Complete genome sequence of Sphingobium sp. strain PAMC 28499 reveals a potential for degrading pectin with comparative genomics approach.

BACKGROUND: Spingobium sp. PAMC 28499 is isolated from the glaciers of Uganda. Uganda is a unique region where hot areas and glaciers coexist, with a variety of living creatures surviving, but the survey on them is very poor. The genetic character and complete genome information of Sphingobium strains help with environmental studies and the development of better to enzyme industry. OBJECTIVE: In this study, complete genome sequence of Spingobium sp. PAMC 28499 and comparative analysis of Spingobium species strains isolated from variety of the region. METHODS: Genome sequencing was performed using PacBio sequel single-molecule real-time (SMRT) sequencing technology. The predicted gene sequences were functionally annotated and gene prediction was carried out using the program NCBI non-redundant database. And using dbCAN2 and KEGG data base were degradation pathway predicted and protein prediction about carbohydrate active enzymes (CAZymes). RESULTS: The genome sequence has 64.5% GC content, 4432 coding protein coding genes, 61 tRNAs, and 12 rRNA operons. Its genome encodes a simple set of metabolic pathways relevant to pectin and its predicted degradation protein an unusual distribution of CAZymes with extracellular esterases and pectate lyases. CAZyme annotation analyses revealed 165 genes related to carbohydrate active, and especially we have found GH1, GH2, GH3, GH38, GH35, GH51, GH51, GH53, GH106, GH146, CE12, PL1 and PL11 such as known pectin degradation genes from Sphingobium yanoikuiae. These results confirmed that this Sphingobium sp. strain PAMC 28499 have similar patterns to RG I pectin-degrading pathway. CONCLUSION: In this study, isolated and sequenced the complete genome of Spingobium sp. PAMC 28499. Also, this strain has comparative genome analysis. Through the complete genome we can predict how this strain can store and produce energy in extreme environment. It can also provide bioengineered data by finding new genes that degradation the pectin.

Development of the production of 2-pyrone-4,6-dicarboxylic acid from lignin extracts, which are industrially formed as by-products, as raw materials.

Lignosulfonate is a by-product of the cooking process by sulfite pulping for paper manufacturing. The treatment of wood chips by various salts of sulfurous acid solubilizes lignin to produce a cellulose-rich wood pulp. Developing a technique for the conversion of lignosulfonate by-product to high value materials has an important industrial utility. Sphingobium sp. strain SYK-6, which was isolated from pulping wastewater, is one of the best enzymatically or genetically characterized bacteria for degrading lignin-derived aromatics. We have previously established a system for the production of 2-pyrone-4,6-dicarboxylic acid (PDC), a novel platform chemical that can produce a variety of bio-based polymers, by introducing of ligA, ligB, and ligC genes from SYK-6 into a mutant strain of Pseudomonas putida PpY1100. In this study, extracts from lignosulfonates, which were desulphonated and depolymerized by alkaline oxidation, were evaluated as substrates for microbiological conversion to PDC by the transgenic bacteria.

Biodegradation of Tetralin: Genomics, Gene Function and Regulation.

Tetralin (1,2,3,4-tetrahydonaphthalene) is a recalcitrant compound that consists of an aromatic and an alicyclic ring. It is found in crude oils, produced industrially from naphthalene or anthracene, and widely used as an organic solvent. Its toxicity is due to the alteration of biological membranes by its hydrophobic character and to the formation of toxic hydroperoxides. Two unrelated bacteria, Sphingopyxis granuli strain TFA and Rhodococcus sp. strain TFB were isolated from the same niche as able to grow on tetralin as the sole source of carbon and energy. In this review, we provide an overview of current knowledge on tetralin catabolism at biochemical, genetic and regulatory levels in both strains. Although they share the same biodegradation strategy and enzymatic activities, no evidences of horizontal gene transfer between both bacteria have been found. Moreover, the regulatory elements that control the expression of the gene clusters are completely different in each strain. A special consideration is given to the complex regulation discovered in TFA since three regulatory systems, one of them involving an unprecedented communication between the catabolic pathway and the regulatory elements, act together at transcriptional and posttranscriptional levels to optimize tetralin biodegradation gene expression to the environmental conditions.

