Paenibacillus Sp Research Articles

Cloning and expression of low temperature active endoglucanase EG5C from Paenibacillus sp. IHB B 3084

The endoglucanase gene designated as EG5C encoding cold active endoglucanase produced by Paenibacillus sp. IHB B 3084 was cloned and expressed in Escherichia coli BL21(DE3). The gene consisting of 1719bp open reading frame encoded a protein of 573 amino acids with a predicted molecular weight of 63.5kDa. The presence of N-terminal catalytic domain of the glycosyl hydrolase family 5 (GH5) and C-terminal carbohydrate binding X2 domain suggested the modular nature of the enzyme. The native signal peptide of EG5C was capable of efficiently secreting the enzyme with near equal activities in the cytoplasmic and extracellular fractions. The recombinant enzyme purified 9.46 fold to homogeneity with 22.33% yield gave 7.758IU/mg specific activity. The enzyme was stable over the broad pH range of 4–12 with more than 50% residual activity. The optimal activity was at 40°C with 70% relative activity at 5°C. The low temperature activity despite the shorter linker region suggested a novel cold adaptation mechanism by the enzyme. The enzyme displayed higher activity on carboxymethylcellulose than avicel which is useful in maintaining the tensile strength of fiber. The efficient secretion and low temperature activity offer prospect for large-scale production and industrial application of the endoglucanase.

Heterologous expression and characterization of detergent stable endoglucanase EG5B from Paenibacillus sp. IHB B 3084

The endoglucanase gene EG5B of 1611bp with a predicted molecular weight of 58.6kDa from Paenibacillus sp. IHB B 3084 was cloned and expressed in Escherichia coli BL21(DE3). The analysis of deduced amino acid sequence revealed a modular structure of the endoglucanase EG5B with an N-terminal catalytic domain of glycosyl hydrolase family 5 and a C-terminal carbohydrate-binding module of family 3. The purified enzyme showed high hydrolytic activity on carboxymethylcellulose, low activity on p-nitrophenyl β-d-cellobioside, avicel and filter paper, and no activity on microcrystalline cellulose, p-nitrophenyl β-d-glucoside, cellobiose and salicin as substrates. The enzyme was mild-alkaline active with optimum activity at pH 7–8 and stable over broad pH range. The temperature optimum was at 50°C with >50% activity over 30–60°C. The enzyme stability with >63% residual activity towards non-ionic and anionic surfactants and commercial detergents suggested its compatibility as an additive to detergents.

Expression of the N2 fixation gene operon of Paenibacillus sp. WLY78 under the control of the T7 promoter in Escherichia coli BL21

To investigate the transcription and translation and nitrogenase activity of the nine N2-fixing-gene (nif) operon (nifBHDKENXhesAnifX) of Paenibacillus sp. WLY78 under the control of the T7 promoter in Escherichia coli BL21 under different conditions. The Paenibacillus nif operon under the control of the T7 promoter is significantly transcribed and effectively translated in E. coli BL21 when grown in medium containing organic N compounds (yeast extract and Tryptone) or NH4+ by using RT-PCR and Western blot analysis. Transcription and translation of foreign nif genes in E. coli are not inhibited by environmental organic or inorganic N compounds or O2. However, contrary to transcription and translation, nitrogenase activity is 4% lower in the recombinant E. coli 78-32 compared to the native Paenibacillus sp. WLY78. The Paenibacillus nif operon under the control of T7 promoter enables E. coli BL21 to synthesize active nitrogenase. This study shows how the nif gene operon can be transferred to non-N2-fixing bacteria or to eukaryotic organelles.

