice-free since 1956 and shows significant differences. In the three sites near the glacier front, Palmaria decipiens was dominant, and three to four species were distributed, whereas in the two sites outside the cove, nine and 14 species appeared, respectively, similar to the assemblage of the other three sites in Maxwell Bay. Palmaria decipiens, a representative opportunistic pioneer species in Antarctica, is dominant because of its physiological adaptation despite the high turbidity and low water temperature of the glacier front. This study shows that macroalgal assemblages in Antarctic fjord-like coves respond to glacial retreat and are valuable in understanding the macroalgal succession in Antarctica.

Mitochondrial genome of the Antarctic microalga Micractinium simplicissimum KSF0127 (Chlorellaceae, Trebouxiophyceae).

We report the first mitochondrial genome of the Antarctic microalga Micractinium simplicissimum KSF0127. The circular mitochondrial genome was 67,923 bp in length and contained 45 protein-coding genes, one ribosomal RNA gene, and 60 transfer RNA genes. The phylogenetic tree was constructed with eight previously reported mitogenome sequences and showed the phylogenetic position of M. simplicissimum KSF0127 within the Chlorellaceae family.

30 years revisit survey for long-term changes in the Antarctic subtidal algal assemblage.

A long-term change of a subtidal macroalgal assemblage has been investigated in Maxwell Bay, King George Island (KGI) of the Antarctic coast by a revisit survey after 30 years. Field surveys were done by SCUBA diving at six sites in 2016-2018 to directly compare with the previous survey conducted in 1988-1993 at the same sites. The total number of macroalgal species was similar between the previous and the present survey, 25 and 27 species respectively. However, the macroalgal assemblage changed substantially with the average similarity of 48.2% between the two surveys. Also, the species-level abundance showed a high variability between surveys. On the other hand, over the 30 years interval there was little overall change at the between-site level hierarchical structure in the subtidal communities of Maxwell Bay. The sites near the penguin rookery consistently showed the highest biodiversity, indicating the importance of land-based nutrients input in Antarctic coastal habitats. A noticeable pattern change over 30 years was the increase of Desmarestia complex and Plocamium cartilagineum and the decrease of Himantothallus grandifolius. Both groups are still dominant, but the shift from Himantothallus to Desmarestia-Plocamium may reflects temperature rise on the Maxwell Bay coast compared to the past.

Co-production of biodiesel and bioethanol using psychrophilic microalga Chlamydomonas sp. KNM0029C isolated from Arctic sea ice.

BACKGROUND: Biofuels, generated using microalgae as sustainable energy, have received a lot of attention. Microalgae can be cultivated at low cost with CO2 and solar energy without competition from edible crops. Psychrophilic microalgae can be a suitable feedstock to produce biofuels without the environmental constraints of low temperatures, because they can grow below 10 °C. However, there is a lack of efficient strategies using psychrophilic microalgae to produce biodiesel and bioethanol. Therefore, the current study aimed to optimize the production of biodiesel and bioethanol from Arctic Chlamydomonas sp. KNM0029C at low temperatures. RESULTS: After incubation in a 20-L photobioreactor, fatty acid methyl ester (FAME) was extracted using modified FAME extraction methods, producing a maximum yield of 0.16-g FAME/g KNM0029C. Residual biomass was pretreated for bioethanol production, and the yields from different methods were compared. The highest bioethanol yield (0.22-g/g residual biomass) was obtained by pretreatment with enzyme (amyloglucosidase) after sonication. Approximately 300-mg biofuel was obtained, including 156-mg FAME biodiesel and 144-mg bioethanol per g dried cells, representing the highest recorded yield from psychrophilic microalgae. CONCLUSIONS: This is the first to attempt at utilizing biomass from psychrophilic Arctic microalga Chlamydomonas sp. KNM0029C for the co-production of bioethanol and biodiesel, and it yielded the highest values among reported studies using psychrophilic organisms. These results can be used as a source for the efficient biofuel production using polar microalgae.

Composition and structure of the marine benthic community in Terra Nova Bay, Antarctica: Responses of the benthic assemblage to disturbances.

