Search

A novel yellow-pigmented bacterial strain (designated BR5-29 T ), was isolated and its taxonomy was studied. Phylogenetic study based on the 16S rRNA and draft genome sequence placed the strain BR5-29 T in a distinct lineage within the family Chitinophagaceae, sharing ≤ 93.4% sequence similarity with members of the closely related genera Ferruginibacter, Flavisolibacter, Flavitalea and Niastella. The novel isolate showed the highest sequence similarity to the genus Ferruginibacter. The draft genome of strain BR5-29 T had a total length of 5,505,520 base pairs. A total of 4585 genes were identified, in which 4537 were CDS and 48 RNA genes were assigned a putative function. The genome annotation of BR5-29 T showed 225 carbohydrate genes which may be responsible for the conversion of major ginsenosides to minor ginsenosides. Strain BR5-29 T contained MK-7 as a predominant quinone, and iso-C 15:0 , iso-C 15:0 G, iso-C 17:0 3-OH, and C 16:1 ω7c and/or C 16:1 ω6c (summed feature 3) as major fatty acids. The polar lipids found in the strain BR5-29 T were phosphatidylglycerol (PG), diphosphatidylglycerol (DPG), phosphatidylethanolamine (PE), five unidentified polar lipids (L1-L5), two unidentified aminolipid and one unidentified aminophospholipid. Our pilot data demonstrate that the novel isolate shares the similar major polar lipid PE, major quinone MK-7 and major fatty acids with the described members of the family Chitinophagaceae. However, the low 16S rRNA gene sequence (< 93.4%), the little high amount of C 12:0 , iso-C 17:0 2-OH and iso-C 15:1 2-OH fatty acids, low DNA G + C content, and the presence of DPG, PG and two unidentified polar lipids (L1 and L3 differentiate the BR5-29 T from its closest phylogenetic neighbors. Thus, the isolate represents a novel genus and species in the family Chitinophagaceae for which the name Ginsengibacter hankyongi gen. nov., sp. nov. is proposed. The type strain is BR5-29 T (= KACC 19446 T  = LMG 30462 T ). Thus, we predict that this novel strain may prove useful for the future research analysis (target gene cloning) and mass production of Rg3.

In this study, a novel ginsenoside transforming bacterium, strain W1-2-3 T , was isolated from mineral water. The 16S rRNA gene sequence analysis showed that strain W1-2-3 T shares 93.7-92.2% sequence similarity with the members of the family Sphingomonadaceae and makes a group with Sphingoaurantiacus capsulatus YLT33 T (93.7%) and S. polygranulatus MC 3718 T (93.4%). The novel isolate efficiently hydrolyses the ginsenoside Rc to Rd. The genome comprises a single circular 2,880,809, bp chromosome with 3211 genes in total, and 1993 protein coding genes. The isolate was observed to grow at 10-37 °C and at pH 6-10 on R2A agar medium; maximum growth was found to occur at 25 °C and pH 7.0. Strain W1-2-3 T was found to contain ubiquinone-10 as the predominant quinone and the fatty acids C 16:1 , C 17:1 ω6c, C 14:0 2-OH, summed feature 3 (C 16:1 ω6c/C 16:1 ω7c) and summed feature 8 (C 18 : 1 ω6c/C 18:1 ω7c). The DNA G+C content was determined to be 65.9 mol%. Strain W1-2-3 T can be distinguished from the other members of the family Sphingomonadaceae by a number of chemotaxonomic and phenotypic characteristics. The major polar lipids of strain W1-2-3 T were identified as phosphatidylethanolamine, an unidentified glycolipid and an unidentified polar lipid. The major poly amine was found to be homospermidine. Based on polyphasic taxonomic analysis, strain W1-2-3 T is concluded to represent a novel species within a new genus, for which the name Hankyongella ginsenosidimutans gen. nov., sp. nov. is proposed. The type strain of Hankyongella ginsenosidimutans is W1-2-3 T (= KACC 18307 T  = LMG 28594 T ).