Factors affecting the sorption of cesium in a nutrient-poor boreal bog

135Cs is among the most important radionuclides in the long-term safety assessments of spent nuclear fuel, due to its long half-life of 2.3 My and large inventory in spent nuclear fuel. Batch sorption experiments were conducted to evaluate the sorption behavior of radiocesium (134Cs) in the surface moss, peat, gyttja, and clay layers of 7-m-deep profiles taken from a nutrient-poor boreal bog. The batch distribution coefficient (Kd) values of radiocesium increased as a function of sampling depth. The highest Kd values, with a geometric mean of 3200 L/kg dry weight (DW), were observed in the bottom clay layer and the lowest in the 0.5–1.0 m peat layer (50 L/kg DW). The maximum sorption in all studied layers was observed at a pH between 7 and 9.5. The in situ Kd values of 133Cs in surface Sphagnum moss, peat and gyttja samples were one order of magnitude higher than the Kd values obtained using the batch method. The highest in situ Kd values (9040 L/kg DW) were recorded for the surface moss layer. The sterilization of fresh surface moss, peat, gyttja and clay samples decreased the sorption of radiocesium by 38%, although the difference was not statistically significant. However, bacteria belonging to the genera Pseudomonas, Paenibacillus, Rhodococcus and Burkholderia isolated from the bog were found to remove radiocesium from the solution under laboratory conditions. The highest biosorption was observed for Paenibacillus sp. V0-1-LW and Pseudomonas sp. PS-0-L isolates. When isolated bacteria were added to sterilized bog samples, the removal of radiocesium from the solution increased by an average of 50% compared to the removal recorded for pure sterilized peat. Our results demonstrate that the sorption of radiocesium in the bog environment is dependent on pH and the type of the bog layer and that common environmental bacteria prevailing in the bog can remove cesium from the solution phase.

Paenibacillin A, a new 2(1H)-pyrazinone ring-containing natural product from the endophytic bacterium Paenibacillus sp. Xy-2

A new 2(1H)-pyrazinone ring-containing natural product, paenibacillin A (1), together with five known diketopiperazine derivatives 2–6 and two known isoflavones 7–8, was isolated from the culture of an endophytic bacterium Paenibacillus sp. Xy-2. The structure of compound 1 was elucidated by extensive spectral methods, including UV, IR, HR-ESI-MS, 1D and 2D NMR and ECD experiments. Compound 1 exhibited moderate cytotoxicity against HL-60 cell line with IC50 value of 50.48 μM.

Influence of different factors on the nitrogenase activity of the engineered Escherichia coli 78-7.

The engineered Escherichia coli 78-7 is a derivative of E. coli JM109 carrying a nitrogen fixation (nif) gene cluster composed of nine genes (nifB, nifH, nifD, nifK, nifE, nifN, nifX, hesA and nifV) and its own σ(70)-dependent nif promoter from a gram-positive bacterium Paenibacillus sp. WLY78. The physiological and biochemical characteristics of the engineered E. coli 78-7 were analyzed by using Biolog GEN III MicroPlate, with E. coli JM109 and JM109/pHY300PLK (E. coli JM109 carrying empty vector) as controls. Analysis of 94 phenotypic tests: 71 carbon source utilization assays and 23 chemical sensitivity tests showed that the engineered E. coli 78-7, E. coli JM109 and JM109/pHY300PLK gave similar patterns of utilization of various substrates as carbon and energy sources. Furthermore, the effect of carbon source, nitrogen source, culture temperature on the nitrogenase activity of the engineered E. coli 78-7 was investigated. Our study demonstrates that the nif capacity of E. coli 78-7 was affected significantly by the different culture condition. The significant nitrogenase activity of E. coli 78-7 were obtained when cells were cultivated in the medium containing 4 g/l glucose (carbon source) and 2 mM glutamate (nitrogen source) and at 30 °C.

Evaluation of bacterial antagonists of Ralstonia solanacearum, causal agent of bacterial wilt of potato

Bacterial wilt of potato caused by Ralstonia solanacearum is one of the most destructive diseases in Kurdistan province, Iran. The objective of the present study was to evaluate antagonistic effects of some rhizobacteria isolated from the rhizosphere of potato plants against R. solanacearum, the agent of potato bacterial wilt. A total of 52 rhizobacteria were isolated and screened for in vitro antagonistic activity against R. solanacearum. Seven isolates with inhibiting effects of the pathogen were identified by phenotypic properties and partial sequencing of 16s rRNA as Pseudomonas fluorescens Pf11, P. fluorescens Pf16, Pseudomonas putida Pp17, Paenibacillus sp. Pb28 and Enterobacter sp. En38, Pseudomonas fluorescens Pp23 and Serratia sp. Se40. Strains Pf11, Pf16, Pp17 and Pb28 significantly inhibited the growth of the pathogen. Strains En38, Pp23 and Se40 showed a moderate or weak inhibition. During greenhouse study, strains were evaluated for their effects in reducing of disease and increasing biomass of potato plants. In according to greenhouse experiment results, isolates Pb28, Pp17 and Pf11significantly reduced disease by 55.56%, 51.50% and 38.58%, respectively. In addition, plant biomass significantly increased in plants treated with Pb28, Pp17, Pf11 and Pf16, compared to the control. Therefore, this study shows that these four strains have potential to be used as biocontrol agents against R. solanacearum. To confirm their effectiveness as commercial biocontrol agent, it is necessary to evaluate their efficiency in the field conditions in the next studies.

