Increasing outer membrane complexity: the case of the lipopolysaccharide lipid A from marine Cellulophaga pacifica.

Gram-negative bacteria living in marine waters have evolved peculiar adaptation strategies to deal with the numerous stress conditions that characterize aquatic environments. Among the multiple mechanisms for efficient adaptation, these bacteria typically exhibit chemical modifications in the structure of the lipopolysaccharide (LPS), which is a fundamental component of their outer membrane. In particular, the glycolipid anchor to the membrane of marine bacteria LPSs, i.e. the lipid A, frequently shows unusual chemical structures, which are reflected in equally singular immunological properties with potential applications as immune adjuvants or anti-sepsis drugs. In this work, we determined the chemical structure of the lipid A from Cellulophaga pacifica KMM 3664T isolated from the Sea of Japan. This bacterium showed to produce a heterogeneous mixture of lipid A molecules that mainly display five acyl chains and carry a single phosphate and a D-mannose disaccharide on the glucosamine backbone. Furthermore, we proved that C. pacifica KMM 3664T LPS acts as a weaker activator of Toll-like receptor 4 (TLR4) compared to the prototypical enterobacterial Salmonella typhimurium LPS. Our results are relevant to the future development of novel vaccine adjuvants and immunomodulators inspired by marine LPS chemistry.

Three novel marine species of the genus Reichenbachiella exhibiting degradation of complex polysaccharides.

Three novel strains designated ABR2-5T, BKB1-1T, and WSW4-B4T belonging to the genus Reichenbachiella of the phylum Bacteroidota were isolated from algae and mud samples collected in the West Sea, Korea. All three strains were enriched for genes encoding up to 216 carbohydrate-active enzymes (CAZymes), which participate in the degradation of agar, alginate, carrageenan, laminarin, and starch. The 16S rRNA sequence similarities among the three novel isolates were 94.0%-94.7%, and against all three existing species in the genus Reichenbachiella they were 93.6%-97.2%. The genome sizes of the strains ABR2-5T, BKB1-1T, and WSW4-B4T were 5.5, 4.4, and 5.0 Mb, respectively, and the GC content ranged from 41.1%-42.0%. The average nucleotide identity and the digital DNA-DNA hybridization values of each novel strain within the isolates and all existing species in the genus Reichenbachiella were in a range of 69.2%-75.5% and 17.7-18.9%, respectively, supporting the creation of three new species. The three novel strains exhibited a distinctive fatty acid profile characterized by elevated levels of iso-C15:0 (37.7%-47.4%) and C16:1 ω5c (14.4%-22.9%). Specifically, strain ABR2-5T displayed an additional higher proportion of C16:0 (13.0%). The polar lipids were phosphatidylethanolamine, unidentified lipids, aminolipids, and glycolipids. Menaquinone-7 was identified as the respiratory quinone of the isolates. A comparative genome analysis was performed using the KEGG, RAST, antiSMASH, CRISPRCasFinder, dbCAN, and dbCAN-PUL servers and CRISPRcasIdentifier software. The results revealed that the isolates harbored many key genes involved in central metabolism for the synthesis of essential amino acids and vitamins, hydrolytic enzymes, carotenoid pigments, and antimicrobial compounds. The KEGG analysis showed that the three isolates possessed a complete pathway of dissimilatory nitrate reduction to ammonium (DNRA), which is involved in the conservation of bioavailable nitrogen within the ecosystem. Moreover, all the strains possessed genes that participated in the metabolism of heavy metals, including arsenic, copper, cobalt, ferrous, and manganese. All three isolated strains contain the class 2 type II subtype C1 CRISPR-Cas system in their genomes. The distinguished phenotypic, chemotaxonomic, and genomic characteristics led us to propose that the three strains represent three novel species in the genus Reichenbachiella: R. ulvae sp. nov. (ABR2-5T = KCTC 82990T = JCM 35839T), R. agarivorans sp. nov. (BKB1-1T = KCTC 82964T = JCM 35840T), and R. carrageenanivorans sp. nov. (WSW4-B4T = KCTC 82706T = JCM 35841T).

