Genome-based taxonomy and functional prediction of Sphingomonas fuzhouensis sp. nov. and Massilia phyllosphaerae sp. nov. isolated from Pennisetum sp. with plant growth-promoting potential.

Two facultatively aerobic strains, designated SGZ-02T and SGZ-792T, were isolated from plant Pennisetum sp., exhibiting the highest 16S rRNA gene sequence similarities with the type strains of Sphingomonas zeae LMG 28739T (98.6%) and Massilia forsythiae NBRC 114511T (98.4%), respectively. SGZ-02T grew between 5 and 45 °C, pH 5.0-11.0 and tolerated NaCl concentrations of 0-4% (w/v), whereas SGZ-792T thrived at 5-40 °C, pH 5.0-11.0 and NaCl tolerance to 0-3.5% (w/v). The major quinone of SGZ-02T was ubiquinone-10, with the dominant fatty acids being C16:0 (13.5%), Summed Feature 3 (6.3%), C14:02-OH (5.3%) and Summed Feature 8 (66.3%). SGZ-792T predominantly contained ubiquinone-8, with major fatty acids being C16:0 (20.3%), Summed Feature 3 (5.0%) and Summed Feature 8 (54.7%). Average nucleotide identity and digital DNA-DNA hybridization values between two strains and their closest references strains were below the bacterial species threshold. Based on genotypic and phenotypic characteristics, strains SGZ-02T and SGZ-792T are proposed as novel species within the genera Sphingomonas and Massilia, respectively. The suggested names for the new species are Sphingomonas fuzhouensis sp. nov. (SGZ-02T = GDMCC 1.4033T = JCM 36769T) and Massilia phyllosphaerae sp. nov. (SGZ-792T = GDMCC 1.4211T = JCM 36643T), respectively.

Functional analysis, diversity, and distribution of the ean cluster responsible for 17ß-estradiol degradation in sphingomonads.

17ß-estradiol (E2) is a natural endocrine disruptor that is frequently detected in surface and groundwater sources, thereby threatening ecosystems and human health. The newly isolated E2-degrading strain Sphingomonas colocasiae C3-2 can degrade E2 through both the 4,5-seco pathway and the 9,10-seco pathway; the former is the primary pathway supporting the growth of this strain and the latter is a branching pathway. The novel gene cluster ean was found to be responsible for E2 degradation through the 4,5-seco pathway, where E2 is converted to estrone (E1) by EanA, which belongs to the short-chain dehydrogenases/reductases (SDR) superfamily. A three-component oxygenase system (including the P450 monooxygenase EanB1, the small iron-sulfur protein ferredoxin EanB2, and the ferredoxin reductase EanB3) was responsible for hydroxylating E1 to 4-hydroxyestrone (4-OH-E1). The enzymatic assay showed that the proportion of the three components is critical for its function. The dioxygenase EanC catalyzes ring A cleavage of 4-OH-E1, and the oxidoreductase EanD is responsible for the decarboxylation of the ring A-cleavage product of 4-OH-E1. EanR, a TetR family transcriptional regulator, acts as a transcriptional repressor of the ean cluster. The ean cluster was also found in other reported E2-degrading sphingomonads. In addition, the novel two-component monooxygenase EanE1E2 can open ring B of 4-OH-E1 via the 9,10-seco pathway, but its encoding genes are not located within the ean cluster. These results refine research on genes involved in E2 degradation and enrich the understanding of the cleavages of ring A and ring B of E2.IMPORTANCESteroid estrogens have been detected in diverse environments, ranging from oceans and rivers to soils and groundwater, posing serious risks to both human health and ecological safety. The United States National Toxicology Program and the World Health Organization have both classified estrogens as Group 1 carcinogens. Several model organisms (proteobacteria) have established the 4,5-seco pathway for estrogen degradation. In this study, the newly isolated Sphingomonas colocasiae C3-2 could degrade E2 through both the 4,5-seco pathway and the 9,10-seco pathway. The novel gene cluster ean (including eanA, eanB1, eanC, and eanD) responsible for E2 degradation by the 4,5-seco pathway was identified; the novel two-component monooxygenase EanE1E2 can open ring B of 4-OH-E1 through the 9,10-seco pathway. The TetR family transcriptional regulator EanR acts as a transcriptional repressor of the ean cluster. The cluster ean was also found to be present in other reported E2-degrading sphingomonads, indicating the ubiquity of the E2 metabolism in the environment.