A gram-negative, motile, strictly aerobic, and rod-shaped bacterium designated 176GS2-150 T was isolated from the sponge Hymeniacidon sinapium. The taxonomic position of the novel isolate was confirmed using the polyphasic approach. Strain 176GS2-150 T grew well at 25 °C on marine agar. Based on its 16S rRNA gene sequence, we showed that strain 176GS2-150 T belongs to the family Psychromonadaceae and class Gammaproteobacteria and is related to Corallincola platygyrae JLT2006 T (96.84% sequence similarity). The G + C content of the genomic DNA was 49.0 mol%. The assembled draft genome of strain 176GS2-150 T was 4.2 Mbp and consisted of 14 contigs. The major respiratory quinone was Q-8, and the major fatty acids were summed feature 3 (comprising C 16 :1 ω6c and/or C 16:1 ω7c), summed feature 8 (comprising C 18 :1 ω7c and/or C 18:1 ω6c), C 17:0 iso, C 16:0 , and C 15:0 iso. The polar lipids were phosphatidylglycerol, phosphatidylethanolamine, 3 unidentified phospholipids, and 1 unidentified polar lipid. On the basis of the genotypic and phenotypic characteristics, strain 176GS2-150 T can be placed as a new species within the genus Corallincola; the name Corallincola spongiicola sp. nov. has been proposed, with type strain 176GS2-150 T (= KACC 19890 T  = LMG 31317 T ).

A Gram-stain-negative, facultatively anaerobic, motile, light-yellow, and rod-shaped bacterium, designated as NY-02 T , was isolated from the sludge of a wastewater treatment plant in Hanam City, Republic of Korea. The comparison of 16S rRNA gene sequences revealed that the strain formed a distinct lineage within the genus Simplicispira and was most closely related to S. suum SC1-8 T (99.0%), S. limi EMB325 T (98.3%), S. psychrophila LMG 5408 T (98.2%), and S. piscis RSG39 T (97.4%). Both average nucleotide identity (ANI) and DNA-DNA hybridization (DDH) values between strain NY-02 T , and its closes type strains [S. suum SC1-8 T , S. limi EMB325 T , S. psychrophila LMG 5408 T , and S. piscis RSG39 T ] were lower than the cut-off (≥ 95-96% for ANI and ≥ 70% for DDH) to define a bacterial species. The genome comprises of 3,709,074 bp with a G + C content of 64.2 mol%. Ubiquinone 8 (Q-8) was the predominant quinone. The major fatty acids were C 16:0 and summed feature 3 (C 16:1 ω7c and/or C 16:1 ω6c), and the major polar lipids were phosphatidylglycerol, diphosphatidylglycerol, and phosphatidylethanolamine. The results of the physiological, biochemical, and taxonomic analyses in addition to low ANI and DNA-DNA relatedness values (82.2% and < 34.0%, respectively) indicate that the strain NY-02 T represents a novel species of the genus Simplicispira. The name proposed for strain NY-02 T (= KACC 19731 T  = LMG 31165 T ) is Simplicispira hankyongi sp. nov.

In this study, many bacterial strains were screened for the production of minor ginsenosides, but based on conversion competence among the strains, the strain Niabella ginsenosidivorans BS26 T has the good ginsenoside-transforming ability. Therefore, the strain BS26 T was selected for complete genome sequence analysis to determine the target (glycoside hydrolase) functional genes. Whole genome analysis of strain BS26 T showed 43 glycoside hydrolase genes in total. To determine the target functional gene, 12 sets of six different glycoside hydrolases (3 set of β-glucosidase; 3 set of trehalase; 3 set of arabinofuranosidase; 2 set of xylosidase; and one set of each α-galactosidase and α-fucosidase, respectively) were selected and cloned in E. coli BL21 (DE3) using the pGEX4T-1 vector and were characterized. Among these 12 sets of clones, only one, β-glucosidase (BglNg-767), showed ginsenoside conversion ability. The BglNg-767 comprised 767 amino acids and belonged to glycoside hydrolase family 3 (GH3). The recombinant GST-BglNg-767 was capable of altering the ginsenosides Rb 1 , Rd, and gypenoside XVII (Gyp-XVII) to F 2 ; Rb 2 to C-O; Rb 3 to C-Mx1, and Rc to C-Mc1. Besides, complete genome sequence analysis of strain BS26 T also indicates 30 endopeptidase genes, which may be responsible for self-hydrolysis of the proteins. Therefore, using SDS-PAGE analysis, we predict that the difference between the molecular weight of the expressed protein (around 90 kDa) and the predicted amino-acid sequence (102.7 kDa) is due to self-hydrolysis of the proteins., Competing Interests: Conflict of interestThe authors declare that there are no conflict of interest.

