Mychonastes sp. 246 Suppresses Human Pancreatic Cancer Cell Growth via IGFBP3-PI3K-mTOR Signaling.

Previously, we confirmed that Mychonastes sp. 246 methanolic extract (ME) markedly reduced the viability of BxPC-3 human pancreatic cancer cells. However, the underlying mechanism ME remained unclear. Hence, we attempted to elucidate the anticancer effect of ME on BxPC-3 human pancreatic cancer cells. First, we investigated the components of ME and their cytotoxicity in normal cells. Then, we confirmed the G1 phase arrest mediated growth inhibitory effect of ME using a cell counting assay and cell cycle analysis. Moreover, we found that the migration-inhibitory effect of ME using a Transwell migration assay. Through RNA sequencing, Gene Ontology-based network analysis, and western blotting, we explored the intracellular mechanisms of ME in BxPC-3 cells. ME modulated the intracellular energy metabolism-related pathway by altering the mRNA levels of IGFBP3 and PPARGC1A in BxPC-3 cells and reduced PI3K and mTOR phosphorylation by upregulating IGFBP3 and 4E-BP1 expression. Finally, we verified that ME reduced the growth of three-dimensional (3D) pancreatic cancer spheroids. Our study demonstrates that ME suppresses pancreatic cancer proliferation through the IGFBP3-PI3K-mTOR signaling pathway. This is the first study on the anticancer effect of the ME against pancreatic cancer, suggesting therapeutic possibilities and the underlying mechanism of ME action.

Improvement of Lutein and Zeaxanthin Production in Mychonastes sp. 247 by Optimizing Light Intensity and Culture Salinity Conditions.

In this study, we sought to improve lutein and zeaxanthin production in Mychonastes sp. 247 and investigated the effect of environmental factors on lutein and zeaxanthin productivity in Mychonastes sp. The basic medium selection and N:P ratio were adjusted to maximize cell growth in one-stage culture, and lutein and zeaxanthin production conditions were optimized using a central composite design for two-stage culture. The maximum lutein production was observed at a light intensity of 60 µE/m2/s and salinity of 0.49%, and the maximum zeaxanthin production was observed at a light intensity of 532 µE/m2/s and salinity of 0.78%. Lutein and zeaxanthin production in the optimized medium increased by up to 2 and 2.6 folds, respectively, compared to that in the basic medium. Based on these results, we concluded that the optimal conditions for lutein and zeaxanthin production are different and that optimization of light intensity and culture salinity conditions may help increase carotenoid production. This study presents a useful and potential strategy for optimizing microalgal culture conditions to improve the productivity of lutein and zeaxanthin, which has applications in the functional food field.

Gossypol Induces Apoptosis of Human Pancreatic Cancer Cells via CHOP/Endoplasmic Reticulum Stress Signaling Pathway.

Gossypol, a natural phenolic aldehyde present in cotton plants, was originally used as a means of contraception, but is currently being studied for its anti-proliferative and anti-metastatic effects on various cancers. However, the intracellular mechanism of action regarding the effects of gossypol on pancreatic cancer cells remains unclear. Here, we investigated the anti-cancer effects of gossypol on human pancreatic cancer cells (BxPC-3 and MIA PaCa-2). Cell counting kit-8 assays, annexin V/propidium iodide staining assays, and transmission electron microscopy showed that gossypol induced apoptotic cell death and apoptotic body formation in both cell lines. RNA sequencing analysis also showed that gossypol increased the mRNA levels of CCAAT/enhancer-binding protein homologous protein (CHOP) and activating transcription factor 3 (ATF3) in pancreatic cancer cell lines. In addition, gossypol facilitated the cleavage of caspase-3 via protein kinase RNA-like ER kinase (PERK), CHOP, and Bax/Bcl-2 upregulation in both cells, whereas the upregulation of ATF was limited to BxPC-3 cells. Finally, a three-dimensional culture experiment confirmed the successful suppression of cancer cell spheroids via gossypol treatment. Taken together, our data suggest that gossypol may trigger apoptosis in pancreatic cancer cells via the PERK-CHOP signaling pathway. These findings propose a promising therapeutic approach to pancreatic cancer treatment using gossypol.