Sphingomonas flavescens sp. nov., isolated from soil.

An aerobic bacterium, designated as PT-12T, was isolated from soil collected from agriculture field, and its taxonomic position was validated through a comprehensive polyphasic methodology. The strain was identified as Gram-stain-negative, non-motile, rod-shaped, and catalase- and oxidase-positive. The yellow-colored colonies showed growth ability at temperature range of 18-37 °C, NaCl content of 0-1.0% (w/v), and at a pH of 6.0-8.0. The 16S rRNA gene and phylogenetic analysis showed that strain PT-12T affiliated with the genus Sphingomonas in the family Sphingomonadaceae, and displayed the highest 16S rRNA nucleotide sequence similarity with Sphingomonas limnosediminicola 03SUJ6T (98.4%). The genome size of strain PT-12T was 2,656,862 bp and the DNA G + C content estimated from genome was 63.5%. The highest values of average nucleotide identity (ANI) and digital DNA-DNA hybridization (dDDH) were observed between strain PT-12T and Sphingomonas segetis YJ09T, accounting to 76.2% and 20.2%, respectively. In addition, both ANI and dDDH values between strain PT-12T and other phylogenetically related neighbors ranged between 69.6% and 76.2% and 18.4% and 20.2%, respectively. Chemotaxonomic features exhibited Q-10 as the only ubiquinone; homospermidine as the major polyamine; summed feature 8 (C18:1ω7c and/or C18:1ω6c), C16:0, and 10-methyl C18:0 as the notable fatty acids; and phosphatidylethanolamine, phosphatidylglycerol, diphosphatidylglycerol, phosphatidylcholine, and sphingoglycolipid as dominating polar lipids. Overall, the comprehensive polyphasic data supported that strain PT-12T represents a novel bacterial species within the genus Sphingomonas. Accordingly, we propose the name Sphingomonas flavescens sp. nov. The type strain is PT-12T (= KCTC 92114T = NBRC 115717T).

Metagenomic and proteomic insights into the self-adaptive cell surface hydrophobicity of Sphingomonas sp. strain PAH02 reducing the migration of cadmium-phenanthrene co-pollutant in rice.

Cell surface hydrophobicity (CSH) dominates the interactions between rhizobacteria and pollutants at the soil-water interface, which is critical for understanding the dissipation of pollutants in the rhizosphere microzone of rice. Herein, we explored the effects of self-adaptive CSH of Sphingomonas sp. strain PAH02 on the translocation and biotransformation behaviour of cadmium-phenanthrene (Cd-Phe) co-pollutant in rice and rhizosphere microbiome. We evidenced that strain PAH02 reduced the adsorption of Cd-Phe co-pollutant on the rice root surface while enhancing the degradation of Phe and adsorption of Cd via its self-adaptive CSH in the hydroponic experiment. The significant upregulation of key protein expression levels such as MerR, ARHDs and enoyl-CoA hydratase/isomerase, ensures self-adaptive CSH to cope with the stress of Cd-Phe co-pollutant. Consistently, the bioaugmentation of strain PAH02 promoted the formation of core microbiota in the rhizosphere soil of rice (Oryza sativa L.), such as Bradyrhizobium and Streptomyces and induced gene enrichment of CusA and PobA that are strongly associated with pollutant transformation. Consequently, the contents of Cd and Phe in rice grains at maturity decreased by 17.2% ± 0.2% and 65.7% ± 0.3%, respectively, after the bioaugmentation of strain PAH02. These findings present new opportunities for the implementation of rhizosphere bioremediation strategies of co-contaminants in paddy fields.

Structural conservation in the glutathione binding in Sphingomonas sp. glutaredoxin Grx3 and variations for cold adaptation.