The community structure and assemblages of marine benthic organisms were investigated in coastal areas near the Jang Bogo Antarctic Research Station in Terra Nova Bay during the 2012-2018 summer seasons. We also examined the recovery pattern of marine benthic organisms following disturbance due to the construction of the Jang Bogo Station. A total of 26 taxa were identified in the study area during the experimental period. Species number and diversity indices (richness, evenness, and diversity) were relatively low compared to data previously reported from Terra Nova Bay. Sphaerotylus antarcticus, Clavularia frankliniana, Hydractinia sp., Iridaea cordata, Fragilariopsis spp., Alcyonium antarcticum, and Metalaeospira pixelli were the dominant species in this area. Of these, the diatom Fragilariopsis spp. were the most abundant species, indicating their key role in maintaining the marine benthic community and controlling biogeochemical cycling. During the construction of the Jang Bogo Station, sediment coverage increased and diatoms declined due to the release of sediment into the coastal area. In February 2014, one month after the disturbance due to cyclone, the diatom coverage increased dramatically and thereby species number, richness index, and diversity index steadily rose from 2015 to 2018. However, non-metric multidimensional scaling ordination analysis of species similarities among sampling times showed that community structure had not completely recovered by 2018. Thus, long-term monitoring is required to elucidate the post-disturbance settlement mechanisms of marine benthic organisms at the study area in Terra Nova Bay.

Type II Ice-Binding Proteins Isolated from an Arctic Microalga Are Similar to Adhesin-Like Proteins and Increase Freezing Tolerance in Transgenic Plants.

Microalgal ice-binding proteins (IBPs) in the polar region are poorly understood at the genome-wide level, although they are important for cold adaptation. Through the transcriptome study with the Arctic green alga Chloromonas sp. KNF0032, we identified six Chloromonas IBP genes (CmIBPs), homologous with the previously reported IBPs from Antarctic snow alga CCMP681 and Antarctic Chloromonas sp. They were organized with multiple exon/intron structures and low-temperature-responsive cis-elements in their promoters and abundantly expressed at low temperature. The biological functions of three representative CmIBPs (CmIBP1, CmIBP2 and CmIBP3) were tested using in vitro analysis and transgenic plant system. CmIBP1 had the most effective ice recrystallization inhibition (IRI) activities in both in vitro and transgenic plants, and CmIBP2 and CmIBP3 had followed. All transgenic plants grown under nonacclimated condition were freezing tolerant, and especially 35S::CmIBP1 plants were most effective. After cold acclimation, only 35S::CmIBP2 plants showed slightly increased freezing tolerance. Structurally, the CmIBPs were predicted to have ß-solenoid forms with parallel ß-sheets and repeated TXT motifs. The repeated TXT structure of CmIBPs appears similar to the AidA domain-containing adhesin-like proteins from methanogens. We have shown that the AidA domain has IRI activity as CmIBPs and phylogenetic analysis also supported that the AidA domains are monophyletic with ice-binding domain of CmIBPs, and these results suggest that CmIBPs are a type of modified adhesins.

Draft Genome Sequence of Arthrobacter oryzae TNBS02, a Bacterium Containing Heavy Metal Resistance Genes, Isolated from Soil of Antarctica.

Arthrobacter oryzae TNBS02 was isolated from soil at Terra Nova Bay of Victoria Land, Antarctica. The genome consists of a chromosome with 4,248,670 bp which contains a total of 3,994 genes. It was identified that this strain has many heavy metal resistance genes similar to other Arthrobacter strains isolated from contaminated environments.

Lipid Production by Arctic Microalga Chlamydomonas sp. KNF0008 at Low Temperatures.

A lipid-producing microalga, Chlamydomonas sp. KNF0008, collected from the Arctic was capable of growing at temperatures ranging from 4 to 20 °C, and the highest cell density was measured at 15 °C and 100 µmol photons m-2 s-1 light intensity under continuous shaking and external aeration. KNF0008 showed the elevated accumulation of lipid bodies under nitrogen-deficient conditions, rather than under nitrogen-sufficient conditions. Fatty acid production of KNF0008 was 4.2-fold (104 mg L-1) higher than that of C. reinhardtii CC-125 at 15 °C in Bold's Basal Medium. The dominant fatty acids were C16:0, C16:4, C18:1, and C18:3, and unsaturated fatty acids (65.69%) were higher than saturated fatty acids (13.65%) at 15 °C. These results suggested that Arctic Chlamydomonas sp. KNF0008 could possibly be utilized for production of biodiesel during periods of cold weather because of its psychrophilic characteristics.