A Gram-stain negative, aerobic, motile, non-spore-forming and rod-shaped bacterial strain, designated YIM 730227 T , was isolated from a soil sample, collected from Karak district, Khyber-Pakhtun-Khwa, Pakistan. The bacterium was characterized using a polyphasic taxonomic approach. Pairwise comparison of the 16S rRNA gene sequences showed that strain YIM 730227 T is closely related to Phenylobacterium lituiforme FaiI3 T (97.5% sequence similarity), Phenylobacterium muchangponense A8 T (97.4%), Phenylobacterium panacis DCY109 T (97.1%), Phenylobacterium immobile E T (97.1%) and Phenylobacterium composti 4T-6 T (97.0%), while also sharing 98.0% sequence similarity with Phenylobacterium hankyongense HKS-05 T after NCBI blast, showing it represents a member of the family Caulobacteraceae. The major respiratory quinone was Q-10 and the major fatty acids were C 16:0, summed feature 8 (comprising C 18:1 ω7c and/or C 18:1 ω6c), C 18:1 ω7c 11-methyl and C 17:0 . The polar lipids were phosphatidylglycerol, unidentified glycolipids, phospholipid and unidentified lipid. The G + C content of the genomic DNA was 68.2 mol%. The DNA-DNA relatedness values of strain YIM 730227 T with P. hankyongense HKS-05 T , P. lituiforme FaiI3 T , P. muchangponense A8 T , P. panacis DCY109 T , P. immobile E T and P. composti 4T-6 T were 31.3 ± 0.6, 26.1 ± 0.2, 24.3 ± 0.1, 21.8 ± 0.9, 19.8 ± 0.6 and 18.2 ± 1.1%, respectively, values lower than 70%. Besides the morphological and chemotaxonomic characteristics, phylogenetic analyses of 16S rRNA gene sequences and the biochemical characteristics indicated that the strain YIM 730227 T represents a novel member of the genus Phenylobacterium, for which the name Phenylobacterium terrae sp. nov. (type strain YIM 730227 T = KCTC62324 T  = CGMCC 1.16326 T ) is proposed.

A Gram-stain negative, strictly aerobic, non-motile, non-spore-forming, and rod-shaped bacterial strain designated FW-3(T) was isolated from fresh water and its taxonomic position was investigated by using a polyphasic approach. Strain FW-3(T) was found to grow at 10-37 °C and at pH 7.0 in the absence of NaCl on nutrient agar. On the basis of 16S rRNA gene sequence similarity, strain FW-3(T) was shown to belong to the family Acetobacteraceae and to be related to Roseomonas lacus TH-G33(T) (97.2 % sequence similarity) and Roseomonas terrae DS-48(T) (96.4 %). The G+C content of the genomic DNA was determined to be 68.0 %. The major menaquinone was determined to be Q-10 and the major fatty acids were identified as summed feature 7 (comprising C18:1 ω9c/ω12t/ω7c as defined by the MIDI system; 55.4 %), and C18:1 2OH (29.8 %). DNA and chemotaxonomic data supported the affiliation of strain FW-3(T) to the genus Roseomonas. Strain FW-3(T) could be differentiated genotypically and phenotypically from the recognized species of the genus Roseomonas. The novel isolate therefore represents a novel species, for which the name Roseomonas sediminicola sp. nov. is proposed, with the type strain FW-3(T) (=KACC 16616(T) = JCM 18210(T)).