Glutaredoxin 3 (Grx3), a redox protein with a thioredoxin-fold structure, maintains structural integrity and glutathione (GSH) binding capabilities across varying habitat temperatures. The cis-Pro loop, essential for GSH binding, relies on the Arg-Asp salt bridge (α2-α3) and Gln-His hydrogen bond (ß3-ß4) for its conformation. In some psychrophilic Grx3 variants, Arg in α2 is replaced with Tyr, and His in ß4 is replaced with Phe. This study examines the roles of these bonds in Grx3's structure, function, and cold adaptation, using SpGrx3 from the Arctic bacterium Sphingomonas sp. Despite its cold habitat, SpGrx3 maintains the Arg51-Asp69 salt bridge and Gln56-His63 hydrogen bond. The R51Y substitution disrupts the α2-α3 salt bridge, while the H63F and H63Y substitutions hinder the salt bridge through cation-π interactions with Arg51, involving Phe63/Tyr63, thereby enhancing flexibility. Conversely, mutations that disrupt the hydrogen bond (Q56A, H63A, and H63F) reduce thermal stability. In the psychrophilic Grx3 configuration A48T/R51Y/H63F, a Thr48-Gln56 hydrogen bond stabilizes the cis-Pro loop, enhancing flexibility by disrupting both bonds. Furthermore, all mutants exhibit reduced α-helical content and catalytic efficiency. In summary, the highly conserved Arg51-Asp69 salt bridge and Gln56-His63 hydrogen bond are crucial for stabilizing the cis-Pro loop and catalytic activity in SpGrx3. His63 is favored as it avoids cation-π interactions with Arg51, unlike Phe63/Tyr63. Psychrophilic Grx3 variants have adapted to cold environments by reducing GSH binding and increasing structural flexibility. These findings deepen our understanding of the structural conservation in Grx3 for GSH binding and the critical alterations required for cold adaptation.

A novel UV-resistant bacterium Sphingomonas endolithica sp. nov., and genomic analysis, isolated from the north slope of Mount Everest.

Gram-stain-negative, aerobic, rod-shaped, non-motile bacterium strain ZFBP2030T was isolated from a rock on the North slope of Mount Everest. This strain contained a unique ubiquinone-10 (Q-10) as a predominant respiratory quinone. Among the tested fatty acids, the strain contained summed feature 8, C14:0 2OH, and C16:0, as major cellular fatty acids. The polar lipid profile contained phosphatidyl glycerol, phosphatidyl ethanolamine, three unidentified phospholipids, two unidentified aminolipids, and six unidentified lipids. The cell-wall peptidoglycan was a meso-diaminopimelic acid, and cell-wall sugars were ribose and galactose. Phylogenetic analyses based on 16S rRNA gene sequence revealed that strain ZFBP2030T was a member of the genus Sphingomonas, exhibiting high sequence similarity to the 16S rRNA gene sequences of Sphingomonas aliaeris DH-S5T (97.9%), Sphingomonas alpina DSM 22537T (97.3%) and Sphingomonas hylomeconis CCTCC AB 2013304T (97.0%). The 16S rRNA gene sequence similarity between ZFBP2030T and other typical strains was less than 97.0%. The average amino acid identity values, average nucleotide identity, and digital DNA-DNA hybridization values between strain ZFBP2030T and its highest sequence similarity strains were 56.9-79.9%, 65.1-82.2%, and 19.3-25.8%, respectively. The whole-genome size of the novel strain ZFBP2030T was 4.1 Mbp, annotated with 3838 protein-coding genes and 54 RNA genes. Moreover, DNA G + C content was 64.7 mol%. Stress-related functions predicted in the subsystem classification of the strain ZFBP2030T genome included osmotic, oxidative, cold/heat shock, detoxification, and periplasmic stress responses. The overall results of this study clearly showed that strain ZFBP2030T is a novel species of the genus Sphingomonas, for which the name Sphingomonas endolithica sp. nov. is proposed. The type of strain is ZFBP2030T (= EE 013T = GDMCC 1.3123T = JCM 35386T).

Sphingomonas abietis sp. nov., an Endophytic Bacterium Isolated from Korean Fir.

PAMB 00755T, a bacterial strain, was isolated from Korean fir leaves. The strain exhibits yellow colonies and consists of Gram-negative, non-motile, short rods or ovoid-shaped cells. It displays optimal growth conditions at 20°C, 0% NaCl, and pH 6.0. Results of 16S rRNA gene-based phylogenetic analyses showed that strain PAMB 00755T was most closely related to Sphingomonas chungangi MAH-6T (97.7%) and Sphingomonas polyaromaticivorans B2-7T (97.4%), and ≤96.5% sequence similarity to other members of the genus Sphingomonas. The values of average nucleotide identity (79.9-81.3%), average amino acid identity (73.3-75.9%), and digital DNA-DNA hybridization (73.3-75.9%) were significantly lower than the threshold values for species boundaries; these overall genome-related indexes (OGRI) analyses indicated that the strain represents a novel species. Genomic analysis revealed that the strain has a 4.4-Mbp genome encoding 4,083 functional genes, while the DNA G+C content of the whole genome is 66.1%. The genome of strain PAMB 00755T showed a putative carotenoid biosynthetic cluster responsible for its antioxidant activity. The respiratory quinone was identified as ubiquinone 10 (Q-10), while the major fatty acids in the profile were identified as C18:1ω7c and/or C18:1ω6c (summed feature 8). The major polar lipids of strain PAMB 00755T were diphosphatidylglycerol, phosphatidylethanolamine, sphingoglycolipid, and phosphatidylcholine. Based on a comprehensive analysis of genomic, phenotypic, and chemotaxonomic characteristics, we proposed the name Sphingomonas abietis sp. nov. for this novel species, with PAMB 00755T as the type strain (= KCTC 92781T = GDMCC 1.3779T).