Pushing the Boundaries of Structural Heterogeneity with the Lipid A of Marine Bacteria Cellulophaga.

Marine bacteria, which are often described as chemical gold, are considered an exceptional source of new therapeutics. Considerable research interest has been given to lipopolysaccharides (LPSs), the main components of the Gram-negative outer membrane. LPS and its lipid A portion from marine bacteria are known to exhibit a tricky chemistry that has been often associated with intriguing properties such as behaving as immune adjuvants or anti-sepsis molecules. In this scenario, we report the structural determination of the lipid A from three marine bacteria within the Cellulophaga genus, which showed to produce an extremely heterogenous blend of tetra- to hexa-acylated lipid A species, mostly carrying one phosphate and one D-mannose on the glucosamine disaccharide backbone. The ability of the three LPSs in activating TLR4 signaling revealed a weaker immunopotential by C. baltica NNO 15840T and C. tyrosinoxydans EM41T , while C. algicola ACAM 630T behaved as a more potent TLR4 activator.

Three marine species of the genus Fulvivirga, rich sources of carbohydrate-active enzymes degrading alginate, chitin, laminarin, starch, and xylan.

Bacteroidota is a group of marine polysaccharide degraders, which play a crucial role in the carbon cycle in the marine ecosystems. In this study, three novel gliding strains, designated as SS9-22T, W9P-11T, and SW1-E11T, isolated from algae and decaying wood were proposed to represent three novel species of the genus Fulvivirga. We identified a large number of genes encoding for carbohydrate-active enzymes, which potentially participate in polysaccharide degradation, based on whole genome sequencing. The 16S rRNA sequence similarities among them were 94.4-97.2%, and against existing species in the genus Fulvivirga 93.1-99.8%. The complete genomes of strains SS9-22T, W9P-11T, and SW1-E11T comprised one circular chromosome with size of 6.98, 6.52, and 6.39 Mb, respectively; the GC contents were 41.9%, 39.0%, and 38.1%, respectively. The average nucleotide identity and the digital DNA-DNA hybridization values with members in the genus Fulvivirga including the isolates were in a range of 68.9-85.4% and 17.1-29.7%, respectively, which are low for the proposal of novel species. Genomic mining in three genomes identified hundreds of carbohydrate-active enzymes (CAZymes) covering up to 93 CAZyme families and 58-70 CAZyme gene clusters, exceeding the numbers of genes present in the other species of the genus Fulvivirga. Polysaccharides of alginate, chitin, laminarin, starch, and xylan were degraded in vitro, highlighting that the three strains are rich sources of CAZymes of polysaccharide degraders for biotechnological applications. The phenotypic, biochemical, chemotaxonomic, and genomic characteristics supported the proposal of three novel species in the genus Fulvivirga, for which the names Fulvivirga ulvae sp. nov. (SS9-22T = KCTC 82072T = GDMCC 1.2804T), Fulvivirga ligni sp. nov. (W9P-11T = KCTC 72992T = GDMCC 1.2803T), and Fulvivirga maritima sp. nov. (SW1-E11T = KCTC 72832T = GDMCC 1.2802T) are proposed.

The Discovery of the Fucoidan-Active Endo-1→4-α-L-Fucanase of the GH168 Family, Which Produces Fucoidan Derivatives with Regular Sulfation and Anticoagulant Activity.

Sulfated polysaccharides of brown algae, fucoidans, are known for their anticoagulant properties, similar to animal heparin. Their complex and irregular structure is the main bottleneck in standardization and in defining the relationship between their structure and bioactivity. Fucoidan-active enzymes can be effective tools to overcome these problems. In the present work, we identified the gene fwf5 encoding the fucoidan-active endo-fucanase of the GH168 family in the marine bacterium Wenyingzhuangia fucanilytica CZ1127T. The biochemical characteristics of the recombinant fucanase FWf5 were investigated. Fucanase FWf5 was shown to catalyze the endo-type cleavage of the 1→4-O-glycosidic linkages between 2-O-sulfated α-L-fucose residues in fucoidans composed of the alternating 1→3- and 1→4-linked residues of sulfated α-L-fucose. This is the first report on the endo-1→4-α-L-fucanases (EC 3.2.1.212) of the GH168 family. The endo-fucanase FWf5 was used to selectively produce high- and low-molecular-weight fucoidan derivatives containing either regular alternating 2-O- and 2,4-di-O-sulfation or regular 2-O-sulfation. The polymeric 2,4-di-O-sulfated fucoidan derivative was shown to have significantly greater in vitro anticoagulant properties than 2-O-sulfated derivatives. The results have demonstrated a new type specificity among fucanases of the GH168 family and the prospects of using such enzymes to obtain standard fucoidan preparations with regular sulfation and high anticoagulant properties.