A Gram-positive, coccoid to rod-shaped, non-spore-forming bacterium, designated Gsoil 958(T), was isolated from soil of a ginseng field located in Pocheon province in South Korea. This bacterium was characterized in order to determine its taxonomic position by using a polyphasic approach. Strain Gsoil 958(T) was observed to grow well at 25-30 °C and at pH 7.0 on R2A and nutrient agar without NaCl supplementation. Strain Gsoil 958(T) was determined to have β-glucosidase activity and the ability to transform ginsenoside Rb1 (one of the dominant active components of ginseng) to F2 via gypenoside XVII and Rd. On the basis of 16S rRNA gene sequence similarity, strain Gsoil 958(T) was shown to belong to the family Nocardioidaceae and related most closely to Nocardioides koreensis MSL-09(T) (97.6 % 16S rRNA gene sequence similarity), Nocardioides aquiterrae GW-9(T) (97.0 %), and Nocardioides sediminis MSL-01(T) (97.0 %). The sequence similarities with other validly named species within the genus Nocardioides were less than 96.8 %. Strain Gsoil 958(T) was characterized chemotaxonomically as having LL-2,6-diaminopimelic acid in the cell-wall peptidoglycan, MK-8(H4) as the predominant menaquinone, and iso-C16:0, iso-C16:1 H, iso-C14:0, iso-C15:0 were identified as the major fatty acids. The G + C content of genomic DNA was determined to be 70.8 mol %. The chemotaxonomic properties and phenotypic characteristics supported the affiliation of strain Gsoil 958(T) to the genus Nocardioides. The results of both physiological and biochemical tests allowed for differentiation of strain Gsoil 958(T) from the recognized Nocardioides species. Therefore, strain Gsoil 958(T) is considered to represent a novel species of the genus Nocardioides, for which the name Nocardioides panaciterrulae sp. nov. is proposed, with the type strain Gsoil 958(T) (KACC 14271(T) = KCTC 19471(T) = DSM 21350(T)).

A Gram-negative, strictly aerobic, non-motile, non-spore-forming and rod-shaped bacterial strain designated KHI67(T) was isolated from sediment of the Gapcheon River in South Korea and its taxonomic position was investigated by using a polyphasic approach. Strain KHI67(T) was observed to grow optimally at 25-30 °C and at pH 7.0 on nutrient and R2A agar. On the basis of 16S rRNA gene sequence similarity, strain KHI67(T) was shown to belong to the family Sphingomonadaceae and was related to Sphingomonas faeni MA-olki(T) (97.6 % sequence similarity), Sphingomonas aerolata NW12(T) (97.5 %) and Sphingomonas aurantiaca MA101b(T) (97.3 %). The G + C content of the genomic DNA was determined to be 65.6 %. The major ubiquinone was found to be Q-10, the major polyamine was identified as homospermidine and the major fatty acids identified were summed feature 8 (comprising C18:1 ω7c/ω6c; 37.0 %), C16:0 (13.0 %), summed feature 3 (comprising C16:1 ω7c/C16:1 ω6c; 12.8 %) and C14:0 2OH (9.3 %). DNA and chemotaxonomic data supported the affiliation of strain KHI67(T) to the genus Sphingomonas. The DNA-DNA relatedness values between strain KHI67(T) and its closest phylogenetic neighbours were below 15 %. Strain KHI67(T) could be differentiated genotypically and phenotypically from the recognised species of the genus Sphingomonas. The isolate therefore represents a novel species, for which the name Sphingomonas ginsenosidivorax sp. nov. is proposed, with the type strain KHI67(T) (=KACC 14951(T) = JCM 17076(T) = LMG 25801(T)).

A novel α-L-arabinofuranosidase (Abf22-3) that could biotransform ginsenoside Rc into Rd was obtained from the ginsenoside converting Leuconostoc sp. strain 22-3, isolated from the Korean fermented food kimchi. The gene, termed abf22-3, consisting of 1,527 bp and encoding a protein with a predicted molecular mass of 58,486 Da was cloned into the pMAL-c2x (TEV) vector. A BLAST search using the Abf22-3's amino acid sequence revealed significant homology to that of family 51 glycoside hydrolases. The over-expressed recombinant Abf22-3 in Escherichia coli BL21 (DE3) catalyzed the hydrolysis of the arabinofuranoside moiety attached to the C-20 position of ginsenoside Rc under optimal conditions of pH 6.0 and 30 °C. This result indicated that Abf22-3 selectively converts ginsenoside Rc into Rd, but did not catalyze the hydrolysis of glucopyranosyl groups from Rc or other ginsenosides such as Rb1 and Rb2. Over-expressed recombinant enzymes were purified by two steps with amylose-affinity and DEAE-cellulose chromatography and then characterized. The kinetic parameters for α-L-arabinofuranosidase showed apparent Km and Vmax values of 0.95 ± 0.02 μM and 1.2 ± 0.1 μmol min(-1) mg of protein(-1) against p-nitrophenyl-α-L-arabinofuranoside, respectively. Using a purified MBP-Abf22-3 (10 μg/ml), 0.1 % of ginsenoside Rc was completely converted to ginsenoside Rd within 20 min.