Regulation Mechanism of Nicotine Catabolism in Sphingomonas melonis TY by a Dual Role Transcriptional Regulator NdpR.

A gene cluster ndp, responsible for nicotine degradation via a variant of the pyridine and pyrrolidine pathways, was previously identified in Sphingomonas melonis TY, but the regulation mechanism remains unknown. The gene ndpR within the cluster was predicted to encode a TetR family transcriptional regulator. Deletion of ndpR resulted in a notably shorter lag phase, higher maximum turbidity, and faster substrate degradation when cultivated in the presence of nicotine. Real-time quantitative PCR and promoter activity analysis in wild-type TY and TYΔndpR strains revealed that genes in the ndp cluster were negatively regulated by NdpR. However, complementation of ndpR to TYΔndpR did not restore transcription repression, but, instead, the complemented strain showed better growth than TYΔndpR. Promoter activity analysis indicates that NdpR also functions as an activator in the transcription regulation of ndpHFEGD. Further analysis through electrophoretic mobility shift assay and DNase I footprinting assay revealed that NdpR binds five DNA sequences within ndp and that NdpR has no autoregulation. These binding motifs overlap with the -35 or -10 box or are located distal upstream of the corresponding transcriptional start site. Multiple sequence alignment of these five NdpR-binding DNA sequences found a conserved motif, with two of the binding sequences being partially palindromic. 2,5-Dihydroxypyridine acted as a ligand of NdpR, preventing NdpR from binding to the promoter region of ndpASAL, ndpTB, and ndpHFEGD. This study revealed that NdpR binds to three promoters in the ndp cluster and is a dual-role transcriptional regulator in nicotine metabolism. IMPORTANCE Gene regulation is critical for microorganisms in the environment in which they may encounter various kinds of organic pollutants. Our study revealed that transcription of ndpASAL, ndpTB, and ndpHFEGD is negatively regulated by NdpR, and NdpR also exhibits a positive regulatory effect on PndpHFEGD. Furthermore, 2,5-dihydroxypyridine was identified as the effector molecular for NdpR and can both prevent the binding of free NdpR to the promoter and release NdpR from the promoters, which is different from previously reported NicR2. Additionally, NdpR was found to have both negative and positive transcription regulatory effects on the same target, PndpHFEGD, while only one binding site was identified, which is notably different from the previously reported TetR family regulators. Moreover, NdpR was revealed to be a global transcriptional regulator. This study provides new insight into the complex gene expression regulation of the TetR family.

Interaction Between Chronic Endometritis Caused Endometrial Microbiota Disorder and Endometrial Immune Environment Change in Recurrent Implantation Failure.

Objective: To investigate the Interaction between chronic endometritis (CE) caused endometrial microbiota disorder and endometrial immune environment change in recurrent implantation failure (RIF). Method: Transcriptome sequencing analysis of the endometrial of 112 patients was preform by using High-Throughput Sequencing. The endometrial microbiota of 43 patients was analyzed by using 16s rRNA sequencing technology. Result: In host endometrium, CD4 T cell and macrophage exhibited significant differences abundance between CE and non-CE patients. The enrichment analysis indicated differentially expressed genes mainly enriched in immune-related functional terms. Phyllobacterium and Sphingomonas were significantly high infiltration in CE patients, and active in pathways related to carbohydrate metabolism and/or fat metabolism. The increased synthesis of lipopolysaccharide, an important immunomodulator, was the result of microbial disorders in the endometrium. Conclusion: The composition of endometrial microorganisms in CE and non-CE patients were significantly different. Phyllobacterium and Sphingomonas mainly regulated immune cells by interfering with the process of carbohydrate metabolism and/or fat metabolism in the endometrium. CE endometrial microorganisms might regulate Th17 response and the ratio of Th1 to Th17 through lipopolysaccharide (LPS).

