Ontogeny and function of the circadian clock in intestinal organoids.

Circadian rhythms regulate diverse aspects of gastrointestinal physiology ranging from the composition of microbiota to motility. However, development of the intestinal circadian clock and detailed mechanisms regulating circadian physiology of the intestine remain largely unknown. In this report, we show that both pluripotent stem cell-derived human intestinal organoids engrafted into mice and patient-derived human intestinal enteroids possess circadian rhythms and demonstrate circadian phase-dependent necrotic cell death responses to Clostridium difficile toxin B (TcdB). Intriguingly, mouse and human enteroids demonstrate anti-phasic necrotic cell death responses to TcdB. RNA-Seq analysis shows that ~3-10% of the detectable transcripts are rhythmically expressed in mouse and human enteroids. Remarkably, we observe anti-phasic gene expression of Rac1, a small GTPase directly inactivated by TcdB, between mouse and human enteroids, and disruption of Rac1 abolishes clock-dependent necrotic cell death responses. Our findings uncover robust functions of circadian rhythms regulating clock-controlled genes in both mouse and human enteroids governing organism-specific, circadian phase-dependent necrotic cell death responses, and lay a foundation for human organ- and disease-specific investigation of clock functions using human organoids for translational applications.

Histone Deacetylase Inhibition by Gut Microbe-Generated Short-Chain Fatty Acids Entrains Intestinal Epithelial Circadian Rhythms.

BACKGROUND & AIMS: The circadian clock orchestrates ∼24-hour oscillations of gastrointestinal epithelial structure and function that drive diurnal rhythms in gut microbiota. Here, we use experimental and computational approaches in intestinal organoids to reveal reciprocal effects of gut microbial metabolites on epithelial timekeeping by an epigenetic mechanism. METHODS: We cultured enteroids in media supplemented with sterile supernatants from the altered Schaedler Flora (ASF), a defined murine microbiota. Circadian oscillations of bioluminescent PER2 and Bmal1 were measured in the presence or absence of individual ASF supernatants. Separately, we applied machine learning to ASF metabolomics to identify phase-shifting metabolites. RESULTS: Sterile filtrates from 3 of 7 ASF species (ASF360 Lactobacillus intestinalis, ASF361 Ligilactobacillus murinus, and ASF502 Clostridium species) induced minimal alterations in circadian rhythms, whereas filtrates from 4 ASF species (ASF356 Clostridium species, ASF492 Eubacterium plexicaudatum, ASF500 Pseudoflavonifactor species, and ASF519 Parabacteroides goldsteinii) induced profound, concentration-dependent phase shifts. Random forest classification identified short-chain fatty acid (SCFA) (butyrate, propionate, acetate, and isovalerate) production as a discriminating feature of ASF "shifters." Experiments with SCFAs confirmed machine learning predictions, with a median phase shift of 6.2 hours in murine enteroids. Pharmacologic or botanical histone deacetylase (HDAC) inhibitors yielded similar findings. Further, mithramycin A, an inhibitor of HDAC inhibition, reduced SCFA-induced phase shifts by 20% (P < .05) and conditional knockout of HDAC3 in enteroids abrogated butyrate effects on Per2 expression. Key findings were reproducible in human Bmal1-luciferase enteroids, colonoids, and Per2-luciferase Caco-2 cells. CONCLUSIONS: Gut microbe-generated SCFAs entrain intestinal epithelial circadian rhythms by an HDACi-dependent mechanism, with critical implications for understanding microbial and circadian network regulation of intestinal epithelial homeostasis.

Thermomonas paludicola sp. nov., isolated from a lotus wetland.

