Reconstruction of Long-Chain Polyunsaturated Acid Synthesis Pathways in Marine Red Microalga Porphyridium cruentum Using Lipidomics and Transcriptomics.

The marine red microalga Porphyridium can simultaneously synthesize long-chain polyunsaturated fatty acids, including eicosapentaenoic acid (C20:5, EPA) and arachidonic acid (C20:4, ARA). However, the distribution and synthesis pathways of EPA and ARA in Porphyridium are not clearly understood. In this study, Porphyridium cruentum CCALA 415 was cultured in nitrogen-replete and nitrogen-limited conditions. Fatty acid content determination, transcriptomic, and lipidomic analyses were used to investigate the synthesis of ARA and EPA. The results show that membrane lipids were the main components of lipids, while storage lipids were present in a small proportion in CCALA 415. Nitrogen limitation enhanced the synthesis of storage lipids and ω6 fatty acids while inhibiting the synthesis of membrane lipids and ω3 fatty acids. A total of 217 glycerolipid molecular species were identified, and the most abundant species included monogalactosyldiglyceride (C16:0/C20:5) (MGDG) and phosphatidylcholine (C16:0/C20:4) (PC). ARA was mainly distributed in PC, and EPA was mainly distributed in MGDG. Among all the fatty acid desaturases (FADs), the expressions of Δ5FAD, Δ6FAD, Δ9FAD, and Δ12FAD were up-regulated, whereas those of Δ15FAD and Δ17FAD were down-regulated. Based on these results, only a small proportion of EPA was synthesized through the ω3 pathway, while the majority of EPA was synthesized through the ω6 pathway. ARA synthesized in the ER was likely shuttled into the chloroplast by DAG and was converted into EPA by Δ17FAD.

The inhibition of enterovirus 71 induced apoptosis by Durvillaea antarctica through P53 and STAT1 signaling pathway.

The infection of enterovirus 71 (EV71) resulted in hand, foot, and mouth disease and may lead to severe nervous system damage and even fatalities. There are no effective drugs to treat the EV71 virus and it is crucial to find novel drugs against it. Polysaccharide isolated from Durvillaea antarctica green algae has an antiviral effect. In this study, D. antarctica polysaccharide (DAPP) inhibited the infection of EV71 was demonstrated by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT), reverse transcription polymerase chain reaction, flow cytometry, and western blot. MTT assay showed that DAPP had no toxicity on Vero cells at the concentration 250 µg/ml. Furthermore, DAPP significantly reduced the RNA level of EV71 in a dose-dependent manner. Moreover, DAPP inhibited the Vero cells apoptosis induced by EV71 via the P53 signaling pathway. Meanwhile, the expression of signal transducer and activator of transcription 1 and mammalian target of rapamycin were increased and the proinflammatory cytokines were significantly inhibited by DAPP. Taken together, these results suggested that DAPP could be a potential pharmaceutical against the infection of EV71 virus.

Inhibition of enterovirus 71 infection by polysaccharides extracted from Picochlorum sp. 122 via the AKT and ATM/ATR signaling pathways.

Enterovirus 71 (EV71) poses a major threat to public health globally due to severe and even fatal hand, foot, and mouth disease (HFMD). However, no effective antiviral agents are available to treat HFMD caused by this virus. Polysaccharides have been shown to exhibit antiviral activity, and polysaccharides extracted from Picochlorum sp. 122 (PPE) could potentially be used to treat HFMD, but reports on their antiviral activity are limited. In this study, the antiviral activity of PPE against EV71 was verified in Vero cells. PPE was shown to limit EV71 infection, as demonstrated using an MTT assay and by observing the cellular cytopathic effect. In addition, a decrease in VP1 RNA and protein levels indicated that PPE effectively inhibits proliferation of EV71 in Vero cells. An annexin V affinity assay also indicated that PPE protects host cells from apoptosis through the AKT and ATM/ATR signalling pathways. These results demonstrate that PPE has potential as an antiviral drug to treat HFMD caused by EV71.

Growth and Biochemical Composition of Porphyridium purpureum SCS-02 under Different Nitrogen Concentrations.

