Novosphingobium naphthalenivorans sp. nov., a naphthalene-degrading bacterium isolated from polychlorinated-dioxin-contaminated environments

Three strains of strictly aerobic, Gram-negative, naphthalene-degrading bacteria isolated from polychlorinated-dioxin-contaminated soil and sediment were characterized. These isolates grew well with naphthalene as the sole carbon and energy source, degrading it completely within 24 h of incubation. The isolates also degraded dibenzofuran co-metabolically in the presence of naphthalene with the concomitant production of yellow intermediate metabolite(s). A 16S rRNA gene sequence analysis revealed that the isolates affiliated to the genus Novosphingobium with Novosphingobium pentaromativorans and Novosphingobium subarcticum as their nearest phylogenetic neighbors (97.4-97.5% similarity). The isolates had a genomic DNA G+C ratio of 64.5-64.6 mol% and formed a genetically coherent group distinguishable from any established species of the genus Novosphingobium at a DNA-DNA hybridization level of less than 46%. The cellular fatty acids were characterized by the predominance of 18 : 1omega7c with significant proportions of 16 : 0, 16 : 1omega7c, 17 : 1omega6c and 2-OH 14 : 0. Sphingoglycolipids were present. The major respiratory quinone was ubiquinone-10. Spermidine was detected as the major polyamine. The distinct taxonomic position of the isolates within the Novosphingobium was also demonstrated by physiological and biochemical testing. Based on these phylogenetic and phenotypic data, we propose Novosphingobium naphthalenivorans sp. nov. to accommodate the novel isolates. The type strain is strain TUT562(T) (DSM 18518(T), JCM 13951(T), NBRC 102051(T)).     

Isolation and characterization of Novosphingobium sp. strain MT1, a dominant polychlorophenol-degrading strain in a groundwater bioremediation system.

A high-rate fluidized-bed bioreactor has been treating polychlorophenol-contaminated groundwater in southern Finland at 5 to 8 degrees C for over 6 years. We examined the microbial diversity of the bioreactor using three 16S ribosomal DNA (rDNA)-based methods: denaturing gradient gel electrophoresis, length heterogeneity-PCR analysis, and restriction fragment length polymorphism analysis. The molecular study revealed that the process was dependent on a stable bacterial community with low species diversity. The dominant organism, Novosphingobium sp. strain MT1, was isolated and characterized. Novosphingobium sp. strain MT1 degraded the main contaminants of the groundwater, 2,4,6-trichlorophenol, 2,3,4,6-tetrachlorophenol, and pentachlorophenol, at 8 degrees C. The strain carried a homolog of the pcpB gene, coding for the pentachlorophenol-4-monooxygenase in Sphingobium chlorophenolicum. Spontaneous deletion of the pcpB gene homolog resulted in the loss of degradation ability. Phenotypic dimorphism (planktonic and sessile phenotypes), low growth rate (0.14 to 0.15 h(-1)), and low-copy-number 16S rDNA genes (single copy) were characteristic of strain MT1 and other MT1-like organisms isolated from the bioreactor.     

<Emphasis Type="Italic">Novosphingobium profundi</Emphasis> sp. nov. isolated from a deep-sea seamount

A marine bacterial strain, F72^T, was isolated from a solitary scleractinian coral, collected in Yap seamounts in the Pacific Ocean. Strain F72^T is a Gram-negative, light-yellow-pigmented, motile, rod-shaped bacterium. Phylogenetic analysis based on 16S rRNA gene sequences showed that strain F72^T is related to the genus Novosphingobium and has high 16S rRNA gene sequence similarities with the type strains of Novosphingobium pentaromativorans US6-1^T (97.7 %), Novosphingobium panipatense SM16^T (97.6 %), Novosphingobium mathurense SM117^T (97.2 %) and Novosphingobium barchaimii LL02^T (97.1 %). Ubiquinone Q-10 was detected as the dominant quinone. The predominant cellular fatty acids were C_18:1ω7c and C_17:1ω6c. The genomic DNA G+C content of strain F72^T was 63.4 mol %. The polar lipids profile contained phosphatidylglycerol, diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylmethylethanolamine, phosphatidylcholine, sphingoglycolipid and one uncharacterized lipid. Strain F72^T shared DNA relatedness of 25 % with N. pentaromativorans JCM 12182^T, 31 % with N. panipatense DSM 22890^T, 21 % with N. mathurense DSM 23374^T and 26 % with N. barchaimii DSM 25411^T. Combined data from phenotypic, phylogenetic and DNA–DNA relatedness studies demonstrated that the strain F72^T is a representative of a novel species of the genus Novosphingobium, for which we propose the name Novosphingobium profundi sp. nov. (type strain F72^T = KACC 18566^T = CGMCC 1.15390^T).     