A Gram-stain-negative, aerobic, rod-shaped and motile bacterium, designated IMCC34681T, was isolated from a lotus wetland in the Republic of Korea. Cellular growth occurred at 10-37 °C (optimum, 30 °C), pH 6-9 (optimum, pH 7) and with 0-2 % (w/v) NaCl (optimum, 0.5 % NaCl). The results of 16S rRNA gene sequence analysis indicated that IMCC34681T represented a member of the genus Thermomonas, sharing 95.3-96.9 % similarities with type strains of species of the genus. The whole-genome sequence of IMCC34681T was 2.72 Mbp in size with 66.2 % DNA G+C content. The IMCC34681T genome shared the highest average nucleotide identity (ANI) value, 82.8 %, with that of Thermomonas brevis KACC 16975T among species of the genus Thermomonas, indicating that the strain represents a novel genomic species. The major respiratory quinone of the strain was ubiquinone-8 (Q-8) and the predominant cellular fatty acids were iso-C15 : 0 (25.7 %) and iso-C14 : 0 (20.8 %). The strain harboured diphosphatidylglycerol, phosphatidylglycerol, phosphatidylethanolamine and an unidentified lipid as major fatty polar lipids. On the basis of the phylogenetic, phenotypic, chemotaxonomic and genomic characteristics, IMCC34681T was assigned to the genus Thermomonas as the type strain of a novel species, for which the name Thermomonas paludicola sp. nov. is proposed. The type strain is IMCC34681T (=KACC 21793T=NBRC 114635T).

Permianibacter fluminis sp. nov., isolated from a freshwater stream.

A Gram-stain-negative, aerobic, non-motile and rod-shaped bacterium, designated as IMCC34836T, was isolated from a freshwater stream. Phylogenetic analysis based on 16S rRNA gene sequences revealed that strain IMCC34836T was most closely related to Permianibacter aggregans HW001T (of the family Pseudomonadaceae) with 95.6 % sequence similarity and formed a robust clade with P. aggregans HW001T. The draft genome sequence of strain IMCC34836T was 4.4 Mbp in size with 59.1 mol% DNA G+C content. Average nucleotide identity and digital DNA-DNA hybridization values between strain IMCC34836T and P. aggregans HW001T were 71.2 and 22.0 %, respectively, indicating that the new strain represents a novel species. The strain contained iso-C15 : 0, summed feature 3 (C16 : 1 ω6c and/or C16 : 1 ω7c) and summed feature 9 (iso-C17 : 1 ω9c and/or C16 : 1 10-methyl) as the major fatty acids and harboured phosphatidylethanolamine, two unidentified aminophospholipids and three unidentified lipids as major polar lipids. The isoprenoid quinone detected in the strain was ubiquinone-8. Based on the phylogenetic and phenotypic characteristics, strain IMCC34836T is considered to represent a novel species of the genus Permianibacter, for which the name Permianibacter fluminis sp. nov. is proposed. The type strain is IMCC34836T (=KACC 21755T=NBRC 114416T).

Uliginosibacterium aquaticum sp. nov., Isolated from a Freshwater Lake.

A Gram-stain-negative, aerobic, chemoheterotrophic, rod-shaped bacterium motile by a polar flagellum, designated IMCC34675T, was isolated from Chungju Lake, an artificial freshwater reservoir in Republic of Korea. The 16S rRNA gene sequence analysis indicated that strain IMCC34675T belongs to the genus Uliginosibacterium, sharing ≤ 97.1% sequence similarities with the type strains of the genus. Whole genome sequencing of strain IMCC34675T revealed a 4.1 Mbp of genome size with 62.4% of the DNA G + C content. The IMCC34675T genome shared 73.3% of average nucleotide identity and 19.9% of digital DNA-DNA hybridization values to the genome of Uliginosibacterium gangwonense DSM 18521T, the type species of the genus. The major fatty acids of strain IMCC34675T were summed feature 3 (comprising C16:1ω6c and/or C16:1ω7c) and C16:0. The respiratory quinone detected in the strains was ubiquinone-8 (Q-8). The major polar lipids were phosphatidylethanolamine, phosphatidylglycerol, diphosphatidylglycerol, one unidentified aminophospholipid, one aminolipid, and five unidentified lipids. Based on the phylogenetic and phenotypic characterization, strain IMCC34675T was considered to represent a novel species within the genus Uliginosibacterium, for which the name Uliginosibacterium aquaticum sp. nov. is proposed with IMCC34675T (= KACC 21758T = NBRC 114418T) as the type strain.

Sphingorhabdus lacus sp. nov. and Sphingorhabdus profundilacus sp. nov., isolated from freshwater environments.