Biotransformation of ginsenosides was examined using lactic acid bacteria isolated from several kinds of kimchi. A Gram-positive, facultatively anaerobic, non-motile, non-spore-forming, and rod-shaped lactic acid bacterial strain, designated EMML 3041(T), was determined to have ginsenoside-converting activity and its taxonomic position was investigated using a polyphasic approach. Strain EMML 3041(T) displayed β-glucosidase activity that was responsible for its ability to transform ginsenoside Rb1 (one of the dominant active components of ginseng) to F2 via gypenoside XVII, ginsenoside Rb2 to compound Y via compound O, ginsenoside Rc to compound Mc via compound Mc1, and ginsenoside Rd to ginsenoside F2. On the basis of the 16S rRNA gene sequence similarity, strain EMML 3041(T) was shown to belong to the genus Lactobacillus and is closely related to Lactobacillus versmoldensis KU-3(T) (98.3 % sequence similarity). Polyphasic taxonomy study confirmed that the strain EMML 3041(T) represents a novel species, for which the name Lactobacillus ginsenosidimutans sp. nov. is proposed, with EMML 3041(T) (=KACC 14527(T) = JCM 16719(T)) as the type strain.

Strain Gsoil 348(T) was isolated from a ginseng field soil sample by selecting micro-colonies from one-fifth strength modified R2A agar medium after a long incubation period. 16S rRNA gene sequence analysis indicated that the strain is related to members of the phylum Armatimonadetes (formerly called candidate phylum OP10). Strain Gsoil 348(T) is mesophilic, strictly aerobic, non-motile and rod-shaped. It only grows in low nutrient media. The major respiratory quinones are menaquinones MK-11 and MK-10, and the main fatty acids are iso-C(15:0), iso-C(17:0), C(16:0) and C(16:1) ω11c. The G+C content is 61.4 mol%. The 16S rRNA gene sequences in public databases belonging to the phylum Armatimonadetes were clustered here into 6 groups. Five of these groups constituted a coherent cluster distinct from the sequences of other phyla in phylogenetic trees that were constructed using multiple-outgroup sequences from 49 different phyla. On the basis of polyphasic taxonomic analyses, it is proposed that strain Gsoil 348(T) (= KACC 14959(T) = JCM 17079(T)) should be placed in Fimbriimonas ginsengisoli gen. nov., sp. nov., as the cultured representative of the Fimbriimonadia class. nov., corresponding with Group 4 of the phylum Armatimonadetes.

A Gram-positive, aerobic or facultative anaerobic, motile, spore-forming bacterial strain, designated Gsoil 1638T, was isolated from a soil sample of a ginseng field in Pocheon province (South Korea), and was characterized taxonomically by using a polyphasic approach. It grew well on nutrient agar medium, utilized a fairly narrow spectrum of carbon sources and tolerated 10% NaCl. The isolate was positive for catalase and oxidase tests but negative for the degradation of macromolecules such as casein, collagen, starch, chitin, CM-cellulose, xylan and DNA. The G + C content of the genomic DNA was 50.7 mol%. The predominant isoprenoid quinone was menaquinone 7 (MK-7). The major fatty acids were anteiso-C15:0 (44%) and C16:0 (25%). Comparative 16S rRNA gene sequence analysis showed that strain Gsoil 1638T fell within the radiation of the cluster comprising Paenibacillus species and joined Paenibacillus anaericanus DSM 15890T with a bootstrap value of 100%. These two strains shared 99.5% 16S rRNA gene sequence similarity with each other. The phylogenetic distance from any other validly described species within the genus Paenibacillus was less than 96.2%. DNA-DNA relatedness value between strain Gsoil 1638T and its closest phylogenetic neighbor, Paenibacillus anaericanus, was 62%. On the basis of its phenotypic properties and phylogenetic distinctiveness, strain Gsoil 1638T (= KCTC 13931T = LMG 23406T = CCUG 52472T) was classified in the genus Paenibacillus as the type strain of a novel species, for which the name Paenibacillus ginsengisoli sp. nov. is proposed.