Microalgae of the genus Porphyridium show great potential for large-scale commercial cultivation, as they accumulate large quantities of B-phycoerythrin (B-PE), long chain polyunsaturated fatty acids (LC-PUFAs) and exopolysaccharide (EPS). The present study aimed to adjust culture nitrogen concentrations to produce Porphyridium biomass rich in B-PE, LC-PUFAs and EPS. Porphyridium purpureum SCS-02 was cultured in ASW culture medium with low nitrogen supply (LN, 3.5 mM), medium nitrogen supply (MN, 5.9 mM) or high nitrogen supply (HN, 17.6 mM). HN significantly enhanced the accumulation of biomass, intracellular protein, B-PE and eicosapentaenoic acid. LN increased the intracellular carbohydrate and arachidonic acid content, and promoted the secretion of EPS. The total lipids content was almost unaffected by nitrogen concentration. Based on these results, a semi-continuous two-step process was proposed, which included the production of biomass rich in B-PE and LC-PUFAs with sufficient nitrogen, and induced EPS excretion with limited nitrogen and strong light.

Pelagibacterium lentulum sp. nov., a marine bacterium from the culture broth of Picochlorum sp. 122.

A Gram-stain-negative, motile, non-spore-forming, rod-shaped bacterium, designated strain B2T, was isolated from the culture broth of a marine microalga, Picochlorum sp. 122. Phylogenetic analyses based on 16S rRNA gene sequences indicated that strain B2T forms a robust cluster with members of the genus Pelagibacterium, and shares the highest sequence similarity of 96.80 % with Pelagibacterium halotolerans CGMCC 1.7692T. Optimal growth of strain B2T was observed at 33 °C, at pH 8 and in the presence of 1 % (w/v) NaCl. The predominant ubiquinone of strain B2T was Q-10, and the G+C content of the genomic DNA was 58.6 mol%. The major fatty acid profile comprised C18 : 1ω7c/ω6c, C19 : 0 cyclo ω8c and C16 : 0. The major polar lipids of strain B2T were diphosphatidylglycerol, phosphatidylglycerol, two unidentified glycolipids and seven unidentified lipids. Comprehensive analyses based on a polyphasic characterization of strain B2T indicated that it represents a novel species of the genus Pelagibacterium, for which the name Pelagibacterium lentulum sp. nov. is proposed. The type strain is B2T (=MCCC 1K03218T=CGMCC 1.15896T=KCTC 52551T).

Butyratibacter algicola gen. nov., sp. nov., a marine bacterium from the culture broth of Picochlorum sp. 122.

A Gram-stain-negative, motile, non-spore-forming, ovoid or rod-shaped bacterium, designated strain B15T, was isolated from the culture broth of a marine microalga, Picochlorum sp. 122. Phylogenetic analyses based on 16S rRNA gene sequences indicated that strain B15T forms a stable cluster with Lutibaculum baratangense KCTC 22669T (95.4 % 16S rRNA gene sequence similarity), Tepidamorphus gemmatus CB-27AT (94.9 %) and Microbaculum marinum HSF11T (94.6 %) in the family Rhodobiaceae. Optimal growth of strain B15T was observed at 33 °C, pH 8-9 and in the presence of 3 % (w/v) NaCl. The only detected ubiquinone of strain B15T was Q-10, and the G+C content of the genomic DNA was 66.3 mol%. The major fatty acid profile comprised C19 : 0 cyclo ω8c, C18 : 1ω7c/ω6c and C17 : 1 iso I/anteiso B. The major polar lipids of strain B15T were diphosphatidylglycerol, phosphatidylglycerol, phosphatidylcholine, phosphatidylethanolamine, phosphatidylmonomethylethanolamine, two unidentified aminolipids, and one unidentified lipid. Comprehensive analyses based on polyphasic characterization of strain B15T indicated that it represents a novel species of a new genusin the family Rhodobiaceae, for which the name Butyratibacter algicola gen. nov., sp. nov. is proposed. The type strain of the type species is B15T (=KCTC 52552T=MCCC 1K03221T).