Two Gram-stain-negative, aerobic, non-motile bacteria, designated IMCC1753T and IMCC26285T, were isolated from a shallow eutrophic pond and a deep oligotrophic lake, respectively. Results of 16S rRNA gene sequence analysis indicated that the two strains shared 99.8 % sequence similarity and were most closely related to Sphingorhabdus contaminans JC216T(98.7-98.8 %). The whole genome sequences of strains IMCC1753T and IMCC26285T were 3.5 and 2.9 Mbp in size with 56.6 and 55.5 mol% DNA G+C content, respectively. Average nucleotide identity (ANI) and digital DNA-DNA hybridization (dDDH) values between the two strains were 82.2 and 25.8 %, respectively, indicating that they are separate species. The two strains showed ≤98.8 % 16S rRNA gene sequence similarities and ≤82.2 % ANI and ≤28.7 % dDDH values to closely related species of the genus Sphingorhabdus, indicating that the two strains each represent novel species. Major fatty acid constituents of strain IMCC1753T were C17 : 1 ω6c, C17 : 1 ω8c and summed features 3 (C16 : 1 ω6c and/or C16 : 1 ω7c) and 8 (C18 : 1 ω6c and/or C18 : 1 ω7c); those of strain IMCC26285T were summed features 3 and 8. The predominant isoprenoid quinone detected in both strains was ubiquinone-10 and the most abundant polyamine was spermidine. Both strains contained phosphatidylethanolamine, phosphatidylmethylethanolamine, phosphatidylcholine, phosphatidylglycerol, diphosphatidylglycerol and sphingoglycolipid as major polar lipids. On the basis of the phylogenetic and phenotypic characteristics, strains IMCC1753T and IMCC26285T were considered to represent two distinct novel species in the genus Sphingorhabdus, for which the names Sphingorhabdus lacus (IMCC1753T=KCTC 52480T=KACC 18985T=NBRC 112442T) and Sphingorhabdus profundilacus (IMCC26285T=KCTC 52479T=KACC 18986T=NBRC 112454T) are proposed, respectively.

Natronospirillum operosum gen. nov., sp. nov., a haloalkaliphilic satellite isolated from decaying biomass of a laboratory culture of cyanobacterium Geitlerinema sp. and proposal of Natronospirillaceae fam. nov., Saccharospirillaceae fam. nov. and Gynuellaceae fam. nov.

A novel haloalkaliphilic bacterium, designated G-116T, was isolated from the decaying biomass of a laboratory culture of cyanobacterium Geitlerinema species. Cells of strain G-116T were Gram-stain-negative, motile spirilla. Strain G-116T showed high halotolerance to 20 % (w/v) NaCl (optimum growth at 3.5-6.0 %, w/v) and obligately alkaliphilic growth within the pH range 7.3-10.4 (optimum growth at pH 8.7-8.9). The major fatty acids identified were C16:0, summed feature 8 (C18:1 ω7c/C18 :1 ω6c), summed feature 3 (C16:1 ω7c/C16 :1 ω6c) and C19:0 cyclo ω8c. The polar lipids consisted of phosphatidylethanolamine, phosphatidylglycerol, diphosphatidylglycerol, an unidentified phospholipid, three unidentified amino lipids and five unidentified lipids. The predominant respiratory quinone was ubiquinone-8 (Q-8). The G+C content of the genomic DNA was 60.4 mol%. Phylogenetic analysis based on 16S rRNA gene sequences revealed that the closest genus with a validly published name is a monotypic Salinispirillum and strain G-116T clustered with Salinispirillum marinum GCWy1T with a 16S rRNA gene sequence similarity of 94.3 %. Based on the data obtained from phenotypic and chemotaxonomic studies and the phylogenetic analysis, the isolate is proposed to be a representative of a novel genus and a novel species, Natronospirillum operosum gen. nov., sp. nov. Together with S. marinum they form a separate clade, for which a novel family, Natronospirillaceae fam. nov., is proposed. In addition, Saccharospirillaceae fam. nov. and Gynuellaceae fam. nov. are proposed to encompass the genera Saccharospirillum and Reinekea, and the genus Gynuella, respectively. All three novel families are within the order Oceanospirillales of the class Gammaproteobacteria. The type strain of the type species, Natronospirillum operosum gen. nov., sp. nov. is G-116T (=VKM B-3134T=KCTC 62956T).

Rhodoferax lacus sp. nov., isolated from a large freshwater lake.