Colonization and Maize Growth Promotion Induced by Phosphate Solubilizing Bacterial Isolates.

Phosphorus (P) limits the production of maize, one of the major food crops in China. Phosphate-solubilizing bacteria (PSB) have the capacity to solubilize phosphate complexes into plant absorbable and utilizable forms by the process of acidification, chelation, and exchange reactions. In this study, six bacteria, including one Paenibacillus sp. B1 strain, four Pseudomonas sp. strains (B10, B14, SX1, and SX2) and one Sphingobium sp. SX14 strain, were those isolated from the maize rhizosphere and identified based on their 16S rRNA sequences. All strains could solubilize inorganic P (Ca3(PO4)2, FePO4 and AlPO4), and only B1 and B10 organic P (lecithin). All strains, except of SX1, produced IAA, and SX14 and B1 showed the highest level. B1 incited the highest increase in root length and the second increase in shoot and total dry weight, shoot length, and total P and nitrogen (N), along with increased root length. In addition, by confocal laser scanning microscopy (CLSM), we found that green fluorescent protein (GFP)-labeled B1 mainly colonized root surfaces and in epidermal and cortical tissue. Importantly, B1 can survive through forming spores under adverse conditions and prolong quality guarantee period of bio-fertilizer. Therefore, it can act as a good substitute for bio-fertilizer to promote agricultural sustainability.

11-Hydroxylation of Protoberberine by the Novel Berberine-Utilizing Aerobic Bacterium Sphingobium sp. Strain BD3100.

Protoberberine alkaloids, including berberine, palmatine, and berberrubine, are produced by medicinal plants and are known to have various pharmacological effects. We isolated two berberine-utilizing bacteria, Sphingobium sp. strain BD3100 and Rhodococcus sp. strain BD7100, from soil collected at a natural medicine factory. BD3100 had the unique ability to utilize berberine or palmatine as the sole carbon and energy source. BD3100 produced demethyleneberberine in berberine-supplemented medium. In a resting-cell incubation with berberine, BD3100 produced 11-hydroxyberberine; the structure of 11-hydroxyberberine was determined by detailed analysis of NMR and MS spectroscopic data. α-Naphthoflavone, miconazole, and ketoconazole, which are known inhibitors of cytochrome P450, interfered with BD3100 metabolism of berberine in resting cells. Inhibition by miconazole led to the production of a new compound, 11-hydroxydemethyleneberberine. In a resting-cell incubation with palmatine, BD3100 generated 11-hydroxypalmatine. This work represents the first report of the isolation and characterization of novel berberine-utilizing aerobic bacteria for the production of 11-hydroxylation derivatives of berberine and palmatine.

Tributyl phosphate biodegradation to butanol and phosphate and utilization by a novel bacterial isolate, Sphingobium sp. strain RSMS.

A Sphingobium sp. strain isolated from radioactive solid waste management site (RSMS) completely degraded 7.98 g/L of tributyl phosphate (TBP) from TBP containing suspensions in 3 days. It also completely degraded 20 mM dibutyl phosphate (DBP) within 2 days. The strain tolerated high levels of TBP and showed excellent stability with respect to TBP degradation over several repeated subcultures. On solid minimal media or Luria Bertani media supplemented with TBP, the RSMS strain showed a clear zone of TBP degradation around the colony. Gas chromatography and spectrophotometry analyses identified DBP as the intermediate and butanol and phosphate as the products of TBP biodegradation. The RSMS strain utilized both TBP and DBP as the sole source of carbon and phosphorous for its growth. The butanol released was completely utilized by the strain as a carbon source thereby leaving no toxic residue in the medium. Degradation of TBP or DBP was found to be suppressed by high concentration of glucose which also inhibited TBP or DBP dependent growth. The results highlight the potential of Sphingobium sp. strain RSMS for bioremediation of TBP and for further molecular investigation.