Mitochondrial and Plastid Genomes from Coralline Red Algae Provide Insights into the Incongruent Evolutionary Histories of Organelles.

Mitochondria and plastids are generally uniparentally inherited and have a conserved gene content over hundreds of millions of years, which makes them potentially useful phylogenetic markers. Organelle single gene-based trees have long been the basis for elucidating interspecies relationships that inform taxonomy. More recently, high-throughput genome sequencing has enabled the construction of massive organelle genome databases from diverse eukaryotes, and these have been used to infer species relationships in deep evolutionary time. Here, we test the idea that despite their expected utility, conflicting phylogenetic signal may exist in mitochondrial and plastid genomes from the anciently diverged coralline red algae (Rhodophyta). We generated complete organelle genome data from five coralline red algae (Lithothamnion sp., Neogoniolithon spectabile, Renouxia sp., Rhodogorgon sp., and Synarthrophyton chejuensis) for comparative analysis with existing organelle genome data from two other species (Calliarthron tuberculosum and Sporolithon durum). We find strong evidence for incongruent phylogenetic signal from both organelle genomes that may be explained by incomplete lineage sorting that has maintained anciently derived gene copies or other molecular evolutionary processes such as hybridization or gene flow during the evolutionary history of coralline red algae.

Genome sequencing of the winged midge, Parochlus steinenii, from the Antarctic Peninsula.

Background: In the Antarctic, only two species of Chironomidae occur naturally-the wingless midge, Belgica antarctica , and the winged midge, Parochlus steinenii . B. antarctica is an extremophile with unusual adaptations. The larvae of B. antarctica are desiccation- and freeze-tolerant and the adults are wingless. Recently, the compact genome of B. antarctica was reported and it is the first Antarctic eukaryote to be sequenced. Although P. steinenii occurs naturally in the Antarctic with B. antarctica , the larvae of P. steinenii are cold-tolerant but not freeze-tolerant and the adults are winged. Differences in adaptations in the Antarctic midges are interesting in terms of evolutionary processes within an extreme environment. Herein, we provide the genome of another Antarctic midge to help elucidate the evolution of these species. Results: The draft genome of P. steinenii had a total size of 138 Mbp, comprising 9513 contigs with an N50 contig size of 34,110 bp, and a GC content of 32.2%. Overall, 13,468 genes were predicted using the MAKER annotation pipeline, and gene ontology classified 10,801 (80.2%) predicted genes to a function. Compared with the assembled genome architecture of B. antarctica , that of P. steinenii was approximately 50 Mbp longer with 6.2-fold more repeat sequences, whereas gene regions were as similarly compact as in B. antarctica . Conclusions: We present an annotated draft genome of the Antarctic midge, P. steinenii . The genomes of P. steinenii and B. antarctica will aid in the elucidation of evolution in harsh environments and provide new resources for functional genomic analyses of the order Diptera.

Enzymatic modification by point mutation and functional analysis of an omega-6 fatty acid desaturase from Arctic Chlamydomonas sp.

Arctic Chlamydomonas sp. is a dominant microalgal strain in cold or frozen freshwater in the Arctic region. The full-length open reading frame of the omega-6 fatty acid desaturase gene (AChFAD6) was obtained from the transcriptomic database of Arctic Chlamydomonas sp. from the KOPRI culture collection of polar micro-organisms. Amino acid sequence analysis indicated the presence of three conserved histidine-rich segments as unique characteristics of omega-6 fatty acid desaturases, and three transmembrane regions transported to plastidic membranes by chloroplast transit peptides in the N-terminal region. The AChFAD6 desaturase activity was examined by expressing wild-type and V254A mutant (Mut-AChFAD6) heterologous recombinant proteins. Quantitative gas chromatography indicated that the concentration of linoleic acids in AChFAD6-transformed cells increased more than 3-fold [6.73 ± 0.13 mg g-1 dry cell weight (DCW)] compared with cells transformed with vector alone. In contrast, transformation with Mut-AChFAD6 increased the concentration of oleic acid to 9.23 ± 0.18 mg g-1 DCW, indicating a change in enzymatic activity to mimic that of stearoyl-CoA desaturase. These results demonstrate that AChFAD6 of Arctic Chlamydomonas sp. increases membrane fluidity by enhancing denaturation of C18 fatty acids and facilitates production of large quantities of linoleic fatty acids in prokaryotic expression systems.