Induction of Apoptosis in Human Lung Epithelial Cell by Sphingomonas sp. Shah, a Recently Identified Cell Culture Contaminant.

Sphingomonas sp. Shah is a bacterium that was first isolated from mammalian cell cultures. According to ribotyping data it is very much homologous to the clinically important pathogen Sphingomonas paucimobilis, which has generated pseudo-outbreaks. Using a tissue culture system, Sphingomonas sp. Shah was discovered to induce apoptosis in human lung epithelial carcinoma. Apoptosis of infected cells was determined by numerous criteria including (1) visual alterations in cellular morphology, (2) initiation of nuclear marginalization and chromatin compaction condensation, (3) the attendance of a high percentage of cells with subG1 DNA content, and (4) caspase-3 activation. In the current study we demonstrate the induction of apoptosis in mammalian lung epithelial cells upon infection with Sphingomonas sp. Shah and provide insight into the molecular processes triggering apoptosis.

Sphingomonas sabuli sp. nov., a carotenoid-producing bacterium isolated from beach sand.

A Gram-stain-negative, aerobic and non-motile bacterium, strain sand1-3T, was isolated from beach sand collected from Haeundae Beach located in Busan, Republic of Korea. Based on the results of 16S rRNA gene sequence and phylogenetic analyses, Sphingomonas daechungensis CH15-11T (97.0 %), Sphingomonas edaphi DAC4T (96.8 %), Sphingomonas xanthus AE3T (96.5 %) and Sphingomonas oryziterrae YC6722T (96.0 %) were selected for comparing phenotypic and chemotaxonomic characteristics. Cells of strain sand1-3T grew at 7-50 °C (optimum, 30-35 °C), pH 5.0-8.0 (optimum, pH 7.0-8.0) and in the presence of 0-0.5 % (w/v) NaCl (optimum, 0 %). Major polar lipids included diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylglycerol, sphingoglycolipid, one unidentified glycolipid and one unidentified phosphoglycolipid. The major fatty acids were summed feature 8 (C18 : 1 ω6c and/or C18 : 1 ω7c) and C18 : 1 2-OH. Moreover, the sole respiratory quinone and major polyamine were identified as ubiquinone-10 and homospermidine, respectively. The genomic DNA G+C content was 65.9 mol%. The digital DNA-DNA hybridization, average nucleotide identity and average amino acid identity values of strain sand1-3T and its reference strains with publicly available genomes were 17.9-18.9 %, 72.0-75.3 % and 63.3-76.5 % respectively. Based on polyphasic evidence, we propose Sphingomonas sabuli sp. nov. as a novel species within the genus Sphingomonas. The type strain is sand1-3T (=KCTC 82358T=NBRC 114538T).

Description of Nocardioides piscis sp. nov., Sphingomonas piscis sp. nov. and Sphingomonas sinipercae sp. nov., isolated from the intestine of fish species Odontobutis interrupta (Korean spotted sleeper) and Siniperca scherzeri (leopard mandarin fish).

A polyphasic taxonomic approach was used to characterize three novel bacterial strains, designated as HDW12AT, HDW-15BT, and HDW15CT, isolated from the intestine of fish species Odontobutis interrupta or Siniperca scherzeri. All isolates were obligate aerobic, non-motile bacteria, and grew optimally at 30°C. Phylogenetic analysis based on 16S rRNA sequences revealed that strain HDW12AT was a member of the genus Nocardioides, and closely related to Nocardioides allogilvus CFH 30205T (98.9% sequence identities). Furthermore, strains HDW15BT and HDW15CT were members of the genus Sphingomonas, and closely related to Sphingomonas lutea JS5T and Sphingomonas sediminicola Dae 20T (97.1% and 97.9% sequence identities), respectively. Strain HDW12AT contained MK-8 (H4), and strains HDW15BT and HDW15CT contained Q-10 as the respiratory quinone. Major polar lipid components of strain HDW12AT were diphosphatidylglycerol, phosphatidylglycerol, and phosphatidylinositol, and those of strains HDW15BT and HDW15CT were sphingoglycolipid, diphosphatidylglycerol, phosphatidylglycerol, phosphatidylethanolamine, and phosphatidylcholine. The G + C content of strains HDW12AT, HDW15BT, and HDW15CT were 69.7, 63.3, and 65.5%, respectively. The results of phylogenetic, phenotypic, chemotaxonomic, and genotypic analyses suggest that strain HDW12AT represents a novel species within the genus Nocardioides, and strains HDW15BT and HDW15CT represent two novel species within the genus Sphingomonas. We propose the names Nocardioides piscis for strain HDW12AT (= KACC 21336T = KCTC 49321T = JCM 33670T), Sphingomonas piscis for strain HDW15BT (= KACC 21341T = KCTC 72588T = JCM 33738T), and Sphingomonas sinipercae for strain HDW15CT (= KACC 21342T = KCTC 72589T = JCM 33739T).