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.

Enhanced biodegradation of hexachlorocyclohexane (HCH) in contaminated soils via inoculation with Sphingobium indicum B90A.

Soil pollution with hexachlorocyclohexane (HCH) has caused serious environmental problems. Here we describe the targeted degradation of all HCH isomers by applying the aerobic bacterium Sphingobium indicum B90A. In particular, we examined possibilities for large-scale cultivation of strain B90A, tested immobilization, storage and inoculation procedures, and determined the survival and HCH-degradation activity of inoculated cells in soil. Optimal growth of strain B90A was achieved in glucose-containing mineral medium and up to 65% culturability could be maintained after 60 days storage at 30 degrees C by mixing cells with sterile dry corncob powder. B90A biomass produced in water supplemented with sugarcane molasses and immobilized on corncob powder retained 15-20% culturability after 30 days storage at 30 degrees C, whereas full culturability was maintained when cells were stored frozen at -20 degrees C. On the contrary, cells stored on corncob degraded gamma-HCH faster than those that had been stored frozen, with between 15 and 85% of gamma-HCH disappearance in microcosms within 20 h at 30 degrees C. Soil microcosm tests at 25 degrees C confirmed complete mineralization of [(14)C]-gamma-HCH by corncob-immobilized strain B90A. Experiments conducted in small pits and at an HCH-contaminated agricultural site resulted in between 85 and 95% HCH degradation by strain B90A applied via corncob, depending on the type of HCH isomer and even at residual HCH concentrations. Up to 20% of the inoculated B90A cells survived under field conditions after 8 days and could be traced among other soil microorganisms by a combination of natural antibiotic resistance properties, unique pigmentation and PCR amplification of the linA genes. Neither the addition of corncob nor of corncob immobilized B90A did measurably change the microbial community structure as determined by T-RFLP analysis. Overall, these results indicate that on-site aerobic bioremediation of HCH exploiting the biodegradation activity of S. indicum B90A cells stored on corncob powder is a promising technology.

Sandarakinorhabdus limnophila gen. nov., sp. nov., a novel bacteriochlorophyll a-containing, obligately aerobic bacterium isolated from freshwater lakes.

Three strains (so36, so42T and wo26) representing a novel Gram-negative, obligately aerobic, bacteriochlorophyll a-containing species of the alpha-4 subgroup of the Proteobacteria were isolated from freshwater lakes using a high-throughput cultivation technique. The non-motile and slender rod-shaped cells formed orange-red-pigmented colonies. The main carotenoids were nostoxanthin and keto-nostoxanthin. According to the absorption spectrum, two different photosynthetic light-harvesting complexes, an LHI complex and a B800-830-type peripheral LHII complex, were present in the cells. The predominant fatty acids of strain so42T were hexadecenoic acid (16 : 1omega7c) and octadecenoic acid (18 : 1omega7c), whereas 17 : 1omega6c and 14 : 0 iso 2-OH were present in smaller amounts. The main polar lipids were phosphatidylethanolamine, phosphatidylglycerol, phosphatidylcholine, diphosphatidylglycerol, glycolipid and sphingoglycolipids. The major respiratory lipoquinone was ubiquinone-10, whereas ubiquinone-9 was present in smaller amounts. The three strains were cytochrome oxidase-negative and catalase-positive and formed alkaline and acid phosphatases. The strains grew chemoorganoheterotrophically in mineral media supplemented with various organic acids, amino acids or complex substrates such as peptone and yeast extract. The G+C content of the genomic DNA of strain so42T was 64.3 mol%. The three novel isolates contained the same 16S rRNA gene sequence. The 16S rRNA gene sequence similarity to the closest phylogenetic relative Sandaracinobacter sibiricus was only 92.8 %. Accordingly, the three strains represent a new genus and species, for which the name Sandarakinorhabdus limnophila gen. nov., sp. nov., is proposed, with strain so42T (=DSM 17366T = CECT 7086T) as the designated type strain.