Recent advances in exopolysaccharides from Paenibacillus spp.: production, isolation, structure, and bioactivities.

This review provides a comprehensive summary of the most recent developments of various aspects (i.e., production, purification, structure, and bioactivity) of the exopolysaccharides (EPSs) from Paenibacillus spp. For the production, in particular, squid pen waste was first utilized successfully to produce a high yield of inexpensive EPSs from Paenibacillus sp. TKU023 and P. macerans TKU029. In addition, this technology for EPS production is prevailing because it is more environmentally friendly. The Paenibacillus spp. EPSs reported from various references constitute a structurally diverse class of biological macromolecules with different applications in the broad fields of pharmacy, cosmetics and bioremediation. The EPS produced by P. macerans TKU029 can increase in vivo skin hydration and may be a new source of natural moisturizers with potential value in cosmetics. However, the relationships between the structures and activities of these EPSs in many studies are not well established. The contents and data in this review will serve as useful references for further investigation, production, structure and application of Paenibacillus spp. EPSs in various fields.

Novel bioemulsifier produced by a Paenibacillus strain isolated from crude oil.

BackgroundSurface active compounds produced by microorganisms are attracting a pronounced interest due to their potential advantages over their synthetic counterparts, and to the fact that they could replace some of the synthetics in many environmental and industrial applications.ResultsBioemulsifier production by a Paenibacillus sp. strain isolated from crude oil was studied. The bioemulsifier was produced using sucrose with and without adding hydrocarbons (paraffin or crude oil) under aerobic and anaerobic conditions at 40°C. It formed stable emulsions with several hydrocarbons and its emulsifying ability was not affected by exposure to high salinities (up to 300 g/l), high temperatures (100°C-121°C) or a wide range of pH values (2–13). In addition, it presented low toxicity and high biodegradability when compared with chemical surfactants. A preliminary chemical characterization by Fourier Transform Infrared Spectroscopy (FT-IR), proton and carbon nuclear magnetic resonance (1H NMR and 13C CP-MAS NMR) and size exclusion chromatography indicated that the bioemulsifier is a low molecular weight oligosaccharide-lipid complex.ConclusionThe production of a low molecular weight bioemulsifier by a novel Paenibacillus strain isolated from crude oil was reported. To the best of our knowledge, bioemulsifier production by Paenibacillus strains has not been previously reported. The features of this novel bioemulsifier make it an interesting biotechnological product for many environmental and industrial applications.Graphical Novel bioemulsifier from Paenibacillus sp.

Study on the effect of magnetic field treatment of newly isolated Paenibacillus sp.

BackgroundSymbiotic nitrogen fixation in plants occurs in roots with the help of some bacteria which help in soil nitrogen fertility management. Isolation of significant environment friendly bacteria for nitrogen fixation is very important to enhance yield in plants.ResultsIn this study effect of different magnetic field intensity and treatment time was studied on the morphology, physiology and nitrogen fixing capacity of newly isolated Paenibaccilus sp. from brown soil. The bacterium was identified by 16S rDNA sequence having highest similarity (99%) with Paenibacillus sp as revealed by BLAST. Different magnetic intensities such as 100mT, 300mT and 500mT were applied with processing time of 0, 5, 10, 20 and 30 minutes. Of all these treatment 300mT with processing time of 10 minutes was found to be most suitable treatment. Results revealed that magnetic treatment improve the growth rate with shorter generation time leading to increased enzyme activities (catalase, peroxidase and superoxide dismutase) and nitrogen fixing efficiencies. High magnetic field intensity (500mT) caused ruptured cell morphology and decreased enzyme activities which lead to less nitrogen fixation.ConclusionIt is concluded that appropriate magnetic field intensity and treatment time play a vital role in the growth of soil bacteria which increases the nitrogen fixing ability which affects the yield of plant. These results were very helpful in future breading programs to enhance the yield of soybean.Electronic supplementary materialThe online version of this article (doi:10.1186/s40529-015-0083-9) contains supplementary material, which is available to authorized users.