Draft genome sequence of Novosphingobium nitrogenifigens Y88(T)

Novosphingobium nitrogenifigens was originally isolated from pulp and paper mill wastewater, a low-nitrogen, high-carbon environment. N. nitrogenifigens is the first known nitrogen-fixing, polyhydroxyalkanoate-accumulating sphingomonad, and we report the annotated draft genome sequence of the type strain Y88(T) here.     

De novo sequencing and assembly of the whole genome of Novosphingobium sp. strain PP1Y

Novosphingobium sp. strain PP1Y is a marine bacterium specifically adapted to use fuels as an energy source. We sequenced and assembled its entire genome using the Roche 454 genome sequencer system, which led to the identification of two plasmids and one megaplasmid, besides a 3.9-Mb circular chromosome.     

大西洋洋中脊深海多环芳烃降解菌群的优势菌分析

摘要：【目的】为了分析大西洋洋中脊深海海水及表层沉积物中多环芳烃(PAHs)降解菌群中的优势菌。【方法】采用富集培养法和平板涂布法从深海样品中分离可培养细菌及PAHs降解菌。通过16S rRNA基因测序完成系统发育分析。采用变性梯度凝胶电泳（DGGE）及DNA测序分析降解菌群中的优势菌。【结果】总共分离到16株细菌,包括一株PAHs降解菌Novosphingobium sp. 4D。系统发育分析发现,可培养细菌中两个最大的类群分别与Alcanivorax dieselolei NO1A（5/16）和Tistrella mobilis TISTR 1108T（5/16）亲缘关系最近。DGGE结果表明,在菌群MC2D中菌株4L（以及4M、4N, Alcanivorax dieselolei NO1A, 99.21％）、4D（Novosphingobium pentaromativorans US6-1T,97.07％）和4B（以及4E、4H、4K,Tistrella mobilis TISTR 1108T,＞99％）是降解菌群中的优势菌。而降解菌群MC3CO中的优势菌是菌株5C（以及5H,Alcanivorax dieselolei NO1A,＞99％）、条带5-8代表的未培养菌株（Novosphingobium aromaticivorans DSM 12444T,99.41%）、5J（Tistrella mobilis TISTR 1108T,99.52％）和5F（以及5G,Thalassospira lucentensis DSM 14000T,＜97％）。【结论】本研究发现在大西洋洋中脊深海海水及表层沉积物中Alcanivorax、Novosphingobium、Thalassospira、Tistrella属的细菌是PAHs降解菌群中的优势菌,其中的主要降解菌是Novosphingobium属的细菌。     

Genome sequence of benzo(a)pyrene-degrading bacterium Novosphingobium pentaromativorans US6-1

Novosphingobium pentaromativorans US6-1 showed a good ability to degrade high-molecular-weight polycyclic aromatic hydrocarbons. We report the draft genome sequence of strain US6-1, which contains a main chromosome (5,096,413 bp, G+C content of 63.1%) and two plasmids (188,476 and 60,085 bp). The majority of the aromatic-hydrocarbon-degrading genes are encoded in the larger plasmid.     

Novosphingobium hassiacum sp. nov., a new species isolated from an aerated sewage pond

The taxonomy of two strains W-51T and W-52 isolated from a wastewater treatment plant was investigated in a polyphasic approach. The yellow pigmented gram-negative organism contained a quinone system with mainly ubiquinone Q-10, and the polar lipid profile contained a sphingoglycolipid suggesting that both strains belonged to the the alpha-4 subclass of the Proteobacteria. The polar lipid profile consisted furthermore of phosphatidylethanolamine, diphosphatidylglycerol, and phosphatidylcholine and of minor amounts of phosphatidylglycerol and phosphatidylmonomethylethanolamine. Sequencing of the 16S rRNA gene supported the allocation into the genus Novosphingobium, together with the type strains of N. subterraneum, N. aromaticivorans, N. stygium, and N. capsulatum, showing similarities of 97.3%, 97.0%, 95.7% and 96.2%, respectively. This allocation was supported by the polyamine profile, which consisted mainly of spermidine. The analysis of the fatty acids revealed 2-OH 13:0, 2-OH 14:0 and 2-OH 15:0, with 2-OH 15:0 as predominant hydroxylated fatty acid. W-51T and W-52 were almost identical with respect to their phenotypic including the majority of the chemotaxonomic properties, identical in their 16S rRNA sequences, and showed 86% DNA-DNA similarity. Both strains were able to reduce nitrate and on the basis of further physiological features, a clear differentiation from all other Novosphingobium species was possible. The DNA-DNA similarities of W-51T to the type strains of N. subterraneum, N. aromaticivorans, and N. capsulatum were below 56%. For these reasons, it is proposed to create a new species with the name Novosphingobium hassiacum sp. nov.     