Algicella marina gen. nov., sp. nov., a novel marine bacterium isolated from a Pacific red alga.

A novel Gram-staining negative, strictly aerobic, rod-shaped, and non-motile bacterium, designated strain 9Alg 56T, was isolated from the red alga Tichocarpus crinitus. The phylogenetic analysis based on 16S rRNA gene sequences placed the novel strain within the family Rhodobacteraceae, the order Rhodobacterales, the class Alphaproteobacteria, the phylum Pseudomonadota. The nearest neighbors of the new strain were Pontivivens insulae KCTC 42458T, Oceanibium sediminis KCTC 62076T, Halovulum dunhuangense YYQ-30T and Monaibacterium marinum C7T with 16S rRNA gene sequence similarity of 94.7, 94.4%, 93.1 and 92.7%, respectively. The AAI/ANI/dDDH values between 9Alg 56T and the five species of the closest genera (Pontivivens, Oceanibium, Halovulum, Monaibacterium, and 'Oceanomicrobium') were 58.63-63.91%/ 75.91-77.37%/ 19.3-20.4%. The prevalent fatty acids of strain 9Alg 56T were C18:1 ω7c, C18:0 and C14:0 3-OH. The polar lipid profile consisted of phosphatidylethanolamine, phosphatidylglycerol, phosphatidic acid, phosphatidylcholine, and two unidentified lipids. The DNA G+C content of strain 9Alg 56T was 61.5 mol%. A combination of the genotypic and phenotypic data showed that the algal isolate represents a novel genus and species, for which the name Algicella marina gen. nov., sp. nov. is proposed. The type strain is 9Alg 56T (= KCTC 72005T = KMM 6775T).

Aureibaculum algae sp. nov. isolated from the Pacific red alga Ahnfeltia tobuchiensis.

A Gram stain-negative, aerobic, rod-shaped, motile by gliding and yellow-orange-pigmented bacterium, designated strain 10Alg 115T, was isolated from the red alga Ahnfeltia tobuchiensis. The phylogenetic analysis based on 16S rRNA gene sequences placed the novel strain within the family Flavobacteriaceae, phylum Bacteroidetes. The nearest neighbor of the new isolate was Aureibaculum marinum KCTC 62204T with sequence similarity of 98.1%. The average nucleotide similarity and digital DNA-DNA hybridization values between the novel strain and Aureibaculum marinum KCTC 62204T were 80% and 22.3%, respectively. The prevalent fatty acids of strain 10Alg 115T were iso-C15:0, iso-C15:1 G, iso-C17:0 3-OH, iso-C16:0 3-OH and C15:0. The polar lipid profile consisted of phosphatidylethanolamine, two unidentified aminolipids and two unidentified lipids. The DNA G + C content of the type strain calculated from the whole-genome sequence was 32.2 mol%. A combination of the genotypic and phenotypic data showed that the algal isolate represents a novel species of the of genus Aureibaculum, for which the name Aureibaculum algae sp. nov. is proposed. The type strain is 10Alg 115T (= KCTC 62086T = KMM 6764T).

Aquimarina algiphila sp. nov., a chitin degrading bacterium isolated from the red alga Tichocarpus crinitus.