Gelatiniphilus marinus gen. nov., sp. nov., a bacterium from the culture broth of a microalga, Picochlorum sp. 122, and emended description of the genus Hwangdonia.

A Gram-stain-negative, non-motile, non-spore-forming, rod-shaped bacterium, designated strain GYP-24T, was isolated from the culture broth of a marine microalga, Picochlorum sp. 122. Phylogenetic analyses based on 16S rRNA gene sequences indicated that strain GYP-24T forms a robust cluster with H.wangdoniaseohaensis KCTC 32177T (95.8 % sequence similarity) in the family Flavobacteriaceae. Growth of strain GYP-24T was observed at 15, 22, 28, 30, 33 and 37 °C (optimal 30-33 °C), pH 6.0-10.0 (optimal pH 7.0-8.0) and in the presence of 0.5-4 % (w/v) NaCl (optimal 2-3 %). The only menaquinone of strain GYP-24T was MK-6, and the G+C content of the genomic DNA was 36.9 mol%. The major fatty acid profile comprised iso-C17 : 0 3-OH, summed feature 3 (C16 : 1 ω7c/ω6c), iso-C15 : 1 G and iso-C15 : 0. The major polar lipids of strain GYP-24T were phosphatidylethanolamine, one unidentified phospholipid, three unidentified aminolipids and three unidentified lipids. Comprehensive analyses based on polyphasic characterization of GYP-24T indicated that it represents a novel species of a new genus, for which the name Gelatiniphilus marinus gen. nov., sp. nov. is proposed. The type strain is GYP-24T (=KCTC 42903T=MCCC 1K01730T). An emended description of the genus Hwangdonia is also given.

Pyruvatibacter mobilis gen. nov., sp. nov., a marine bacterium from the culture broth of Picochlorum sp. 122.

A Gram-stain-negative, aerobic bacterium, designated strain GYP-11T, was isolated from the culture broth of a marine microalga, Picochloruma sp. 122. Cells were dimorphic rods; free living cells were motile by means of a single polar flagellum, and star-shaped-aggregate-forming cells were attached with stalks and non-motile. Sodium pyruvate or Tween 20 was required for growth on marine agar 2216.16S rRNA gene sequence analysis revealed that this isolate shared 94.07 % similarity with its closest type strain, Parvibaculum hydrocarboniclasticum EPR92T. Phylogenetic analyses indicated that strain GYP-11T represents a distinct lineage in a robust clade consisting of strain GYP-11T, alphaproteobacterium GMD21A06 and Candidatus Phaeomarinobacter ectocarpi Ec32. This clade was close to the genera Parvibaculum and Tepidicaulis in the order Rhizobiales. Chemotaxonomic and physiological characteristics, including cellular fatty acids and carbon source profiles, also readily distinguished strain GYP-11T from all established genera and species. Thus, it is concluded that strain GYP-11T represents a novel species of a new genus in the order Rhizobiales, for which the name Pyruvatibacter mobilis gen. nov., sp. nov. is proposed. The type strain of Pyruvatibacter mobilis is GYP-11T ( = CGMCC 1.15125T = KCTC 42509T).

A Saponification Method for Chlorophyll Removal from Microalgae Biomass as Oil Feedstock.

Microalgae oil is an optimal feedstock for nutraceutical, pharmaceutical and biodiesel production, but its high levels of chlorophyll limit its large-scale application. To date, few effective approaches have been developed to remove chlorophyll from microalgae oil. The main purpose of this study was to present a preprocessing method of algae oil feedstock (Scenedesmus) to remove chlorophyll by saponification. The results showed that 96% of chlorophyll in biomass was removed. High quality orange transparent oil could be extracted from the chlorophyll reduced biomass. Specifically, the proportion of neutral lipids and saturation levels of fatty acids increased, and the pigments composition became carotenoids-based. The critical parameters of chlorophyll reduced biodiesel conformed to the standards of the USA, China and EU. Sodium copper chlorophyllin could be prepared from the bleaching effluent. The results presented herein offer a useful pathway to improve the quality of microalgae oil and reduce the cost of microalgae biodiesel.

Maribacter flavus sp. nov., isolated from a cyanobacterial culture pond.