Pacificamonas flava gen. nov., sp. nov., a Novel Member of the Family Sphingomonadaceae Isolated from the Southeastern Pacific

Strain JLT2015^T was isolated from surface seawater of the Southeastern Pacific. The strain was Gram-negative, aerobic, motile by gliding, and rod shaped. The dominant fatty acids were C_18:1ω7c, C_16:0, and C_16:1ω7c. The major respiratory ubiquinone was Q-10, and the predominant polyamine pattern was spermidine. The components of the polar lipid profile were phosphatidylethanolamine, phosphatidylglycerol, diphosphatidylglycerol, and sphingoglycolipid. The DNA G+C content was 64.2 %. Phylogenetic analysis based on 16S rRNA gene sequence revealed strain JLT2015^T belonged to belong to the family Sphingomonadaceae, exhibiting 94.7 % 16S rRNA gene sequence similarity with Novosphingobium pentaromativorans. On the basis of the taxonomic data presented, together with phylogenetic and genetic characteristics, strain JLT2015^T is considered to represent a novel genus, for which the name Pacificamonas flava gen. nov., sp. nov. is proposed. The type strain of Pacificamonas flava is JLT2015^T (=LMG27364^T = CGMCC1.12401^T).     

The unique aromatic catabolic genes in sphingomonads degrading polycyclic aromatic hydrocarbons (PAHs)

Many members of the sphingomonad genus isolated from different geological areas can degrade a wide variety of polycyclic aromatic hydrocarbons (PAHs) and related compounds. These sphingomonads such as Sphingobium yanoikuyae strain B1, Novosphingobium aromaticivorans strain F199, and Sphingobium sp. strain P2 have been found to possess a unique group of genes for aromatic degradation, which are distantly related with those in pseudomonads and other genera reported so far both in sequence homology and gene organization. Genes for aromatics degradation in these sphingomonads are complexly arranged; the genes necessary for one degradation pathway are scattered through several clusters. These aromatic catabolic gene clusters seem to be conserved among many other sphingomonads such as Sphingobium yanoikuyae strain Q1, Sphingomonas paucimobilis strain TNE12, S. paucimobilis strain EPA505, Sphingobium agrestis strain HV3, and Sphingomonas chungbukensis strain DJ77. Furthermore, some genes for naphthalenesulfonate degradation found in Sphingomonas xenophaga strain BN6 also share a high sequence homology with their homologues found in these sphingomonads. On the other hand, protocatechuic catabolic gene clusters found in fluorene-degrading Sphingomonas sp. strain LB126 appear to be more closely related with those previously found in lignin-degrading S. paucimobilis SYK-6 than the genes in this group of sphingomonads. This review summarizes the information on the distribution of these strains and relationships among their aromatic catabolic genes.     

Biological control and plant growth promoting capacity of rhizobacteria on pepper under greenhouse and field conditions

Plant growth promoting rhizobacteria Ochrobactrum lupini KUDC1013 and Novosphingobium pentaromativorans KUDC1065 isolated from Dokdo Island, S. Korea are capable of eliciting induced systemic resistance (ISR) in pepper against bacterial spot disease. The present study aimed to determine whether plant growth-promoting rhizobacteria (PGPR) strains including strain KUDC1013, strain KUDC1065, and Paenibacillus polymyxa E681 either singly or in combinations were evaluated to have the capacity for potential biological control and plant growth promotion effect in the field trials. Under greenhouse conditions, the induced systemic resistance (ISR) effect of treatment with strains KUDC1013 and KUDC1065 differed according to pepper growth stages. Drenching of 3-week-old pepper seedlings with the KUDC-1013 strain significantly reduced the disease symptoms. In contrast, treatment with the KUDC1065 strain significantly protected 5-week-old pepper seedlings. Under field conditions, peppers treated with PGPR mixtures containing E681 and KUDC1013, either in a two-way combination, were showed greater effect on plant growth than those treated with an individual treatment. Collectively, the application of mixtures of PGPR strains on pepper might be considered as a potential biological control under greenhouse and field conditions.     