A strictly aerobic, Gram-stain-negative, rod-shaped, motile by gliding and yellow-orange pigmented flavobacterium, designated strain 9Alg 151T, was isolated from the Pacific red alga Tichocarpus crinitus. Phylogenetic analysis based on 16S rRNA gene sequences showed that the novel strain fell into the genus Aquimarina of the family Flavobacteriaceae with a 16S rRNA gene sequence similarity range of 94.2-98.2 % to the recognized species of the genus. Strain 9Alg 151T grew in the presence of 0.5-5 % NaCl and at 5-34 °C, and hydrolysed aesculin, agar, gelatin, starch, Tween 40, DNA and chitin. The predominant fatty acids were iso-C17 : 0 3-OH, iso-C15 : 0, iso-C15 : 1 G, iso-C15 : 0 3-OH, iso-C16 : 0, iso-C17 : 1ω8c and summed feature 3. The polar lipid profile comprised phosphatidylethanolamine, three unidentified aminolipids and three unidentified lipids. The major respiratory quinone was MK-6. The genomic DNA G+C content was 32.6 mol%. On the basis of 16S rRNA gene sequence data, and chemotaxonomic and phenotypic characteristics, strain 9Alg 151T represents a novel species of the genus Aquimarina, for which the name Aquimarina algiphila sp. nov. is proposed. The type strain is 9Alg 151T (=KCTC 23622T=KMM 6462T).

Polaribacter staleyi sp. nov., a polysaccharide-degrading marine bacterium isolated from the red alga Ahnfeltia tobuchiensis.

A Gram-stain-negative, rod-shaped, motile by gliding and yellow-pigmented bacterium, designated strain 10Alg 139T, was isolated from the Pacific red alga Ahnfeltiato buchiensis. The phylogenetic analysis based on 16S rRNA gene sequences showed that the novel strain belonged to the genus Polaribacter, a member of the family Flavobacteriaceae, the phylum Bacteroidetes, with highest sequence similarity to Polaribacter butkevichii KMM 3938T (99.3 %) and 93.3-98.6 % to other recognized Polaribacter species. The prevalent fatty acids of strain 10Alg 139T were iso-C15 : 0 3-OH, C15 : 0 3-OH, iso-C15:0, iso-C13 : 0, C15 : 0 and C15 : 1ω6c. The polar lipid profile consisted of the major lipids phosphatidylethanolamine, two unidentified aminolipids and four unidentified lipids. The main respiratory quinone was menaquinone 6. The DNA G+C content of the type strain is 31.8 mol%. The new isolate and the type strains of recognized species of the genus Polaribacter were readily distinguished based on a number of phenotypic characteristics. A combination of the genotypic and phenotypic data showed that the isolate from alga represents a novel species of the genus Polaribacter, for which the name Polaribacterstaleyi sp. nov. is proposed. The type strain is 10Alg 139T (=KCTC 52773T=KMM 6729T).

Olleya algicola sp. nov., a marine bacterium isolated from the green alga Ulva fenestrata.

A strictly aerobic, Gram-stain-negative, rod-shaped, motile by gliding and yellow-pigmented bacterium, designated strain 3Alg 18T, was isolated from the Pacific green alga Ulva fenestrata. Phylogenetic analysis based on 16S rRNA gene sequences showed that the novel strain was affiliated to the family Flavobacteriaceae of the phylum Bacteroidetes, being most closely related to the type strains of recognized species of the genus Olleya, with 16S rRNA gene sequence similarity of 97.9-99.3 %. Strain 3Alg 18T grew in the presence of 0.5-5 % (w/v) NaCl and at 4-37 °C, and hydrolysed aesculin, casein, gelatin, starch and Tweens 20, 40 and 80. The prevalent fatty acids were iso-C15 : 0, iso-C15 : 1 G, iso-C15 : 0 3-OH, iso-C16:0 2-OH, iso-C17 : 0 3-OH, summed feature 3, iso-C16 : 0 3-OH, anteiso-C15 : 0 and C15 : 0. The polar lipid profile contained phosphatidylethanolamine, three unidentified aminolipids and four unidentified lipids. The major respiratory quinone was menaquinone MK-6. The genomic DNA G+C content was 34.6 mol%. On the basis of 16S rRNA gene sequence data, and chemotaxonomic and phenotypic characteristics, strain 3Alg 18T represents a novel species of the genus Olleya, for which the name Olleya algicola sp. nov. is proposed. The type strain is 3Alg 18T (=KCTC 22024T=KMM 6133T).