A Gram-staining-negative, aerobic, motile with a single polar flagellum and rod-shaped bacterium as a bacterial host of podovirus P26218, designated IMCC26218T, was isolated from Lake Soyang, South Korea. Phylogenetic analysis based on 16S rRNA gene sequences revealed that strain IMCC26218T belonged to the genus Rhodoferax of the family Comamonadaceae and shared 97.7-99.0 % sequence similarities with Rhodoferax species. The draft whole genome sequence of strain IMCC26218T was ca. 4.9 Mbp in size with the DNA G+C content of 62.3 mol%. Average nucleotide identity (ANI) and digital DNA-DNA hybridisation (dDDH) values between strain IMCC26218T and other Rhodoferax were 74.0-77.3 % and 19.5-21.0 %, respectively, showing that the strain represents a new Rhodoferax species. The strain contained summed feature 3 (C16 : 1 ω6c and/or C16 : 1 ω7c) and C16 : 0 as the major fatty acids and phosphatidylethanolamine, three unidentified phospholipids, two unidentified aminolipids and two unidentified lipids as major polar lipids. The predominant isoprenoid quinone of the strain was ubiquinone-8 (Q-8). On the basis of the phylogenetic and phenotypic characteristics, strain IMCC26218T is considered to represent a novel species of the genus Rhodoferax, for which the name Rhodoferax lacus sp. nov. is proposed. The type strain is IMCC26218T (=KACC 18983T=NBRC 112709T).

Prophylactic use of probiotic chocolate modulates intestinal physiological functions in constipated rats.

BACKGROUND: This study investigated the in vivo prophylactic effect of probiotic chocolate on constipation. Rats were administered chocolate containing 2.5 × 1010 CFU g-1 of probiotics daily for 4 weeks and treated with loperamide (5 mg kg-1 ) daily at the fourth week of treatment. RESULTS: Probiotic chocolate treatment significantly (P < 0.05) increased the intestinal motility, colon length, fecal moisture content and number of excreted fecal pellets in constipated rats. Moreover, quantitative real-time polymerase chain reaction data and histological images also revealed that both probiotic chocolate LYC and BB12 treatments were capable of upregulating the mRNA expression levels of colonic ZO-1, occludin and AQP8, leading to the maintenance of the defensive barrier function in the constipated rats compared with the negative controls. Interestingly, these treatments also modulated gut bacterial populations by increasing the abundance levels of Lactobacillus and Bifidobacterium, as well as reducing the abundance level of Enterobacteriaceae. CONCLUSION: The present study demonstrated that probiotic chocolate LYC and BB12 could potentially be used as alternative agents for prophylactic constipation. © 2018 Society of Chemical Industry.

Flavobacterium lacicola sp. nov., isolated from a freshwater lake.

A bacterial strain, designated IMCC25901T, was isolated from a freshwater lake, Soyang, in the Republic of Korea. The strain was Gram-stain-negative, aerobic, non-motile, orange-coloured and short rod-shaped. The 16S rRNA gene sequence analysis showed that strain IMCC25901T was most closely related to Flavobacterium yonginense HMD1001T (97.0 %) and formed a robust phylogenetic clade with other species of the genus Flavobacterium. Growth of strain IMCC25901T was observed at 10-30 °C (optimum, 20 °C), pH 6-8 (optimum, pH 7) and 0-1.0 % NaCl (optimum, 0 %). The DNA G+C content of strain IMCC25901T was 34.2 mol%. The major fatty acid constituents of the strain were anteiso-C15 : 0, summed feature 3 (C16 : 1ω6c and/or C16 : 1ω7c) and iso-C15 : 0. Cells of strain IMCC25901T contained phosphatidylethanolamine, an unidentified phospholipid, two unidentified aminolipids and two unidentified lipids. The isoprenoid quinone detected in the strain was MK-6. On the basis of the taxonomic data obtained in this study, it was concluded that strain IMCC25901T represented a novel species in the genus Flavobacterium, for which the name Flavobacterium lacicola sp. nov. is proposed. The type strain of Flavobacterium lacicola is IMCC25901T (=KCTC 52571T=NBRC 112883T).

Deinococcus lacus sp. nov., a gamma radiation-resistant bacterium isolated from an artificial freshwater pond.