Characterization of polyhydroxyalkanoates (PHAs) biosynthesis by isolated Novosphingobium sp. THA_AIK7 using crude glycerol

Biodiesel-contaminated wastewater was used to screen for PHAs-producing bacteria by using crude glycerol as the sole carbon source. A gram-negative THA_AIK7 isolate was chosen as a potential PHAs producer. The 16S rRNA phylogeny indicated that THA_AIK7 isolate is a member of Novosphingobium genus which is supported by a bootstrap percentage of 100% with Novosphingobium capsulatum. The 1,487 bp of 16S rRNA gene sequence of THA_AIK7 isolate has been deposited in the GenBank database under the accession number HM031593. Polymer content of 45% cell dry weight was achieved in 72 h with maximum product yield coefficient of 0.29 g PHAs g?1 glycerol. Transmission electron micrograph results exhibited the PHAs granules accumulated inside the bacterial cell. PHAs polymer production in mineral salt media supplemented with 2% (w/v) of crude glycerol at initial pH 7 was extracted by the sodium hypochlorite method. Polymer film spectrographs from Nuclear magnetic resonance displayed a pattern of signal virtually identical to spectra of commercial PHB. Thermal analysis by Differential scanning calorimeter showed a melting temperature at 179°C. Molecular weight analysis by Gel permeation chromatography showed two main peaks of 133,000 and 700 g mol?1 with weight-average molecular weight value of 23,800 and number-average molecular weight value of 755. Endotoxinfree of PHAs polymer was preliminarily assessed by a negative result of the gel-clot formation, Pyrotell? Single test vial, at sensitivity of 0.25 EU ml?1. To our knowledge, this is the first reported test of endotoxin-free PHAs naturally produced from gram-negative bacteria which could be used for biomedical application.     

Starvation- and xenobiotic-related transcriptomic responses of the sulfanilic acid-degrading bacterium,Novosphingobium resinovorum SA1

Applied Microbiology and Biotechnology - Novosphingobium resinovorum SA1 was the first single isolate capable of degrading sulfanilic acid, a widely used representative of sulfonated aromatic...     

A pyrene-degrading consortium from deep-sea sediment of the West Pacific and its key member Cycloclasticus sp. P1

A pyrene-degrading bacterial consortium was obtained from deep-sea sediments of the Pacific Ocean. The consortium degraded many kinds of polycyclic aromatic hydrocarbons (PAHs), including naphthalene, phenanthrene, pyrene, acenaphthene, fluorene, anthracene, fluoranthene, 2-methylnaphthalene and 2,6-dimethylnaphthalene, but it did not grow with chrysene and benzo[alpha]pyrene. With methods of plate cultivation and polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE), 72 bacteria belonging to 22 genera were detected from this consortium. Among the detected bacteria, the following genera frequently occurred: Flavobacterium, Cycloclasticus, Novosphingobium, Halomonas, Achromobacter, Roseovarius and Alcanivorax. The first two genera showed the strongest bands in denaturing gradient gel electrophoresis (DGGE) profiles and appeared in all PAH treatments. By now, only one isolate designated P1 was confirmed to be a pyrene degrader. It was identified to be Cycloclasticus spirillensus (100%). Although P1 can degrade pyrene independently, other bacteria, such as Novosphingobium sp. (Band 14), Halomonas sp. (Band 16) and an unidentified bacterium (Band 35), were involved in pyrene degradation in some way; they persist in the consortium in the test of dilution to extinction if only the consortium was motivated with pyrene. However, the secondary most important member Flavobacterium sp. evaded from the community at high dilutions. As a key member of the consortium, P1 distinguished itself by both cell morphology and carbon source range among the isolates of this genus. Based on intermediate analyses of pyrene degradation, P1 was supposed to take an upper pathway different from that previously reported. Together with the results of obtained genes from P1 homology with those responsible for naphthalene degradation, its degradation to pyrene is supposed to adopt another set of genes unique to presently detected. Summarily, an efficient pyrene-degrading consortium was obtained from the Pacific Ocean sediment, in which Cycloclasticus bacterium played a key role. This is the first report to exploit the diversity of pyrene-degrading bacteria in oceanic environments.     