A Gram-stain-negative, yellow, non-spore-forming, strictly aerobic bacterium, designated C3T, was isolated from a cyanobacterial culture pond. Cells were halophilic, rod-shaped and able to move by gliding. Growth of strain C3T was observed at 15-30 °C (optimum 25 °C), pH 6.0-9.0 (optimum pH 7.5), and in the presence of 1-7 % (w/v) NaCl (optimum 2-3 %). Phylogenetic analysis based on 16S rRNA gene sequences revealed that strain C3T formed a distinct lineage within the family Flavobacteriaceae and exhibited the highest similarity (95.21 %) to the type strains of Maribacter dokdonensis, Maribacter arcticus, Maribacter orientalis and Maribacter stanieri, and 'Maribacter caenipelagi' HD-44. The only isoprenoid quinone present within strain C3T was menaquinone 6 (MK-6). The G+C content of genomic DNA was 41.5 mol%. The major polar lipids were phosphatidylethanolamine and three unidentified lipids. The predominant cellular fatty acids (>5 % of the total fatty acids) were iso-C15 : 1 G, iso-C15 : 0, iso-C15 : 0 3-OH, iso-C17 : 0 3-OH, and summed feature 3 (comprising C16 : 1ω7c and/or C16 : 1ω6c). On the basis of the phenotypic, chemotaxonomic and phylogenetic characteristics, strain C3T represents a novel species of the genus Maribacter, for which the name Maribacter flavus sp. nov. is proposed. The type strain is C3T ( = KCTC 42508T = CGMCC 1.15112T).

Aliikangiella marina gen. nov., sp. nov., a marine bacterium from the culture broth of Picochlorum sp. 122, and proposal of Kangiellaceae fam. nov. in the order Oceanospirillales.

A Gram-stain-negative, non-motile, non-spore-forming, long rod-shaped bacterium, designated strain GYP-15T, was isolated from the culture broth of a marine microalga, Picochloruma sp. 122. Phylogenetic analyses revealed that strain GYP-15T shared 90.6 % 16S rRNA gene sequence similarity with its closest relative, Kangiella aquimarina KCTC 12183T, and represents a distinct phylogenetic lineage in a robust clade consisting of GYP-15T and members of the genera Kangiella and Pleionea in the order Oceanospirillales. Chemotaxonomic and physiological characteristics, including major cellular fatty acids, NaCl tolerance and pattern of carbon source utilization, could also readily distinguish strain GYP-15T from all established genera and species. Thus, it is concluded that strain GYP-15T represents a novel species of a new genus, for which the name Aliikangiella marina gen. nov., sp. nov. is proposed. The type strain of Aliikangiella marina is GYP-15T ( = MCCC 1K01163T = KCTC 42667T). Based on phylogenetic results, 16S rRNA gene signature nucleotide pattern and some physiological characteristics, the three genera Kangiella, Pleionea and Aliikangiella are proposed to make up a novel family, Kangiellaceae fam. nov., in the order Oceanospirillales.

Oceanobacillus bengalensis sp. nov., a bacterium isolated from seawater of the Bay of Bengal.

A Gram-stain positive, motile, and subterminal endospore-forming rod-shaped bacterium, designated strain Ma-21(T), was isolated from seawater of the Bay of Bengal. Strain Ma-21(T) was found to grow optimally at 37 °C and pH 8.0 with 3% (w/v) NaCl. Phylogenetic analyses showed that strain Ma-21(T) forms a distinct phylogenetic lineage close to Oceanobacillus chungangensis CAU 1051(T), Oceanobacillus caeni S-11(T), Oceanobacillus arenosus CAU 1183(T), Oceanobacillus halophilum GD01(T) and Ornithinibacillus heyuanensis GIESS003(T) in the family Bacillaceae. The cell wall of strain Ma-21(T) was found to contain meso-diaminopimelic acid as the diagnostic diamino acid, which is in line with those of members of the genus Oceanobacillus. The genomic DNA G+C content was determined to be 35.9 mol%. The only respiratory quinone detected was MK-7. The major cellular fatty acids were identified as anteiso-C(15:0) and anteiso-C(17:0). The major polar lipids were found to be diphosphatidylglycerol and phosphatidylglycerol. On the basis of phylogenetic, chemotaxonomic and phenotypic properties, strain Ma-21(T) is suggested to represent a novel species in the genus Oceanobacillus, for which the name Oceanobacillus bengalensis sp. nov. is proposed. The type strain is Ma-21(T) (=CGMCC 1.12799(T) = KCTC 33416(T) = MCCC 1K00260(T)).