A Gram-stain-negative, pink-coloured, non-motile and gamma radiation-resistant bacterium, designated strain IMCC1711T, was isolated from a freshwater sample collected from an artificial pond (Inkyong Pond). The 16S rRNA gene sequence analysis showed that strain IMCC1711T was most closely related to Deinococcus piscis 3axT (94.2 %) and formed a robust phylogenetic clade with other species of the genus Deinococcus. Optimal growth of strain MCC1711T was observed at 25 °C and pH 7.0 without NaCl. Strain IMCC1711T exhibited high resistance to gamma radiation. The DNA G+C content of strain IMCC1711T was 59.1 mol% and MK-8 was the predominant isoprenoid quinone. Major fatty acid constituents of the strain were C17 : 1ω8c, C16 : 0, summed feature 3 (C16 : 1ω6c and/or C16 : 1ω7c) and C15 : 1ω6c. The major polar lipids constituted phosphatidylethanolamine, one unidentified phosphoglycolipid and two unidentified glycolipids. On the basis of taxonomic data obtained in this study, it was concluded that strain IMCC1711T represented a novel species of the genus Deinococcus, for which the name Deinococcus lacus sp. nov. is proposed. The type strain of Deinococcus lacus is IMCC1711T (KCTC 52494T=KACC 18979T=NBRC 112440T).

Microorganism-ionizing respirator with reduced breathing resistance suitable for removing airborne bacteria.

In this paper, we have demonstrated the feasibility of using microorganism-ionizing respirators with reduced breathing resistance to remove airborne bacteria. Using a miniaturized corona ionizer and two pairs of separator electrodes, airborne bacteria were ionized and removed from the airflow. Two microorganism-ionizing respirator designs were experimentally evaluated with flow rates ranging from ∼10 to 20 L/min and yielded airborne bacterial removal efficiencies of ∼75%-100%. Further, they were in close agreement with the analytical airborne particle removal efficiencies, at a similar range of flow rates. These flow rates also correspond to the breathing rates of standing and walking adults. More importantly, the breathing resistance could be reduced by more than 50% for flow rates of ∼200 L/min. Using manganese (IV) oxide coated mesh, the ozone concentration in the air outflow was reduced to less than 0.1 ppm, at a flow rate of ∼20 L/min, thus enabling safe use. The power consumption was less than 1 W.

Exopolysaccharide from Lactobacillus plantarum LRCC5310 offers protection against rotavirus-induced diarrhea and regulates inflammatory response.

We aimed to determine the effects of Lactobacillus strains against rotaviral infections. Rotaviruses are the major causative agent of acute gastroenteritis in infants and children worldwide. However, to date, no specific antiviral drugs for the treatment of rotavirus infection have been developed. We identified 263 Lactobacillus strains from 35 samples of the traditional Korean fermented vegetable food kimchi. Among them, Lactobacillus plantarum LRCC5310, more specifically the exopolysaccharides produced by these cells, were shown to have an antiviral effect against human rotavirus Wa strain in vitro. In vivo, the oral administration of exopolysaccharides for 2 d before and 5 d after mouse infection with the murine rotavirus epidemic diarrhea of infant mice strain led to a decrease in the duration of diarrhea and viral shedding and prevented the destruction of enteric epithelium integrity in the infected mice. We demonstrated here that the exopolysaccharides extracted from L. plantarum LRCC5310 can be used for the effective control of rotavirus infection.

Methionine-Restricted Diet Increases miRNAs That Can Target RUNX2 Expression and Alters Bone Structure in Young Mice.

Dietary methionine restriction (MR) increases longevity and improves healthspan in rodent models. Young male C57BL/6J mice were placed on MR to assess effects on bone structure and formation. Mice were fed diets containing 0.86% or 0.12% methionine for 5 weeks. Fasting blood plasma was analyzed for metabolic and bone-related biomarkers. Tibiae were analyzed by histomorphometry, while femurs were analyzed by micro-CT and biomechanically using 4-point bending. MR mice had reduced plasma glucose and insulin, while FGF21 and FGF23 increased. Plasma levels of osteocalcin and osteoprotegrin were unaffected, but sclerostin and procollagen I decreased. MR induced bone marrow fat accretion, antithetical to the reduced fat depots seen throughout the body. Cortical bone showed significant decreases in Bone Tissue Density (BTD). In trabecular bone, mice had decreased BTD, bone surface, trabecula and bone volume, and trabecular thickness.. Biomechanical testing showed that on MR, bones were significantly less stiff and had reduced maximum load and total work, suggesting greater fragility. Reduced expression of RUNX2 occurred in bone marrow of MR mice. These results suggest that MR alters bone remodeling and apposition. In MR mice, miR-31 in plasma and liver, and miR-133a, miR-335-5p, and miR-204 in the bone marrow was elevated. These miRNAs were shown previously to target and regulate Osterix and RUNX2 in bone, which could inhibit osteoblast differentiation and function. Therefore, dietary MR in young animals alters bone structure by increasing miRNAs in bone and liver that can target RUNX2. J. Cell. Biochem. 118: 31-42, 2017. © 2016 Wiley Periodicals, Inc.