Lacinutrix cladophorae sp. nov., a flavobacterium isolated from the green alga Cladophora stimpsonii, transfer of Flavirhabdus iliipiscaria Shakeela et al. 2015 to the genus Lacinutrix as Lacinutrix iliipiscaria comb. nov. and emended description of the genus Lacinutrix.

A strictly aerobic, Gram-stain-negative, rod-shaped, motile by gliding and yellow-pigmented bacterium, designated strain 7Alg 4T, was isolated from the green alga Cladophora stimpsonii. Phylogenetic analysis based on 16S rRNA gene sequences revealed that the novel strain was affiliated to the family Flavobacteriaceae of the phylum Bacteroidetes, and was most closely related to the recognized species of the genera Lacinutrixand Flavirhabdus, with 16S rRNA gene sequence similarities of 95.1-98.1 and 97.0 %, respectively. Strain 7Alg 4T grew in the presence of 1-5 % NaCl and at 4-32 °C, and hydrolysed aesculin, gelatin, starch and Tween 80. The prevalent fatty acids were iso-C15 : 1 G, iso-C15 : 0, iso-C17 : 0 3-OH, iso-C15 : 0 3-OH and C15 : 0. The polar lipid profile was characterized by the presence of phosphatidylethanolamine, three unidentified aminolipids and four unidentified lipids. The major respiratory quinone was MK-6. The DNA G+C content was 31.9 mol%. On the basis of the differences in 16S rRNA gene sequences, chemotaxonomic and phenotypic characteristics, it is suggested that strain 7Alg 4T represents a novel species of the genus Lacinutrix, for which the name Lacinutrixcladophorae sp. nov. is proposed. The type strain is 7Alg 4T (=KCTC 23036T=KMM 6381T). Reclassification of Flavirhabdus iliipiscaria as Lacinutrix iliipiscaria comb. nov. and an emend of the genus Lacinutrix are also proposed.

Amylibacter ulvae sp. nov., a new alphaproteobacterium isolated from the Pacific green alga Ulva fenestrata.

A strictly aerobic, Gram-stain-negative, rod-shaped and non-motile bacterium, designated strain 6Alg 255(T), was isolated from the green alga Ulva fenestrata. Phylogenetic analysis based on 16S rRNA gene sequencing revealed that the novel strain affiliated to the family Rhodobacteraceae of the class Alphaproteobacteria being most closely related to Amylibacter marinus LMG 28364(T) with 16S rRNA gene sequence similarity of 97.2 %. Strain 6Alg 255(T) grew with 0.5-6.0 % NaCl and at 4-33 °C, hydrolysed aesculin, casein, gelatin and urea. The DNA G + C content was 50.4 mol%. The prevalent fatty acids were C18:1 ω7c and C16:0. The polar lipid profile was characterized by the presence of phosphatidylethanolamine, phosphatidylglycerol, phosphatidylcholine and unidentified aminolipid. The major respiratory quinone was Q-10. The significant molecular distinctiveness between the novel isolate and its nearest neighbour was strongly supported by the differences in physiological and biochemical tests. Therefore, strain 6Alg 255(T) represents a novel species of the genus Amylibacter, for which the name Amylibacter ulvae sp. nov. is proposed. The type strain is 6Alg 255(T) (=KCTC 32465(T) = KMM 6515(T)).

Lutibacter holmesii sp. nov., a marine bacterium of the family Flavobacteriaceae isolated from the sea urchin Strongylocentrotus intermedius, and emended description of the genus Lutibacter.