深海多环芳烃降解菌新鞘氨醇杆菌H25的降解特性及降解基因

[目的]为了从深海环境中筛选新的多环芳烃降解菌,了解其降解基因及降解特性.[方法]以原油作为碳源从印度洋深海海水样品中富集筛选出降解能力较强的多环芳烃降解菌,并根据已报道的相关菌属的多环芳烃起始双加氧酶大亚基序列及侧翼序列设计兼并引物进行扩增.[结果]获得了1株能够高效降解原油、柴油及多种多环芳烃的菌株H25.经16S rDNA序列系统发育分析表明它属于新鞘氨醇杆菌属(Novosphingobium)(96%).并从该菌株中扩增获得2条相似度为91.0%双加氧酶基因片段.2条序列在NCBI上Blastn分析表明均与菌株N.aromaticivorans DSM12444T的降解质粒pNL1上的双加氧酶大亚基具有最高相似度,分别为99.6%和91.0%.根据pNL1上的双加氧酶序列设计引物获得了包含H25双加氧酶大亚基及上下游序列的2个基因片段H25 Ⅰ(2.9kb)和H25Ⅱ(4.5kb).另外,单碳降解实验表明H25对联苯、2-甲基萘、2,6-二甲基萘、菲、二苯并噻吩、二苯并呋喃等均有较好的降解能力.[结论]H25菌株是Novosphingobium属可能的新种.深海细菌在大洋环境多环芳烃污染的自然净化中起到一定作用,并在环境生物修复中有较大的应用前景.     

Engineering of the cytochrome P450 monooxygenase system for benzyl maltol hydroxylation

Maltol derivatives are utilized in a variety of fields due to their metal-chelating abilities, and modification of the 2-methyl side chain is known to effectively expand their functional diversity. In the present study, microbial enzymes were screened for hydroxylating activity towards the 2-methyl group in a maltol derivative, 3-benzyloxy-2-methyl-4-pyrone (BMAL). Novosphingobium sp. SB32149 was found to have the ability to convert BMAL into 3-benzyloxy-2-hydroxymethyl-4-pyrone (BMAL-OH). The enzymes responsible, a cytochrome P450 monooxygenase (P450nov), a ferredoxin (FDXnov), and a ferredoxin reductase (FDRnov), were identified in the SB32149 strain. In the reaction with recombinant Escherichia coli expressing P450nov, FDXnov, and FDRnov, BMAL-OH was successfully produced from BMAL. Moreover, using the directed evolution approach, four amino acid substitutions, L188P/F218L/L237M in P450nov and A10T in FDXnov, were found to enhance BMAL-OH production. Consequently, up to 5.2 g/L BMAL-OH was obtained from 8.0 g/L BMAL by bioconversion using a 250-mL jar fermenter, indicating that this strain may be useful for synthesis of maltol derivatives which could have potential applications in various fields.     

碳、氮、磷源生态因子对新鞘氨醇杆菌降解微囊藻毒素的影响及分子机理

【背景】微藻囊毒素(Microcystins,MCs)是蓝藻暴发后产生的主要有害物质之一,在河口、湖泊和近海水体中均存在。【目的】研究水体营养物质浓度对新鞘氨醇杆菌(Novo sphingobium sp.ERN07)降解能力的影响,以求达到快速降解MCs的目的。【方法】通过正交实验、基因转录分析等方法鉴定菌株降解特性,了解生物降解和功能基因的表达关系,认识其降解机理。【结果】菌株去除MCs的能力受水体中碳、氮和磷源浓度的影响,由碳、氮源组成的双营养物质对生物降解抑制作用比单源弱;与标准MSM培养基相比,添加碳源、氮源和减少磷源会抑制生物降解MCs,表现为降解基因表达下调。不同营养条件下,mlr在转录水平上的基因表达不同,从而产生不同的降解效果。【结论】Novo sphingobium sp. ERN07具有较高的MCs降解能力,在水华发生之前能有效去除水环境中碳源和氮源,可以应用于治理受MCs污染的水体。     

Degradation of 17beta-estradiol by a gram-negative bacterium isolated from activated sludge in a sewage treatment plant in Tokyo, Japan

A 17beta-estradiol (E2)-degrading bacterium was isolated from activated sludge in a sewage treatment plant in Tokyo, Japan. The isolate was suggested to be a new Novosphingobium species. Gas chromatography-mass spectrometry and (1)H nuclear magnetic resonance analyses of the metabolites of E2 degradation suggested that no toxic products accumulated in the culture medium.     

Prokaryotic origins for the mitochondrial alternative oxidase and plastid terminal oxidase nuclear genes

The mitochondrial alternative oxidase is a diiron carboxylate quinol oxidase (Dox) found in plants and some fungi and protists, but not animals. The plastid terminal oxidase is distantly related to alternative oxidase and is most likely also a Dox protein. Database searches revealed that the alpha-proteobacterium Novosphingobium aromaticivorans and the cyanobacteria Nostoc sp. PCC7120, Synechococcus sp. WH8102 and Prochlorococcus marinus subsp. pastoris CCMP1378 each possess a Dox homolog. Each prokaryotic protein conforms to the current structural models of the Dox active site and phylogenetic analyses suggest that the eukaryotic Dox genes arose from an ancestral prokaryotic gene.