Wenzhouxiangella marina gen. nov, sp. nov, a marine bacterium from the culture broth of Picochlorum sp. 122, and proposal of Wenzhouxiangellaceae fam. nov. in the order Chromatiales.

A Gram-stain negative, non-motile, non-phototrophic, non-alkaliphilic, obligately aerobic, chemoheterotrophic, and rod-shaped bacterium, designated strain Ma-11(T), was isolated from the culture broth of a marine microalga, Picochloruma sp. 122. Phylogenetic analyses showed that strain Ma-11(T) has less than 91 % similarity to its closest relative, Thioalkalivibrio sulfidiphilus HL-EbGR7(T), represents a distinct phylogenetic lineage in the order Chromatiales, and could not be assigned to any defined families in this order. Chemotaxonomic, genetic and physiological characteristics, including major fatty acids, genomic G+C content, lack of motility, aerophilicity and chemoheterotrophicity, could readily distinguish strain Ma-11(T) from any established members of the order Chromatiales. Based on the 16S rRNA gene sequence analysis and its signature nucleotide pattern, a new family Wenzhouxiangellaceae fam. nov. comprising the genus Wenzhouxiangella gen. nov. and species Wenzhouxiangella marina sp. nov. is proposed. The type strain is Ma-11(T) (=CGMCC 1.14936(T) = KCTC 42284(T) = MCCC 1K00261(T)).

Physicochemical and crystalline properties of heat-moisture-treated rice starch: combined effects of moisture and duration of heating.

BACKGROUND: Currently there is much interest in the application of physical modification techniques such as heat-moisture treatment (HMT). The effects of HMT on normal and waxy rice starches, subject to different levels of moisture content and duration of heating, were investigated. RESULTS: Water solubility index (determined at 90 °C) decreased after HMT for normal and waxy rice starches, while swelling power (determined at 90 °C) showed inconsistent results (decrease for normal type, increase for waxy type) after HMT. Values in pasting parameters of normal and waxy rice starch increased initially, but the extent of increase slowed down with moisture content and length of treatment increasing. HMT decreased gelatinization temperatures with 4 h and 8 h treatment, but when length of treatment was prolonged to 16 h gelatinization temperature increased. Degree of crystallinity decreased for all treatments, and decreased much more at higher levels of moisture content. CONCLUSION: Variations in levels of moisture content and duration of heating had significant effects on physicochemical and crystalline properties to different extents.

Bacterioplanes sanyensis gen. nov., sp. nov., a PHB-accumulating bacterium isolated from a pool of Spirulina platensis cultivation.

A Gram-negative, poly-3-hydroxybutyrate-accumulating rod bacterium, strain GYP-2(T), was isolated from a pool of marine Spirulina platensis cultivation, Sanya, China. Growth was observed at 10-45 °C and pH 6-10 in the presence of 1-10 % (w/v) NaCl. Phylogenetic analysis based on 16S rRNA gene sequences indicated that the new isolate belonged to Gammaproteobacteria and displayed 93.8-95.3 % 16S rRNA gene sequences similarities to members of the genera Thalassolituus, Oleibacter, and Oceanobacter, but house-keeping gene gyrB (encode DNA gyrase beta subunit) demonstrated that the new isolate was distantly related to Thalassolituus, Oleibacter, and Oceanobacter species (only 77-83 % gene gyrB sequences similarities).The G+C content of genomic DNA was 55 mol%. The major respiratory quinone was Q-9, while that for Oceanobacter kriegii LMG 6238(T) was Q-8. Major polar lipids were diphosphatidylglycerol, phosphatidylglycerol, and phosphatidylethanolamine. On the basis of its physiological, chemotaxonomic, and molecular properties, strain GYP-2(T) is suggested to represent a novel species of a new genus in Gammaproteobacteria, for which the name Bacterioplanes sanyensis gen. nov., sp. nov. is proposed. The type strain is GYP-2(T) (=CGMCC 1.12392(T)=KCTC 32220(T)).