Characterization of Xylanase activity produced by Paenibacillus sp. XJ18 from TNBD Jambi, Indonesia

Lignocellulose waste in nature is increasing due to the increasing activity of agroforestry. Up to 40% of lignocellulose biomass are consisted of xylan. Xylan complete breakdown requires the action of xylanase. Xylanase has been used to breakdown xylan into commercial product such as low calories sugar, prebiotic, and biofuel. Due to its wide application, several variation of xylanase characterization are needed. Our previous studies have collected Paenibacillus sp. XJ18 from TNBD forest, Jambi, Indonesia, to gain a unique enzyme characteristic. In this study the characteristic of crude xylanase from Paenibacillus sp. XJ18 was investigated. The highest activity of xylanase production was at 36 h. The xylanase showed activity in a broad range of pH (4.5-9.0). The highest activity showed at pH 5.0, 90 oC. Crude enzyme extract was unstable and had halftime at its pH and optimum temperature about 67 min. The xylanase activity was increased about 4.59 times after being concentrated by 70% acetone (2.4578 U/mL). Based on TLC result, xylanase from Paenibacillus sp. XJ18 was predicted to produce xylobiose exclusively from extracted corncob xylan.

Improvement and Immobilization of a new Endo-β-1,4-xylanases KRICT PX1 from Paenibacillus sp. HPL-001

A new endo-1,4-beta-xylanase gene (FJ380951), KRICT PX1, isolated from Paenibacillus sp. HPL-001, was expressed in E. coli. Enzyme purification, mutation, immobilization, and molecular simulation were conducted. The P1H1 mutant of one amino acid substitution (168Leu￫Gln) showing the strongest xylanase activity was selected among 250 mutant clones. The specific activity of the P1H1 xylanase was 53.3 U/mg protein at 50°C, an approximately fivefold increase compared to that of the original KRICT PX1 xylanase (10.25 U/mg protein) at the same condition of pH 5~10, and enhanced activity from 20.1 to 36.6 U/mg proteins at 60°C. The structural dynamics of the mutant P1H1 became stronger than that of the wild type, which could be eliminated by a helix deformation with the H-bond missing between Leu168 and Asp164 after mutation. Xylanase could be reused for 5 batches without significant loss of activity after being immobilized on surface functionalized silica-based mesoporous cellular foam. However, xylanase activity declined to 60% at the 10th batch. Further research on practical applications will be necessary for industrial usage.

Recent trends in control methods for bacterial wilt diseases caused by Ralstonia solanacearum.

Previous studies have described the development of control methods against bacterial wilt diseases caused by Ralstonia solanacearum. This review focused on recent advances in control measures, such as biological, physical, chemical, cultural, and integral measures, as well as biocontrol efficacy and suppression mechanisms. Biological control agents (BCAs) have been dominated by bacteria (90%) and fungi (10%). Avirulent strains of R. solanacearum, Pseudomonas spp., Bacillus spp., and Streptomyces spp. are well-known BCAs. New or uncommon BCAs have also been identified such as Acinetobacter sp., Burkholderia sp., and Paenibacillus sp. Inoculation methods for BCAs affect biocontrol efficacy, such as pouring or drenching soil, dipping of roots, and seed coatings. The amendment of different organic matter, such as plant residue, animal waste, and simple organic compounds, have frequently been reported to suppress bacterial wilt diseases. The combined application of BCAs and their substrates was shown to more effectively suppress bacterial wilt in the tomato. Suppression mechanisms are typically attributed to the antibacterial metabolites produced by BCAs or those present in natural products; however, the number of studies related to host resistance to the pathogen is increasing. Enhanced/modified soil microbial communities are also indirectly involved in disease suppression. New promising types of control measures include biological soil disinfection using substrates that release volatile compounds. This review described recent advances in different control measures. We focused on the importance of integrated pest management (IPM) for bacterial wilt diseases.

Identification and Characterization of 1,4-β-D-Glucan Glucohydrolase for the Saccharification of Cell Oligomers from Paenibacillus sp. HPL-001

The 1,4-β-D-glucan glucohydrolase gene (Ggh) which was isolated from Paenibacillus sp. strain HPL-001 (KCTC11365BP) has been cloned and expressed in Escherichia coli, and efficiently purified using affinity column chromatography. Recombinant 1,4-β-Dglucan glucohydrolase (719aa, NCBI accession number KJ573391) was highly active at 40°C in pH 6.0 without significant changes to most salts tested, and exhibited Km 0.893 mg/mL Vmax 33.33 U/mg on pNPG, respectively. All soluble cellooligomers (e.g., cellobiose, cellotriose, cellotetraose, cellopentaose, and cellohexaose) had been virtually hydrolyzed to glucose by this enzyme. When compared with the commercialized-enzymes, the hydrolytic pattern to all substrates was almost same to the Celluclast 1.5L (Novozyme) with about 1/3 strength, however hydrolytic pattern of Glucosidase (Lucigen) and Almonds (Sigma) was significantly decreased with substrates increasing number of glucose polymer from cellotriose to cellohexaose. About 90% of the initial enzyme activity was maintained even after 10 consecutive recycle by the 11-carbon bridge and aldehyde-functionalized MCF-immobilization. 3-D structure of this enzyme has the ligand of cellobiose and cellulose binding in the center of niche according to the sequence information.