Flavobacterium inkyongense sp. nov., isolated from an artificial freshwater pond.

A Gram-stain-negative, non-motile, yellow-pigmented bacterium, designated strain IMCC27201T, was isolated from an artificial freshwater pond (Inkyong) in Korea. Growth of strain IMCC27201T occurred at 10-37 °C (optimum, 30 °C), at pH 7.0-8.0 (optimum, pH 7.0) and with 0-0.25 % (w/v) NaCl (optimum, 0 %). Phylogenetic analysis based on 16S rRNA gene sequences indicated that strain IMCC27201T belonged to the genus Flavobacterium and was related most closely to Flavobacterium columnare IFO 15943T (97.8 % sequence similarity), Flavobacterium terrae R2A1-13T (97.2 %) and Flavobacterium vireti THG-SM1T (96.4 %). Strain IMCC27201T exhibited low DNA-DNA relatedness with F. columnare KCTC 42885T (34.9-40.6 %). The DNA G+C content of strain IMCC27201T was 32.1 mol%. The major fatty acids were iso-C15 : 0, iso-C15 : 1 G and iso-C15 : 0 3-OH. Strain IMCC27201T contained menaquinone-6 (MK-6) as the sole isoprenoid quinone. The polar lipids of strain IMCC27201T consisted of phosphatidylethanolamine, three unidentified aminolipids and two unidentified lipids. Based on 16S rRNA gene phylogeny, phenotypic characterization and DNA-DNA relatedness, strain IMCC27201Trepresents a novel species of the genus Flavobacterium, for which the name Flavobacterium inkyongense sp. nov. is proposed. The type strain is IMCC27201T (=KCTC 52244T=JCM 31385T).

Flavobacterium chuncheonense sp. nov. and Flavobacterium luteum sp. nov., isolated from a freshwater lake.

Two Gram-staining-negative, orange coloured, non-motile, rod-shaped bacterial strains, designated strains IMCC26013T and IMCC26026T, were isolated from a freshwater sample collected from Lake Soyang in Korea. The 16S rRNA gene-based phylogenetic analyses showed that both strains belonged to the genus Flavobacterium and that strains IMCC26013T and IMCC26026T were most closely related to Flavobacterium psychrophilum (96.5 %) and Flavobacterium myungsuense (97.7 %), respectively. DNA G+C contents of strains IMCC26013T and IMCC26026T were 37.8 and 33.7 mol%, respectively. DNA-DNA relatedness between strain IMCC26026T and F. myungsuense HMD1033T was 56.4 %, showing a novel species status of strain IMCC26026T. Major fatty acid constituents (>10 %) of strain IMCC26013T were iso-C15 : 1 G, C15 : 1 ω6c, C17 : 1 ω6c and summed feature 3 (C16 : 1 ω6c and/or C16 : 1 ω7c) and those of strain IMCC26026T were iso-C15 : 0, anteiso-C15 : 0 and summed feature 3. Respiratory quinone detected in the two strains was MK-6. Both strains contained phosphatidylethanolamine as a major polar lipid. On the basis of these results, strains IMCC26013T and IMCC26026T were considered to represent novel species in the genus Flavobacterium, for which the names Flavobacterium chuncheonense (type strain IMCC26013T=KCTC 52573T=NBRC 112526T), and Flavobacterium luteum (type strain IMCC26026T=KCTC 52572T=NBRC 112527T) are proposed, respectively.

Flavobacterium soyangense sp. nov., a psychrotolerant bacterium, isolated from an oligotrophic freshwater lake.