Cadmium-resistant and arginine decarboxylase-producing endophytic Sphingomonas sp. C40 decreases cadmium accumulation in host rice (Oryza sativa Cliangyou 513).

In this study, an cadmium (Cd)-immobilizing and arginine decarboxylase-producing endophytic Sphingomonas sp. strain C40 obtained from the seeds of Oryza sativa Cliangyou 513 was characterized for its Cd availability and Cd uptake in host rice using hydroponic and soil experiments. The Cd concentration decreased by 51-95% compared to the control, while the spermidine concentration increased by 19-25% with Cd compared with no Cd in the strain C40-inoculated solution. Strain C40 decreased the above-ground tissue Cd content by 27-37% and increased spermine and spermidine contents by 28-67% and the expression levels of genes involved in spermine and spermidine production by 29-217% in rice roots compared to the controls. Furthermore, correlation analyses showed the significantly negative correlation between rice root spermine and spermidine contents and above-ground tissue Cd content. In the Cd-added soil, strain C40 promoted the rice biomass by 29-36% and decreased rice root, above-ground tissue, and grain Cd contents by 18, 16, and 33% and total grain Cd uptake by 14% compared with the controls at the maturity stage. Strain C40 decreased the exchangeable Cd content by 27% and increased the Fe and Mn oxides-bound Cd content by 45% in the rice rhizosphere soils at the maturity stage compared with the controls. These results suggested that the endophytic bacterial strain C40 increased rice root polyamine production and their related gene expression and the transformation of available Cd to unavailable Cd, leading to reduced Cd accumulation and translocation from the rice roots to grains.

Heterologous production of new lasso peptide koreensin based on genome mining.

Lasso peptides are a class of ribosomally biosynthesized and posttranslationally modified peptides with a knot structure as a common motif. Based on a genome search, a new biosynthetic gene cluster of lasso peptide was found in the genome of the proteobacterium Sphingomonas koreensis. Interestingly, the amino acid sequence of the precursor peptide gene includes two cell adhesion motif sequences (KGD and DGR). Heterologous production of the new lasso peptide was performed using the cryptic biosynthetic gene cluster of S. koreensis. As a result, a new lasso peptide named koreensin was produced by the gene expression system in the host strain Sphingomonas subterranea. The structure of koreensin was determined by NMR and ESI-MS analysis. The three-dimensional structure of koreensin was obtained based on an NOE experiment and the coupling constants. A variant peptide (koreensin-RGD), which had RGD instead of KGD, was produced by heterologous production with site-directed mutagenesis experiment. Koreensin and koreensin-RGD did not show cell adhesion inhibitory activity, although the molecules possessed cell adhesion motifs. The possible presence of a salt bridge between the motifs in koreensin was indicated, and it may prevent the cell adhesion motif from functioning.

Sphingomonas palmae sp. nov. and Sphingomonas gellani sp. nov., endophytically associated phyllosphere bacteria isolated from economically important crop plants.

In this study, two endophytic bacterial strains designated JS21-1T and S6-262T isolated from leaves of Elaeis guineensis and stem tissues of Jatropha curcas respectively, were subjected for polyphasic taxonomic approach. On R2A medium, colonies of strains JS21-1T and S6-262T are orange and yellow, respectively. Phylogenetic analyses using 16S rRNA gene sequencing and whole-genome sequences placed the strains in distinct clades but within the genus Sphingomonas. The DNA G + C content of JS21-1T and S6-262T were 67.31 and 66.95%, respectively. Furthermore, the average nucleotide identity and digital DNA-DNA hybridization values of strains JS21-1T and S6-262T with phylogenetically related Sphingomonas species were lower than 95% and 70% respectively. The chemotaxonomic studies indicated that the major cellular fatty acids of the strain JS21-1T were summed feature 8 (C18:1 ω7c and/or C18:1 ω6c), C16:0, and C14:0 2OH; strain S6-262T possessed summed feature 3 (C16:1 ω7c and/or iso-C15:0 2-OH) and summed feature 8 (C18:1 ω6c and/or C18:1 ω7c). The major quinone was Q10, and the unique polyamine observed was homospermidine. The polar lipid profile comprised of mixture of sphingoglycolipid, phosphatidylethanolamine, phosphatidylglycerol, diphosphatidylglycerol and certain uncharacterised phospholipids and lipids. Based on this polyphasic evidence, strains JS21-1T and S6-262T represent two novel species of the genus Sphingomonas, for which the names Sphingomonas palmae sp. nov. and Sphingomonas gellani sp. nov. are proposed, respectively. The type strain of Sphingomonas palmae sp. nov. is JS21-1T (= DSM 27348T = KACC 17591T) and the type strain of Sphingomonas gellani sp. nov. is S6-262T (= DSM 27346T =  KACC 17594T).