Optimization of medium using response surface methodology for lipid production by Scenedesmus sp.

Lipid production is an important indicator for assessing microalgal species for biodiesel production. In this work, the effects of medium composition on lipid production by Scenedesmus sp. were investigated using the response surface methodology. The results of a Plackett-Burman design experiment revealed that NaHCO3, NaH2PO4·2H2O and NaNO3 were three factors significantly influencing lipid production, which were further optimized by a Box-Behnken design. The optimal medium was found to contain 3.07 g L⁻¹ NaHCO3, 15.49 mg L⁻¹ NaH2PO4·2H2O and 803.21 mg L⁻¹ NaNO3. Using the optimal conditions previously determined, the lipid production (304.02 mg·L⁻¹) increased 54.64% more than that using the initial medium, which agreed well with the predicted value 309.50 mg L⁻¹. Additionally, lipid analysis found that palmitic acid (C16:0) and oleic acid (C18:1) dominantly constituted the algal fatty acids (about 60% of the total fatty acids) and a much higher content of neutral lipid accounted for 82.32% of total lipids, which strongly proved that Scenedesmus sp. is a very promising feedstock for biodiesel production.

A novel lipid extraction method from wet microalga Picochlorum sp. at room temperature.

A novel method using ethanol was proposed for extracting lipids from wet microalga Picochlorum sp. at room temperature and pressure. In this study, Central Composite design (CCD) was applied to investigate the optimum conditions of lipid extraction. The results revealed that the solvent to biomass ratio had the largest effect on lipid extraction efficiency, followed by extraction time and temperature. A high lipid extraction yield (33.04% of the dry weight) was obtained under the following extraction conditions: 5 mL solvents per gram of wet biomass for 37 min with gentle stirring at room temperature. The extraction yield was comparable to that obtained by the widely used Bligh-Dyer method. Furthermore, no significant differences in the distribution of lipid classes and fatty acid composition were observed according to different extraction methods. In conclusion, these results indicated that the proposed procedure using ethanol could extract lipids from wet biomass efficiently and had giant potential for lipid extraction at large scale.

Multi-Omics Reveals the Effects of Spirulina platensis Powder Replacement of Fish Meal on Intestinal Metabolism and Stress in Zig-Zag Eel (Mastacembelus armatus).

The booming aquaculture industry has created a strong demand for fishmeal and increased environmental pressures. Spirulina, as a potential alternative to fishmeal, has been shown to have growth-promoting and animal health-enhancing properties. In this study, 600 large spiny loaches, divided into five experimental groups, F0, F1, F2, F3, and F4, were reared for 10 weeks using Spirulina platensis powder (SPP) as a substitute for 0%, 5%, 10%, 15%, and 20% of fishmeal, respectively. The results of intestinal physiological indexes showed that superoxide dismutase was lower than F0 in all treatment groups, and the activity of F3 was significantly lower than F0 (p < 0.05). The activity of malondialdehyde was significantly higher than that of F0 in all groups except F3 (p < 0.05). The addition of SPP also led to a decrease in the activity of acid phosphatase in the intestine, which was significantly lower in all treatment groups compared to the F0 group (p < 0.05). The results of serum physiology showed that the activity of superoxide dismutase in serum gradually increased with the increase in the percentage of SPP addition, and the F3 group produced a significant difference from the F0 group (p < 0.05). The transcriptomics results showed that DEGs in the low percentage substitution group (<15%) were mostly enriched in metabolism-related pathways, such as bile secretion; DEGs in the high percentage substitution group (>15%) were mostly enriched in inflammation-related pathways, such as complement p and coagulation cascades. Metabolomics confirmed that nicotinate and nicotinamide metabolism and glycerophospholipid metabolism were the two pathways that were significantly enriched in the treatment groups of fishmeal replacement by SPP. The present study demonstrated that a low percentage (<15%) of fishmeal replacement by SPP in feed mobilized MA digestive metabolism, whereas a high percentage (>15%) of replacement induced intestinal stress. Considering the health and farm efficiency aspects, the proportion of SPP in feed formulation for MA should be less than 15%.