Characterization of Stearoyl-CoA Desaturases from a Psychrophilic Antarctic Copepod, Tigriopus kingsejongensis.

Stearoyl-CoA desaturase is a key regulator in fatty acid metabolism that catalyzes the desaturation of stearic acid to oleic acid and controls the intracellular levels of monounsaturated fatty acids (MUFAs). Two stearoyl-CoA desaturases (SCD, Δ9 desaturases) genes were identified in an Antarctic copepod, Tigriopus kingsejongensis, that was collected in a tidal pool near the King Sejong Station, King George Island, Antarctica. Full-length complementary DNA (cDNA) sequences of two T. kingsejongensis SCDs (TkSCDs) were obtained from next-generation sequencing and isolated by reverse transcription PCR. DNA sequence lengths of the open reading frames of TkSCD-1 and TkSCD-2 were determined to be 1110 and 681 bp, respectively. The molecular weights deduced from the corresponding genes were estimated to be 43.1 kDa (TkSCD-1) and 26.1 kDa (TkSCD-2). The amino acid sequences were compared with those of fatty acid desaturases and sterol desaturases from various organisms and used to analyze the relationships among TkSCDs. As assessed by heterologous expression of recombinant proteins in Escherichia coli, the enzymatic functions of both stearoyl-CoA desaturases revealed that the amount of C16:1 and C18:1 fatty acids increased by greater than 3-fold after induction with isopropyl ß-D-thiogalactopyranoside. In particular, C18:1 fatty acid production increased greater than 10-fold in E. coli expressing TkSCD-1 and TkSCD-2. The results of this study suggest that both SCD genes from an Antarctic marine copepod encode a functional desaturase that is capable of increasing the amounts of palmitoleic acid and oleic acid in a prokaryotic expression system.

[Not Available].

In this study, a new "non-oxytrichid Dorsomarginalia" ciliate, Urosomoida sejongensis n. sp. discovered from freshwater of the King George Island, Antarctica, was investigated using morphological, morphometrical, and molecular methods. Morphology of U. sejongensis is characterized as follows: body shape slender to elongated; cortical granules spherical and colorless, groups of granules formed patchy distribution; ring-shaped structures scattered in cytoplasm; 27-30 adoral membranelles with undulating membranes in Oxytricha pattern; usually 17 frontal-ventral-transverse (FVT) cirri composed of 3 frontal, 1 buccal, 4 frontoventral, 3 postoral ventral, 2 pretransverse ventral and 4 transverse cirri; 1 right and 1 left marginal rows; 3 dorsal kineties with 1 dorsomarginal row, 3 caudal cirri; 1 micronucleus between 2 macronuclear nodules. This new species mainly differs from other congeners by the combination of following morphological features: a micronucleus, cortical granules, and ciliatures (e.g., adoral membranelles, FVT cirri). Urosomoida sejongensis shows a nucleotide similarity of 97.3% with U. agilis, type of this genus, using the SSU rDNA sequence. Molecular phylogeny shows a non-monophyletic relationship among Urosomoida species and emphasizes the need for further morphogenetic studies of this genus and other related species to resolve morphological convergences.

Genetic and Molecular Epidemiological Characterization of a Novel Adenovirus in Antarctic Penguins Collected between 2008 and 2013.