Sphingomonas parva sp. nov., isolated from soil in Jeju Island.

A Gram staining-negative, yellow-colored, rod-shaped, non-motile and aerobic, designated strain 17J27-24T was isolated from a soil sample in Korea. Phylogenetic analysis based on 16S rRNA gene sequences revealed that strain 17J27-24T was related to the members of the family Sphingomonadaceae and formed a distinct monophyletic cluster within the genus Sphingomonas with Sphingomonas deserti (98.3% 16S rRNA gene sequence similarity). Growth was observed at 30 °C (optimum), at pH 7.0 (optimum), and in the absence of NaCl (%). The predominant cellular fatty acids were summed feature 8 (18:1 ω7c and/or 18:1 ω6c) and C17:1 ω6c. The major respiratory quinone was Q10. The polar lipids profile comprised of diphosphatidylglycerol (DPG), phosphatidylglycerol (PG), phosphatidylethanolamine (PE), and sphingoglycolipid (SGL). The DNA G + C content was 77.8 mol%. The average nucleotide identity (ANI) and digital DNA-DNA hybridisation (dDDH) values between 17J27-24T and its phylogenetically closest Sphingomonas deserti (KCTC 62411T) were below the established cut-off < 94% (ANI) and < 70% (dDDH) for species delineation. Moreover, the results of the polyphasic approach confirmed that strain 17J27-24T represents a novel species of the genus Sphingomonas within the family Sphingomonadaceae, for which the name Sphingomonas parva sp. nov. is proposed. The type strain of this species is 17J27-24T (= KCTC 62208T = JCM 3896T). An emended description of the species Sphingomonas parva is provided.

A natural occurring bifunctional CPD/(6-4)-photolyase from the Antarctic bacterium Sphingomonas sp. UV9.

Photolyases are flavoproteins that repair ultraviolet-induced DNA lesions (cyclobutane pyrimidine dimer or CPD, and pyrimidine (6-4) pyrimidone photoproducts or (6-4)-PPs), using blue light as an energy source. These enzymes are substrate specific, meaning that a specific photolyase repairs either a CPD or a (6-4)-PP. In this work, we produced a class II CPD-photolyase (called as PhrSph98) from the Antarctic bacterium Sphingomonas sp. UV9 by recombinant DNA technology and we purified the enzyme using immobilized metal affinity chromatography. By using an immunochemistry assay, with monoclonal antibodies against CPD and (6-4)-PP, we found that PhrSph98 repairs both DNA lesions. The result was confirmed by immunocytochemistry using immortalized non-tumorigenic human keratinocytes. Results from structure prediction, pocket computation, and molecular docking analyses showed that PhrSph98 has the two expected protein domains (light-harvesting antenna and a catalytic domain), a larger catalytic site as compared with photolyases produced by mesophilic organisms, and that both substrates fit the catalytic domain. The results obtained from predicted homology modeling suggest that the electron transfer pathway may occur following this pathway: Y389-W369-W390-F376-W381/FAD. The evolutionary reconstruction of PhrSph98 suggests that this is a missing link that reflects the transition of (6-4)-PP repair into the CPD repair ability for the class II CPD-photolyases. To the best of our knowledge, this is the first report of a naturally occurring bifunctional, CPD and (6-4)-PP, repairing enzyme. KEY POINTS: • We report the first described bifunctional CPD/(6-4)-photoproducts repairing enzyme. The bifunctional enzyme reaches the nuclei of keratinocyte and repairs the UV-induced DNA damage. The enzyme should be a missing link from an evolutionary point of view. The enzyme may have potential uses in the pharmaceutical and cosmetic industries.