Limnovirga soli gen. nov., sp. nov., isolated from river sediment.

A cream-coloured, Gram-stain-negative, rod-shaped bacterium, designated strain KSC-6T, was isolated from soil sampled at the Gapcheon River watershed in Daejeon, Republic of Korea. The organism does not require NaCl for growth and grows at pH 6.0-8.0 (optimum, pH 7.0) and 10-37 °C (optimum, 25 °C). Phylogenetic trees based on the 16S rRNA gene sequences reveal that strain KSC-6T belongs to the family Chitinophagaceae within the order Chitinophagales and is most closely related to Panacibacter ginsenosidivorans Gsoil 1550T (95.9% similarity). The genomic DNA G+C content was 38.9 mol%. The major cellular fatty acids (>8 %) of strain KCS-6T were iso-C15:0, iso-C15 : 1 G and iso-C17 : 0 3-OH. The predominant respiratory quinone was menaquinone 7 and the predominant polar lipids were phosphatidylethanolamine, five unidentified aminolipids and two unidentified lipids. Based on genome analyses, low digital DNA-DNA hybridization, average nucleotide identity and average amino acid identity values with closely related genera, and differential chemotaxonomic and physiological properties, we suggest that strain KCS-6T represents a novel species in a new genus in the family Chitinophagaceae, for which the name Limnovirga soli gen. nov., sp. nov. (type strain KCS-6T=KCCM 43337T=NBRC 114336T) is proposed.

Paucibacter aquatile sp. nov. isolated from freshwater of the Nakdong River, Republic of Korea.

A Gram-negative, aerobic, motile, and rod-shaped bacterial strain designated CR182T was isolated from freshwater of the Nakdong River, Republic of Korea. Optimal growth conditions for this novel strain were found to be: 25-30 °C, pH 6.5-8.5, and 3% (w/v) NaCl. Phylogenetic analysis based on 16S rRNA gene sequence indicates that the strain CR182T belongs to type strains of genus Paucibacter. Strain CR182T showed 98.0% 16S rRNA gene sequence similarity with Paucibacter oligotrophus CHU3T and formed a robust phylogenetic clade with this species. The average nucleotide identity value between strain CR182T and P. oligotrophus CHU3T was 78.4% and the genome-to-genome distance was 22.2% on average. The genomic DNA G+C content calculated from the genome sequence was 66.3 mol%. Predominant cellular fatty acids of strain CR182T were summed feature 3 (C16:1 ω7c and/or C16:1 ω6c) (31.2%) and C16:0 (16.0%). Its major respiratory quinine was ubiquinone Q-8. Its polar lipids consisted of diphosphatidylglycerol, phosphatidylethanolamine, and two unidentified phospholipids. Its genomic DNA G+C content was 66.3%. Based on data obtained from this polyphasic taxonomic study, strain CR182T represents a novel species belonging to genus Paucibacter, for which a name of P. aquatile sp. nov. is proposed. The type strain is CR182T (= KCCM 90284T = NBRC 113032T).

Halostella salina gen. nov., sp. nov., an extremely halophilic archaeon isolated from solar salt.

A novel halophilic archaeon designated strain CBA1114T was isolated from solar salt in the Republic of Korea. Strain CBA1114T, cells of which were coccoid and Gram-stain-negative, grew in the presence of 15-30 % (w/v) NaCl (optimum, 20 %) and at 20-50 °C (optimum, 40 °C) and pH 7.0-9.0 (optimum, pH 8.0). Strain CBA1114T required Mg2+ for growth. Strain CBA1114T had three 16S rRNA genes, rrnA, rrnB and rrnC; levels of similarity between the sequences were 99.7-99.9 %. The 16S rRNA gene sequence of strain CBA1114T showed 91.7 % similarity to that of Haloterrigena thermotolerans PR5T. In multilocus sequence analysis (MLSA), five housekeeping genes, atpB, EF-2, radA, rpoB' and secY, were found to be closely related to those of the members of the genera Halorientalis(89.7 % similarity of the atpB gene sequence), Halomicroarcula(91.9 %, EF-2), Haloterrigena(85.4 %, radA), Natronoarchaeum(89.2 %, rpoB') and Natrinema(75.7 %, secY). A phylogenetic tree generated from the results of MLSA of the five housekeeping genes showed that strain CBA1114T was closely related to species of the genus Halorientalis in the family Halobacteriaceae. The major polar lipids were identified as phosphatidylglycerol, phosphatidylglycerol phosphate methyl ester and unidentified lipids. The G+C content of the genomic DNA of strain CBA1114T was 68.1 mol%. According to the results of phylogenetic, phenotypic and chemotaxonomic analyses, we designate strain CBA1114T (=JCM 30111T=KCTC 4206T) as the type strain of Halostella salina gen. nov., sp. nov., a novel species of a new genus within the family Halobacteriaceae.