Antarctica is considered a relatively uncontaminated region with regard to the infectious diseases because of its extreme environment, and isolated geography. For the genetic characterization and molecular epidemiology of the newly found penguin adenovirus in Antarctica, entire genome sequencing and annual survey of penguin adenovirus were conducted. The entire genome sequences of penguin adenoviruses were completed for two Chinstrap penguins (Pygoscelis antarctica) and two Gentoo penguins (Pygoscelis papua). The whole genome lengths and G+C content of penguin adenoviruses were found to be 24,630-24,662 bp and 35.5-35.6%, respectively. Notably, the presence of putative sialidase gene was not identified in penguin adenoviruses by Rapid Amplification of cDNA Ends (RACE-PCR) as well as consensus specific PCR. The penguin adenoviruses were demonstrated to be a new species within the genus Siadenovirus, with a distance of 29.9-39.3% (amino acid, 32.1-47.9%) in DNA polymerase gene, and showed the closest relationship with turkey adenovirus 3 (TAdV-3) in phylogenetic analysis. During the 2008-2013 study period, the penguin adenoviruses were annually detected in 22 of 78 penguins (28.2%), and the molecular epidemiological study of the penguin adenovirus indicates a predominant infection in Chinstrap penguin population (12/30, 40%). Interestingly, the genome of penguin adenovirus could be detected in several internal samples, except the lymph node and brain. In conclusion, an analysis of the entire adenoviral genomes from Antarctic penguins was conducted, and the penguin adenoviruses, containing unique genetic character, were identified as a new species within the genus Siadenovirus. Moreover, it was annually detected in Antarctic penguins, suggesting its circulation within the penguin population.

Growth and lipid content at low temperature of Arctic alga Chlamydomonas sp. KNM0029C.

Biodiesel produced from microalgae is a promising source of alternative energy. In winter, however, outdoor mass cultivation for biodiesel production is hampered by poor growth. Here, we report that Arctic Chlamydomonas sp. KNM0029C exhibits optimal growth at 4 °C and reaches densities up to 1.4 × 10(7) cells mL(-1). Lipid body formation in the alga was visualized through BODIPY 505/515 staining and fluorescence microscopy. The fatty acid methyl ester (FAME) production level of KNM0029C was 178.6 mg L(-1) culture and 2.3-fold higher than that of C. reinhardtii CC-125 at 4 °C. Analysis of the FAME content showed a predominance of polyunsaturated fatty acids such as C16:3, C18:2, C18:3, and C20:2. C18:3 fatty acids comprised the largest fraction (20.7%), and the content of polyunsaturated fatty acids (39.6%) was higher than that of saturated fatty acids (6.8%) at 4 °C. These results indicate that Chlamydomonas sp. KNM0029C, as a psychrophilic microalga, might represent a favorable source for biodiesel production in cold environments.

Complete mitochondrial genome of the South Polar Skua Stercorarius maccormicki (Charadriiformes, Stercorariidae) in Antarctica.

The South Polar Skua, gull-like seabirds is the most fascinating Antarctic seabirds that lay two eggs at sites free of snow and ice and predominantly hunt pelagic fish and penguins. Blood samples of the South Polar Skua Stercorarius maccormicki was collected during the summer activity near King Sejong station in Antarctica. The complete mitochondrial DNA sequence of S. maccormicki was 16,669 bp, showing conserved genome structure and orientation found in other avian species. The control region of S. maccormicki was 93- and 80 bp shorter compared to those of Chroicocephalus saundersi and Synthliboramphus antiquus respectively. Interestingly, there is a (CAACAAACAA)6 repeat sequence in the control region. Our results of S. maccormicki mt genome including the repeat sequence, may provide useful genetic information for phylogenetic and phylogeographic histories of the southern skua complex.

Complete mitochondrial genome of the Antarctic barnacle Lepas australis (Crustacea, Maxillopoda, Cirripedia).

We present the complete mitochondrial genome of the Antarctic barnacle Lepas australis (Cirripedia, Thoracica, Lepadidae). The genome sequence is 15,502 bp in size. Except for CO1, 12 protein-coding genes (PCGs) start with an ATN initiation codon (ATA, ATG, ATC and ATT). Twelve PCGs were terminated with TAA or TAG stop codon, whereas ND1 possessed an incomplete termination codon (T- -). We compared the mitogenome structure of L. australis to those of other cirripeds and a typical arthropod Homarus americanus. The PCGs in the L. australis mtgenome showed a typical gene arrangement, identical to the arthropod pattern in other cirriped genomes. However, at least 8 tRNA genes were translocated and 2 tRNA genes were inverted in the coding polarity. Unique differences in L. australis mtgenome included translocation of trnS2, trnD and trnI. These results are useful for understanding the phylogenetic relationships among cirripedians, and additional mtgenome information of barnacles including the polar species would allow exploration of the thoracican relationships and mtgenome modifications in the barnacle evolution.