A bacterial strain, designated IMCC26223T, was isolated from an oligotrophic freshwater lake, Lake Soyang, Korea. Cells of strain IMCC26223T were Gram-staining negative, strictly aerobic, non-motile and short-rod-shaped. Growth occurred at pH 6-8 (optimum, pH 7.0), at 4-25 °C (optimum, 15 °C) and with 0-0.5 % (w/v) NaCl (optimum, 0 %). Phylogenetic analyses based on 16S rRNA gene sequences showed that strain IMCC26223T was identified as a member of the genus Flavobacterium and most closely related to Flavobacterium fluvii H7T (97.6 %), Flavobacterium segetis AT1048T (97.5 %) and Flavobacterium weaverense AT1042T (97.2 %). DNA-DNA relatedness between strain IMCC26223T and F. fluvii H7T was 41.5-51.7 % in the reciprocal hybridization. Strain IMCC26223T contained MK-6 as the major respiratory quinone. The major cellular fatty acids consisted of C16 : 0, iso-C15 : 0, anteiso-C15 : 0 and summed feature 3 (C16 : 1ω6c and/or C16 : 1ω7c), and the polar lipids were phosphatidylethanolamine, an unidentified aminolipid and two unidentified lipids. The DNA G+C content of strain IMCC26223T was 34.5 mol%. On the basis of 16S rRNA gene phylogeny and phenotypic characterization, strain IMCC26223T represents a novel species of the genus Flavobacterium, for which the name Flavobacterium soyangense sp. nov., is proposed. The type strain is IMCC26223T (=KCTC 52245T=JCM 31384T).

Micropatterned mammalian cells exhibit phenotype-specific left-right asymmetry.

Left-right (LR) asymmetry (handedness, chirality) is a well-conserved biological property of critical importance to normal development. Changes in orientation of the LR axis due to genetic or environmental factors can lead to malformations and disease. While the LR asymmetry of organs and whole organisms has been extensively studied, little is known about the LR asymmetry at cellular and multicellular levels. Here we show that the cultivation of cell populations on micropatterns with defined boundaries reveals intrinsic cell chirality that can be readily determined by image analysis of cell alignment and directional motion. By patterning 11 different types of cells on ring-shaped micropatterns of various sizes, we found that each cell type exhibited definite LR asymmetry (p value down to 10(-185)) that was different between normal and cancer cells of the same type, and not dependent on surface chemistry, protein coating, or the orientation of the gravitational field. Interestingly, drugs interfering with actin but not microtubule function reversed the LR asymmetry in some cell types. Our results show that micropatterned cell populations exhibit phenotype-specific LR asymmetry that is dependent on the functionality of the actin cytoskeleton. We propose that micropatterning could potentially be used as an effective in vitro tool to study the initiation of LR asymmetry in cell populations, to diagnose disease, and to study factors involved with birth defects in laterality.

Ameliorating effects of Orostachys japonica against high-fat diet-induced obesity and gut dysbiosis.

ETHNOPHARMACOLOGICAL RELEVANCE: Orostachys japonica (rock pine) has been used as a folk remedy to treat inflammation, hepatitis, and cancer in East Asia. AIM OF THE STUDY: The aim of this study was to investigate the effect of rock pine extract (RPE) on high-fat diet-induced obesity in mice and to examine its effects on gut dysbiosis. MATERIALS AND METHODS: The characteristic compound of RPE, kaempferol-3-O-rutinoside, was quantified using high-performance liquid chromatography. The prebiotic potential of RPE was evaluated by assessing the prebiotic activity score obtained using four prebiotic strains and high-fat (HF)-induced obesity C57BL/6 mice model. Analysis included examining the lipid metabolism and inflammatory proteins and evaluating the changes in gut permeability and metabolites to elucidate the potential signaling pathways involved. RESULTS: In vitro, RPE enhanced the proliferation of beneficial probiotic strains, including Lactiplantibacillus and Bifidobacterium. HF-induced model showed that the administration of 100 mg/kg/day of RPE for 8 weeks significantly (p < 0.05) reduced the body weight, serum lipid levels, and insulin resistance, which were associated with notable changes in lipid metabolism and inflammation-related markers. CONCLUSIONS: Our results demonstrate that rock pine consumption could mitigate obesity and metabolic endotoxemia in HF-fed mice through enhancing intestinal environment.