Occurrence of viable, red-pigmented haloarchaea in the plumage of captive flamingoes.

Flamingoes (Phoenicopterus spp.) whose plumage displays elegant colors, inhabit warm regions close to the ocean throughout the world. The pink or reddish color of their plumage originates from carotenoids ingested from carotenoid-abundant food sources, since flamingoes are unable to synthesize these compounds de novo. In this study, viable red-colored archaeal strains classified as extremely halophilic archaea (i.e., haloarchaea) and belonging to the genera Halococcus and Halogeometricum were isolated from the plumage of flamingoes in captivity. Detailed analysis for haloarchaeal community structure in flamingo feathers based on metagenomic data identified several haloarchaeal genera and unclassified sequences of the class Halobacteria at the genus level. Carotenoid pigment analyses showed that a bacterioruberin precursor carotenoid in haloarchaea was identical to one of the pigments found in flamingo plumage. To the best of our knowledge, this is the first report of viable extremophilic archaea in avian plumage, thus contributing to our understanding of the ecology of haloarchaea. The potential influence of haloarchaea as an environmental factor determining avian plumage coloration should be investigated in further studies.

Draft genome sequence of the extremely halophilic archaeon Haladaptatus cibarius type strain D43(T) isolated from fermented seafood.

An extremely halophilic archaeon, Haladaptatus cibarius D43(T), was isolated from traditional Korean salt-rich fermented seafood. Strain D43(T) shows the highest 16S rRNA gene sequence similarity (98.7 %) with Haladaptatus litoreus RO1-28(T), is Gram-negative staining, motile, and extremely halophilic. Despite potential industrial applications of extremely halophilic archaea, their genome characteristics remain obscure. Here, we describe the whole genome sequence and annotated features of strain D43(T). The 3,926,724 bp genome includes 4,092 protein-coding and 57 RNA genes (including 6 rRNA and 49 tRNA genes) with an average G + C content of 57.76 %.

Genome sequence of the haloarchaeon Haloterrigena jeotgali type strain A29(T) isolated from salt-fermented food.

Haloterrigena jeotgali is a halophilic archaeon within the family Natrialbaceae that was isolated from shrimp jeotgal, a traditional Korean salt-fermented food. A29(T) is the type strain of H. jeotgali, and is a Gram-negative staining, non-motile, rod-shaped archaeon that grows in 10 %-30 % (w/v) NaCl. We present the annotated H. jeotgali A29(T) genome sequence along with a summary of its features. The 4,131,621 bp genome with a GC content of 64.9 % comprises 4,215 protein-coding genes and 127 RNA genes. The sequence can provide useful information on genetic mechanisms that enable haloarchaea to endure a hypersaline environment.

Croceitalea litorea sp. nov., isolated from seashore sand.

Strain CBA3205T is a Gram-stain-negative, non-motile and rod-shaped bacterium that was isolated from the seashore sand of Jeju Island in South Korea. Based on the phylogenetic analysis, the most closed related species was Croceitalea eckloniae DOKDO 025T, with 94.8 % sequence similarity for the 16S rRNA gene. Strain CBA3205T was observed to grow optimally at 25-30 °C and at pH 8.5 in the presence of 2-3 % (w/v) NaCl. The major fatty acids of strain CBA3205T were iso-C15 : 0, iso-C15 : 1 G, and iso-C17 : 0 3-OH. The major respiratory quinone was MK-6 and the major polar lipids were two unidentified amino-group-containing phospholipids and an unidentified polar lipid. The G+C content of the genomic DNA of strain CBA3205T was 62.5 mol%. Based on the phenotypic, genotypic and phylogenetic analyses, strain CBA3205T was considered to be a novel species belonging to the genus Croceitalea within the family Flavobacteriaceae, for which the name Croceitalea litorea sp. nov. is proposed. The type strain is CBA3205T ( = KACC 17669T = JCM 19531T).