Pustulibacterium marinum gen. nov., sp. nov., a member of the family Flavobacteriaceae isolated from the Bashi Channel.

A Gram-reaction-negative, non-spore-forming, gliding, non-translucent, colourless or yellow, aerobic and elevated-colony-forming strain, designated E403(T), was isolated from the Bashi Channel and subjected to a polyphasic taxonomic study. Strain E403(T) could grow in the presence of 0.3-8 % (w/v) NaCl, at 16-43 °C and at pH 6-9, and grew optimally at 28 °C, pH 8, in natural seawater medium. The respiratory quinones were MK-6 and MK-7. The major fatty acids were iso-C15 : 0, iso-C17 : 0 3-OH, iso-C15 : 1 G, summed feature 3 (C16 : 1ω7c/C16 : 1ω6c), iso-C15 : 0 3-OH and C16 : 0. The DNA G+C content of strain E403(T) was 37.9 mol%. Phylogenetic analyses based on 16S rRNA gene sequences of members of the family Flavobacteriaceae showed that strain E403(T) formed a distinct evolutionary lineage within the stable cluster containing type strains Zhouia amylolytica HN-171(T) (92.2 % similarity) and Joostella marina En5(T) (92.4 % similarity). In addition to the large 16S rRNA gene sequence differences, E403(T) can also be distinguished from the reference type strains J. marina En5(T) and Sinomicrobium oceani SCSIO 03483(T) by several phenotypic characteristics and chemotaxonomic properties. On the basis of phenotypic, chemotaxonomic and phylogenetic properties, strain E403(T) is suggested to represent a novel species of a new genus in the family Flavobacteriaceae, for which the name Pustulibacterium marinum gen. nov., sp. nov. is proposed. The type strain is E403(T) (= CCTCC AB2012862(T) = CGMCC 1.12333(T) = KCTC 32192(T)).

Nonhongiella spirulinensis gen. nov., sp. nov., a bacterium isolated from a cultivation pond of Spirulina platensis in Sanya, China.

A Gram-negative, aerobic, motile rod strain, designated Ma-20(T), was isolated from a pool of marine Spirulina platensis cultivation, Sanya, China, and was subjected to a polyphasic taxonomy study. Strain Ma-20(T) can grow in the presence of 0.5-11 % (w/v) NaCl, 10-43 °C and pH 6-10, and grew optimally at 30 °C, pH 7.5-9.0 in natural seawater medium. The polar lipids were composed of phosphatidylethanolamine, three unidentified phospholipids and three unidentified polar lipids. The respiratory quinone was ubiquinone 8 (Q-8) and the major fatty acids were C18:1ω6c/C18:1ω7c (summed feature 8, 32.84 %), C16:1ω6c/C16:1ω7c (summed feature 3, 30.76 %), C16:0 (13.54 %), C12:03-OH (4.63 %), and C12:0 (4.09 %). The DNA G+C content of strain Ma-20(T) was 58 mol %. Phylogenetic analyses based on 16S rRNA gene sequences showed that strain Ma-20(T) belonging to Gammaproteobacteria, it shared 88.46-91.55 and 89.21-91.26 % 16S rRNA gene sequence similarity to the type strains in genus Hahella and Marinobacter, respectively. In addition to the large 16S rRNA gene sequence difference, Ma-20(T) can also be distinguished from the reference type strains Hahella ganghwensis FR1050(T) and Marinobacter hydrocarbonoclasticus sp. 17(T) by several phenotypic characteristics and chemotaxonomic properties. On the basis of phenotypic, chemotaxonomic and phylogenetic properties, strain Ma-20(T) is suggested to represent a novel species of a new genus in Gammaproteobacteria, for which the name Nonhongiella spirulinensis gen. nov., sp. nov. is proposed. The type strain is Ma-20(T) (=KCTC 32221(T)=LMG 27470(T)).