Seven Gram-staining-negative, strictly aerobic, pale-yellow-pigmented, rod-shaped and non-motile strains were isolated from the sea urchin Strongylocentrotus intermedius collected from Troitsa Bay, Sea of Japan. Phylogenetic analyses based on 16S rRNA gene sequences showed that these isolates were affiliated with the family Flavobacteriaceae. The novel isolates showed 99.9-100 % 16S rRNA gene sequence similarity to each other and were closely related to the type strains of the recognized members of the genus Lutibacter with sequence similarities of 95.8-98.4 %. The G+C content of the genomic DNA was 35-36 mol%. DNA-DNA relatedness among the sea urchin isolates was 95-99 % and between strain KMM 6277T and its most closely related type strains, Lutibacter agarilyticus KCTC 23842T and Lutibacter litoralis JCM 13034T, was 38 and 27 %, respectively. The prevalent fatty acids were iso-C15 : 0, anteiso-C15 : 0, summed feature 3 (comprising iso-C15 : 0 2-OH and/or C16 : 1 ω7c fatty acids), iso-C15 : 1 and C15 : 0. The polar lipid profile was composed of the phosphatidylethanolamine, one unknown aminolipid and one unknown lipid. The main respiratory isoprenoid quinone was MK-6.The results of phylogenetic, phenotypic and genotypic analyses indicated that the novel strains represent a novel species within the genus Lutibacter, for which the name Lutibacter holmesii sp. nov. is proposed. The type strain is KMM 6277T ( = CCUG 62221T = LMG 26737T).

Winogradskyella litoriviva sp. nov., isolated from coastal seawater.

A Gram-stain-negative, facultatively anaerobic, moderately halophilic and gliding bacterium, designated KMM 6491T, was isolated from coastal seawater collected from Troitsa Bay, the Sea of Japan. Comparative analysis of 16S rRNA gene sequences revealed that strain KMM 6491T was a member of the genus Winogradskyella, with 94.5­97.9 % sequence similarity to recognized species of the genus Winogradskyella. The DNA G+C content of strain KMM 6491T was 31.3 mol% and DNA­DNA relatedness values with the type strains of Winogradskyella echinorum, Winogradskyella damuponensis, Winogradskyella eximia and Winogradskyella pulchriflava were in range of 10­26 %. Strain KMM 6491T contained menaquinone 6 (MK-6) as the single quinone and iso-C15 : 0, iso-C15 : 1, iso-C15 : 0 3-OH, iso-C17 : 0 3-OH, C15 : 0 and summed feature 3 (comprising iso-C15 : 0 2-OH and/or C16 : 1ω7c) anteiso-C15 : 0, as the prevalent fatty acids. The major polar lipids of strain KMM 6491T were phosphatidylethanolamine, two unknown aminolipids and two unknown lipids. Strain KMM 6491T was able to grow with 0.5­7 % NaCl and at 4­34 °C. The novel strain decomposed gelatin and starch and produced acid from d-glucose, maltose, mannose, rhamnose, sucrose, fructose and glycerol. On the basis of the results of the phylogenetic and phenotypic analyses it is suggested that strain KMM 6491T represents a novel species of the genus Winogradskyella, for which the name Winogradskyella litoriviva sp. nov. is proposed. The type strain is KMM 6491T ( = KCTC 23972T = LMG 26984T).

Flavobacterium ahnfeltiae sp. nov., a new marine polysaccharide-degrading bacterium isolated from a Pacific red alga.

A Gram-negative, aerobic, rod-shaped, motile by gliding and yellow-pigmented bacterium, designated strain 10Alg 130(T), that displayed the ability to destroy polysaccharides of red and brown algae, was isolated from the red alga Ahnfeltia tobuchiensis. The phylogenetic analysis based on 16S rRNA gene sequence placed the novel strain within the genus Flavobacterium, the type genus of the family Flavobacteriaceae, the phylum Bacteroidetes, with sequence similarities of 96.2 and 95.7 % to Flavobacterium jumunjiense KCTC 23618(T) and Flavobacterium ponti CCUG 58402(T), and 95.3-92.5 % to other recognized Flavobacterium species. The prevalent fatty acids of strain 10Alg 130(T) were iso-C15:0, iso-C15:0 3-OH, iso-C17:0 3-OH, C15:0 and iso-C17:1ω9c. The polar lipid profile consisted of phosphatidylethanolamine, two unknown aminolipids and three unknown lipids. The DNA G+C content of the type strain was 34.3 mol%. The new isolate and the type strains of recognized species of the genus Flavobacterium could strongly be distinguished by a number of phenotypic characteristics. A combination of the genotypic and phenotypic data showed that the algal isolate represents a novel species of the genus Flavobacterium, for which the name Flavobacterium ahnfeltiae sp. nov. is proposed. The type strain is 10Alg 130(T) (=KCTC 32467(T) = KMM 6686(T)).