A novel adenovirus in Chinstrap penguins (Pygoscelis antarctica) in Antarctica.

Adenoviruses (family Adenoviridae) infect various organ systems and cause diseases in a wide range of host species. In this study, we examined multiple tissues from Chinstrap penguins (Pygoscelis antarctica), collected in Antarctica during 2009 and 2010, for the presence of novel adenoviruses by PCR. Analysis of a 855-bp region of the hexon gene of a newly identified adenovirus, designated Chinstrap penguin adenovirus 1 (CSPAdV-1), showed nucleotide (amino acid) sequence identity of 71.8% (65.5%) with South Polar skua 1 (SPSAdV-1), 71% (70%) with raptor adenovirus 1 (RAdV-1), 71.4% (67.6%) with turkey adenovirus 3 (TAdV-3) and 61% (61.6%) with frog adenovirus 1 (FrAdV-1). Based on the genetic and phylogenetic analyses, CSPAdV-1 was classified as a member of the genus, Siadenovirus. Virus isolation attempts from kidney homogenates in the MDTC-RP19 (ATCC® CRL-8135™) cell line were unsuccessful. In conclusion, this study provides the first evidence of new adenovirus species in Antarctic penguins.

Plastid and mitochondrion genomic sequences from Arctic Chlorella sp. ArM0029B.

BACKGROUND: Chorella is the representative taxon of Chlorellales in Trebouxiophyceae, and its chloroplast (cp) genomic information has been thought to depend only on studies concerning Chlorella vulgaris and GenBank information of C. variablis. Mitochondrial (mt) genomic information regarding Chlorella is currently unavailable. To elucidate the evolution of organelle genomes and genetic information of Chlorella, we have sequenced and characterized the cp and mt genomes of Arctic Chlorella sp. ArM0029B. RESULTS: The 119,989-bp cp genome lacking inverted repeats and 65,049-bp mt genome were sequenced. The ArM0029B cp genome contains 114 conserved genes, including 32 tRNA genes, 3 rRNA genes, and 79 genes encoding proteins. Chlorella cp genomes are highly rearranged except for a Chlorella-specific six-gene cluster, and the ArM0029B plastid resembles that of Chlorella variabilis except for a 15-kb gene cluster inversion. In the mt genome, 62 conserved genes, including 27 tRNA genes, 3 rRNA genes, and 32 genes encoding proteins were determined. The mt genome of ArM0029B is similar to that of the non-photosynthetic species Prototheca and Heicosporidium. The ArM0029B mt genome contains a group I intron, with an ORF containing two LAGLIDADG motifs, in cox1. The intronic ORF is shared by C. vulgaris and Prototheca. The phylogeny of the plastid genome reveals that ArM0029B showed a close relationship of Chlorella to Parachlorella and Oocystis within Chlorellales. The distribution of the cox1 intron at 721 support membership in the order Chlorellales. Mitochondrial phylogenomic analyses, however, indicated that ArM0029B shows a greater affinity to MX-AZ01 and Coccomyxa than to the Helicosporidium-Prototheca clade, although the detailed phylogenetic relationships among the three taxa remain to be resolved. CONCLUSIONS: The plastid genome of ArM0029B is similar to that of C. variabilis. The mt sequence of ArM0029B is the first genome to be reported for Chlorella. Chloroplast genome phylogeny supports monophyly of the seven investigated members of Chlorellales. The presence of the cox1 intron at 721 in all four investigated Chlorellales taxa indicates that the cox1 intron had been introduced in early Chorellales as a cis-splice form and that the cis-splicing intron was inherited to recent Chlorellales and was recently trans-spliced in Helicosporidium.