Halolamina sediminis sp. nov., an extremely halophilic archaeon isolated from solar salt.

An extremely halophilic archaeal strain, halo-7T, was isolated from brine sediment of the Gomso solar saltern, Republic of Korea. Cells of strain halo-7T were pleomorphic, stained Gram-negative, lysed in distilled water and formed red-pigmented colonies. Strain halo-7T grew in the range of 25-45°C (optimum 37-40°C), pH 6.5-9.5 (optimum pH 7.0-8.0), 15-30% (w/v) NaCl (optimum, 20-25%), and 0.05-0.5 M MgCl2 (optimum 0.1-0.3 M). The minimal NaCl concentration to prevent cell lysis of strain halo-7T was 10% (w/v). The major polar lipids of the isolate were phosphatidylglycerol, phosphatidylglycerolphosphate methyl ester, an unidentified phospholipid, an unidentified lipid, and two unidentified glycolipids. Phylogenetic analysis based on 16S rRNA gene sequences showed that strain halo-7T is closely related to the members of the genus Halolamina, Halolamina salina WSY15-H3T (98.7% 16S rRNA gene sequence similarity), Halolamina pelagica TBN21T (98.2%) and Halolamina rubra CBA1107T (97.4%). The genomic DNA G+C content determined for strain halo-7T (68.0 mol%) was slightly higher than those of H. salina JCM 18549T and H. rubra CBA1107T. DNA-DNA hybridization values between strain halo-7T and reference strains were < 25%. Based on phenotypic, chemotaxonomic and phylogenetic properties, we describe a novel species of the genus Halolamina, represented by strain halo-7T, for which we propose the name Halolamina sediminis sp. nov. The type strain is halo-7T ( = JCM 30187T = CECT 8739T).

Draconibacterium filum sp. nov., a new species of the genus of Draconibacterium from sediment of the east coast of the Korean Peninsula.

A Gram-stain negative, long rod shaped, facultatively anaerobic bacterium, designated strain F2(T), was isolated from coastal sediment of the Korean Peninsula. Strain F2(T) was found to grow at 10-40 °C (optimum 30 °C), at pH 6.0-8.5 (optimum pH 7.5) and at 0.0-8.0 % (w/v) NaCl (optimum 3.0 %). Phylogenetic analysis of the 16S rRNA gene sequence showed that strain F2(T) is closely related to Draconibacterium orientale FH5(T) (with 97.9 % 16S rRNA gene similarity) of the family Prolixibacteraceae of the phylum Bacteroidetes. The major isoprenoid quinone was identified as MK-7 and the main fatty acids as iso-C15:0 (24.1 %), anteiso-C15:0 (15.4 %), C16:0 (10.7 %), iso-C17:0 3-OH (7.6 %) and iso-C16:0 3-OH (5.9 %). The major polar lipids were identified as phosphatidylethanolamine, phosphatidylinositol and four unidentified polar lipids. The genomic DNA G+C content of strain F2(T) was determined to be 44.7 mol% and the DNA-DNA relatedness of strain F2(T) with D. orientale DSM 25947(T) was 34.6 ± 4.3 %. Nitrate reduction capability and cell morphology of strain F2(T) are distinct from those of the closest relative, D. orientale DSM 25947(T). Based on these properties, we propose strain F2(T) represents a novel species within the genus Draconibacterium, with the name Draconibacterium filum sp. nov. The type strain of D. filum is F2(T) (=KCTC 32486(T) = JCM 19986(T)).

Vulcanisaeta thermophila sp. nov., a hyperthermophilic and acidophilic crenarchaeon isolated from solfataric soil.