Flavimarina pacifica gen. nov., sp. nov., a new marine bacterium of the family Flavobacteriaceae, and emended descriptions of the genus Leeuwenhoekiella, Leeuwenhoekiella aequorea and Leeuwenhoekiella marinoflava.

A facultatively anaerobic, Gram-stain negative, rod-shaped and yellow pigmented bacterium, designated strain IDSW-73(T), was isolated from a seawater sample and subjected to a polyphasic taxonomic study. Phylogenetic analysis based on 16S rRNA gene sequences revealed that the novel strain formed a distinct phyletic line in the family Flavobacteriaceae and is most closely related to the members of the genus Leeuwenhoekiella, with 16S rRNA gene sequence similarity of 91.4-92.6 %. Strain IDSW-73(T) was found to be able to grow with 0-12 % NaCl and at 4-33 °C; and was able to hydrolyse gelatin, starch and Tweens 20, 40 and 80. The DNA G+C content was determined to be 42.2 mol%. The predominant cellular fatty acids were identified as branched-chain saturated and unsaturated and straight-chain unsaturated fatty acids such as iso-C15:0, iso-C15:1, iso-C17:1 ω9c, C15:1 ω6c, iso-C15:0 3-OH, iso-C17:0 3-OH and summed feature 3 (as defined by MIDI), comprising iso-C15:0 2-OH and/or C16:1 ω7c. The polar lipids found were phosphatidylethanolamine, two unknown aminolipids and one unknown lipid. The major respiratory quinone was identified as MK-6. The significant molecular distinctiveness between the novel isolate and its nearest neighbours were strongly supported by notable differences in physiological and biochemical tests. Therefore, strain IDSW-73(T) is considered to represent a novel genus and species within the family Flavobacteriaceae, for which the name Flavimarina pacifica gen. nov., sp. nov. is proposed. The type strain is IDSW-73(T) (=KCTC 32466(T) = KMM 6759(T)). Emended descriptions of the recognized species of the genus Leeuwenhoekiella are also proposed.

Arenicella chitinivorans sp. nov., a gammaproteobacterium isolated from the sea urchin Strongylocentrotus intermedius.

A strictly aerobic, Gram-stain-negative, rod-shaped, non-motile and yellow-pigmented bacterial strain, designated KMM 6208(T), was isolated from a sea urchin. Phylogenetic analysis based on 16S rRNA gene sequencing revealed that this novel isolate was affiliated to the class Gammaproteobacteria and formed a robust cluster with Arenicella xantha KMM 3895(T) with 98.2 % 16S rRNA gene sequence similarity. Strain KMM 6208(T) grew in the presence of 0.5-5 % NaCl and at a temperature range of 4-38 °C. The isolate was oxidase-positive and hydrolysed aesculin, casein, chitin, gelatin, starch and Tweens 40 and 80. The prevalent fatty acids of strain KMM 6208(T) were C16 : 1ω7c, iso-C16 : 0, iso-C18 : 0, C18 : 1ω7c and C16 : 0. The polar lipids consisted of phosphatidylethanolamine, phosphatidylglycerol, diphosphatidylglycerol and an unidentified aminophospholipid, and the major isoprenoid quinone was Q-8. The DNA G+C content of strain KMM 6208(T) was 46.3 mol%. The DNA-DNA relatedness value of strain KMM 6208(T) with Arenicella xantha KMM 3895(T) was 5 %. Molecular data in a combination with phenotypic findings strongly suggest inclusion of this novel strain in the genus Arenicella as a representative of a novel species for which the name Arenicella chitinivorans sp. nov. is proposed. The type strain is KMM 6208(T) ( = KCTC 12711(T) = LMG 26983(T)).

Litorimonas cladophorae sp. nov., a new alphaproteobacterium isolated from the Pacific green alga Cladophora stimpsoni, and emended descriptions of the genus Litorimonas and Litorimonas taeaensis.