Comprehensive Genome Analysis on the Novel Species Sphingomonas panacis DCY99T Reveals Insights into Iron Tolerance of Ginseng.

Plant growth-promoting rhizobacteria play vital roles not only in plant growth, but also in reducing biotic/abiotic stress. Sphingomonas panacis DCY99T is isolated from soil and root of Panax ginseng with rusty root disease, characterized by raised reddish-brown root and this is seriously affects ginseng cultivation. To investigate the relationship between 159 sequenced Sphingomonas strains, pan-genome analysis was carried out, which suggested genomic diversity of the Sphingomonas genus. Comparative analysis of S. panacis DCY99T with Sphingomonas sp. LK11 revealed plant growth-promoting potential of S. panacis DCY99T through indole acetic acid production, phosphate solubilizing, and antifungal abilities. Detailed genomic analysis has shown that S. panacis DCY99T contain various heavy metals resistance genes in its genome and the plasmid. Functional analysis with Sphingomonas paucimobilis EPA505 predicted that S. panacis DCY99T possess genes for degradation of polyaromatic hydrocarbon and phenolic compounds in rusty-ginseng root. Interestingly, when primed ginseng with S. panacis DCY99T during high concentration of iron exposure, iron stress of ginseng was suppressed. In order to detect S. panacis DCY99T in soil, biomarker was designed using spt gene. This study brings new insights into the role of S. panacis DCY99T as a microbial inoculant to protect ginseng plants against rusty root disease.

Cloning and characterization of a thermostable glutathione reductase from a psychrophilic Arctic bacterium Sphingomonas sp.

Glutathione reductase is an important oxidoreductase that helps maintain redox homeostasis by catalyzing the conversion of glutathione disulfide to glutathione using NADPH as a cofactor. In this study, we cloned and characterized a glutathione reductase (hereafter referred to as SpGR) from Sphingomonas sp. PAMC 26621, an Arctic bacterium. SpGR comprises 449 amino acids, and functions as a dimer. Surprisingly, SpGR exhibits characteristics of thermophilic enzymes, showing optimum activity at 60°C and thermal stability up to 70°C with ∼50% residual activity at 70°C for 2 h. The amino acid composition analysis of SpGR showed a 1.9-fold higher Arg content (6%) and a 2.7-fold lower Lys/Arg ratio (0.75) compared to the Arg content (3.15%) and the Lys/Arg ratio (2.01) of known psychrophilic glutathione reductases. SpGR also exhibits its activity at 4°C, and circular dichroism and fluorescence spectroscopy results indicate that SpGR maintains its secondary and tertiary structures within the temperature range of 4-70°C. Taken together, the results of this study indicate that despite its origin from a psychrophilic bacterium, SpGR has high thermal stability. Our study provides an insight into the role of glutathione reductase in maintaining the reducing power of an Arctic bacterium in a broad range of temperatures.

Comparative genome analysis reveals the evolution of chloroacetanilide herbicide mineralization in Sphingomonas wittichii DC-6.

The environmental fate of the extensively used chloroacetanilide herbicides (CH) has been a cause of increasing concern in the past decade because of their carcinogenic properties. Although microbes play important roles in CH degradation, Sphingomonas wittichii DC-6 was the first reported CH-mineralizing bacterium. In this study, the complete genome of strain DC-6 was sequenced and comparative genomic analysis was performed using strain DC-6 and other three partial CH-degrading bacteria, Sphingobium quisquiliarum DC-2, Sphingobium baderi DE-13, and Sphingobium sp. MEA3-1. 16S rDNA phylogenetic analysis indicated that strain DC-2, MEA3-1, and DE-13 are closely related and DC-6 has relatively distant genetic relationship with the other three strains. The identified CH degradation genes responsible for the upstream and downstream pathway, including cndA, cmeH, meaXY, and meaAB, were all located in conserved DNA fragments (or genetic islands) in the vicinity of mobile element proteins. Protein BLAST in the NCBI database showed that cndA and cmeH were present in the genomes of other sequenced strains isolated from various habitats; however, the gene compositions in these host strains were completely different from those of other sphingomonads, and codon usage of genes for upstream pathway were also different from that of downstream pathway. These results showed that the upstream and downstream pathways of CH degradation in strain DC-6 have evolved by horizontal gene transfer and gene combination. In addition, the genes of the ring-cleavage pathway were not conserved and may have evolved directly from bacterial degradation of hydroxyquinol. The present study provides insights into the evolutionary strategy and microbial catabolic pathway of CH mineralization.