An anaerobic, rod-shaped, hyperthermophilic and acidophilic crenarchaeon, designated strain CBA1501(T), was isolated from solfataric soil of the Mayon volcano in the Republic of the Philippines. Phylogenetic analysis showed that strain CBA1501(T) is affiliated with the genus Vulcanisaeta in the phylum Crenarchaeota. DNA sequence similarities between the 16S rRNA gene of strain CBA1501(T) and those of Vulcanisaeta distributa IC-017(T) and Vulcanisaeta souniana IC-059(T) were 98.5 and 97.4 %, respectively. Strain CBA1501(T) grew between 75-90 °C, over a pH range of 4.0-6.0 and in the presence of 0-1.0 % (w/v) NaCl, with optimal growth occurring at 85 °C, pH 5.0, and with 0 % (w/v) NaCl. Fumarate, malate, oxidized glutathione, sulfur and thiosulfate were used as final electron acceptors, but FeCl3, nitrate and sulfate were not. The DNA G+C content of strain CBA1501(T) was 43.1 mol%. On the basis of polyphasic taxonomic analysis, strain CBA1501(T) represents a novel species of the genus Vulcanisaeta in the phylum Crenarchaeota, for which we propose the name Vulcanisaeta thermophila sp. nov. The type strain is CBA1501(T) ( = ATCC BAA-2415(T) = JCM 17228(T)).

Phylogenetic analysis of microalgae based on highly abundant proteins using mass spectrometry.

The blooms of toxic phototrophic microorganisms, such as microalgae and cyanobacteria, which are typically found in freshwater and marine environments, are becoming more frequent and problematic in aquatic systems. Due to accumulation of toxic algae, harmful algal blooms (HABs) exert negative effects on aquatic systems. Therefore, rapid detection of harmful microalgae is important for monitoring the occurrence of HABs. Mass spectrometry-based methods have become sensitive, specific techniques for the identification and characterization of microorganisms. Matrix-assisted laser desorption/ionization (MALDI) with time-of-flight (TOF) mass spectrometry (MS) allows us to measure a unique molecular fingerprint of highly abundant proteins in a microorganism and has been used for the rapid, accurate identification of bacteria and fungi in clinical microbiology. Here, we tested the specificity of MALDI-TOF MS using microalgal strains (Heterocapsa, Alexandrium, Nannochloropsis, Chaetoceros, Chlorella, and Dunaliella spp.). Our research suggested that this method was comparable in terms of the rapid identification of microalgea to conventional methods based on genetic information and morphology. Thus, this efficient mass spectrometry-based technique may have applications in the rapid identification of harmful microorganisms from aquatic environmental samples.

Draft genome sequence of the extremely halophilic archaeon Halococcus sediminicola CBA1101T isolated from a marine sediment sample.

Halococcus sediminicola CBA1101T (=CECT 8275T, JCM 18965T) is an extremely halophilic strain isolated from a marine sediment collected from the bay of Gangjin in the Republic of Korea. We found that the draft genome of H. sediminicola CBA1101T contains 3,764,367bp, with 62.3% G+C content. This is the one of the few genomes to be sequenced in the genus Halococcus.

Draft genome sequence of Halapricum salinum CBA1105(T), an extremely halophilic archaeon isolated from solar salt.

Halapricum salinum CBA1105(T) (=KCTC 4202(T), JCM 19729(T)) is an extremely halophilic archaeon isolated from solar salt in the Republic of Korea. We present the draft genome of CBA1105(T), which is assembled into 3 contigs containing 3,451,492bp with a G+C content of 63.7%. This is the first genome that has been sequenced in the genus Halapricum.

Draft genome sequence of Halolamina rubra CBA1107(T), an agarolytic haloarchaeon isolated from solar salt.

Halolamina rubra CBA1107(T) (=CECT 8421(T), JCM 19436(T)), an extremely halophilic archaeon, was isolated from non-purified solar salt in the Republic of Korea. H. rubra CBA1107(T) shows agarase activity, and its draft genome contains 2955,064bp with a G+C content of 69.0%. This is the first genome that has been sequenced in the genus Halolamina.

Draft genome sequence of Halorubrum halophilum B8(T), an extremely halophilic archaeon isolated from salt-fermented seafood.

The extremely halophilic archaeon, Halorubrum halophilum B8(T) (=JCM 18963(T), CECT 8278(T)) was isolated from salt-fermented seafood. We report here the draft genome of H. halophilum B8(T), containing 3,677,984bp with a G+C content of 65.1%. The genome consists of 19 genes predicted to encode esterases.