A strictly aerobic, Gram-stain-negative, rod-shaped and red-orange pigmented bacterium, designated strain KMM 6395(T), was isolated from the green alga Cladophora stimpsoni and subjected to a polyphasic taxonomic study. Phylogenetic analysis based on 16S rRNA gene sequencing revealed that the novel strain affiliated to the family Hyphomonadaceae of the class Alphaproteobacteria being most closely related to the type strain of the single species of the genus Litorimonas, Litorimonas taeanensis G5(T), with 16S rRNA gene sequence similarity of 96.8 %. Strain KMM 6395(T) grew with 1-5 % NaCl and at 4-35 °C, hydrolysed starch and Tween 80. The DNA G+C content was 48.7 mol%. The prevalent fatty acids were C18:1 ω7c, C19:1 ω8c and C18:1 ω7c 10-methyl. The polar lipid profile was characterized by the presence of phosphatidylglycerol, monoglycosyldiglyceride, glucuronopyranosyldiglyceride and an unidentified glycolipid. The major respiratory quinone was Q-10. The significant molecular distinctiveness between the novel isolate and its nearest neighbour, L. taeanensis G5(T), were strongly supported by the differences in physiological and biochemical tests. Therefore, strain KMM 6395(T) represents a novel species of the genus Litorimonas, for which the name Litorimonas cladophorae sp. nov. is proposed. The type strain is KMM 6395(T) (=KCTC 23968(T) = LMG 26985(T)). The emended descriptions of the genus Litorimonas and L. taeaensis are also provided.

Echinimonas agarilytica gen. nov., sp. nov., a new gammaproteobacterium isolated from the sea urchin Strongylocentrotus intermedius.

A novel Gram-negative, facultatively anaerobic and motile bacterial strain, designated KMM 6351(T), was isolated from the sea urchin Strongylocentrotus intermedius and examined using a polyphasic taxonomic approach. A phylogenetic analysis based on 16S rRNA gene sequencing revealed that the strain formed a distinct phyletic line in the class Gammaproteobacteria and was most closely related to the genera Aliivibrio, Photobacterium and Vibrio. Strain KMM 6351(T) grows at 4-40 °C and with 0.5-12 % NaCl and decomposes aesculin, agar, gelatin, starch, chitin and DNA. The DNA G+C content of the strain was determined to be 46.1 mol%. The prevalent fatty acids were found to be C(16:0), C(18:1) ω7c, C(12:0) 3-OH and summed feature 3 (comprising C(16:1) ω7c and/or iso-C(15:0) 2-OH fatty acids). The major polar lipids were determined to be diphosphatidylglycerol, phosphatidylglycerol, phosphatidylethanolamine and an unidentified aminolipid. The predominant ubiquinone was found to be Q-8. The results of the phenotypic, chemotaxonomic and genotypic analyses clearly indicated that the novel strain should be assigned to a new genus and species within the class γ-Proteobacteria for which the name Echinimonas agarilytica gen. nov., sp. nov. is proposed. The type strain is KMM 6351(T) (=KCTC 22996(T) = LMG 25420(T)).

Altererythrobacter troitsensis sp. nov., isolated from the sea urchin Strongylocentrotus intermedius.

An aerobic, halotolerant, Gram-negative bacterium was isolated from the sea urchin Strongylocentrotus intermedius and subjected to taxonomic characterization. The strain, designated KMM 6042(T), was rod-shaped, motile and yellow-pigmented. Phylogenetic analysis indicated that the strain was most closely related to the type strain of Altererythrobacter dongtanensis, and the level of 16S rRNA gene sequence similarity between the two was 99.0 %. However, the DNA-DNA relatedness between the two strains was 34.4 ± 7.6 %. Physiological and chemotaxonomic properties clearly distinguished the novel strain from other species of the genus Altererythrobacter. It is thus evident from the phylogenetic and phenotypic analyses that strain KMM 6042(T) merits recognition as a novel species of the genus Altererythrobacter, for which the name Altererythrobacter troitsensis sp. nov. (type strain, KMM 6042(T) = KCTC 12303(T) = JCM 17037(T)) is proposed.