Physiological and genetic screening methods for the isolation of methyl tert-butyl ether-degrading bacteria for bioremediation purposes

Bioremediation of groundwater contaminated with methyl tert-butyl ether (MTBE) has been widely described since their cost/efficient ratios are lower than other physic-chemical methodologies. The present study focused on the isolation and selection of MTBE degrading microorganisms from contaminated soil and groundwater samples based on results from growth on mineral media amended with MTBE and BTEX, presence or absence of the monooxygenase genes and specific ability to degrade MTBE. Three bacterial strains were selected and identified as Rhodococcus ruber, strains EE1 (CECT 8555), EE6 (CECT 8612) and A5 (CECT 8556), showing the ability to degrade 60.0, 36.0 and 10.0 mg l−1 MTBE, respectively. Moreover, all the R. ruber strains showed the presence of genes encoding MTBE-degrading enzymes. One isolated strain was identified as Paenibacillus sp. SH7 (CECT 8558) and demonstrated the greatest MTBE degradation value (100 mg l−1), but together with the last strain selected and identified as Agrobacterium sp. MS2 (CECT 8557) did not result in positive amplification of any of the monooxygenase primers tested. The lowest toxicity (as EC50) was observed after 4-days growth of R. ruber EE6 on MTBE-supplemented mineral medium. The potential application of these strains in bioremediation processes is discussed.

Characterization of a GH family 8 β-1,3-1,4-glucanase with distinctive broad substrate specificity from Paenibacillus sp. X4.

A β-1,3-1,4 glucanase gene of Paenibacillus sp. X4, bglc8H, was cloned and characterized. BGlc8H was predicted to be a protein of 409 amino acid residues, including a signal peptide of 31 amino acids. The mature enzyme was predicted to have 378 amino acid residues; its [corrected] molecular mass and pI were estimated as 41,561 Da and 7.61, respectively. BGlc8H belongs to glycoside hydrolase family 8 (GH8). Site-directed mutants of Glu95 and Asp156 of BGlc8H showed a near-complete loss of activity, indicating that they are catalytically-active residues. Unlike other GH8 members, BGlc8H had broad substrate specificity and hydrolyzed barley-β-D-glucan > chitosan > carboxymethyl-cellulose > and lichenan. BGlc8H had a lower ratio of lichenase/barley-β-D-glucanase activities compared to GH16 enzymes. BGlc8H was optimally active at pH 5 and 50 °C, except for barley-β-D-glucanase (40 °C) and chitosanase (pH 7) activities. BGlc8H hydrolyzed cello-oligosaccharides (G3-G6) to G3 and G2 but not to G1. Ca(2+) increased the activity and thermostability of BGlc8H for lichenan suggesting its use for the saccharification of cellulosic biomass.

Microcosmic study of endosulfan degradation by Paenibacillus sp. ISTP10 and its toxicological evaluation using mammalian cell line

In the present study, an endosulfan degrading strain Paenibacillus sp. ISTP10 was isolated from activated sludge. Soil microcosms were set up with endosulfan (60 mg kg−1 of dry soil) to evaluate the degradation potency of the strain. Soil samples from the microcosms were collected at regular intervals and the organic compounds were extracted with hexane. GC–MS analysis of the soil extract showed the formation of metabolites of endosulfan such as endosulfan diol and endosulfan ether confirming that the strain degrades endosulfan via a hydrolytic pathway. Methyl tetrazolium (MTT) assay for cytotoxicity and alkaline comet assay for genotoxicity were carried out in human hepato-carcinoma cell line HepG2 to evaluate the toxic potential of endosulfan and its degraded metabolites. The bacterium reduced toxicity as determined by an increase in LC50 value by 75.86 fold and a reduction in Olive Tail Moment by 21 fold after 30 days of treatment. The by-products of degradation were found to be less toxic than the parent compound showing the biodegradation and detoxification potential of endosulfan by Paenibacillus sp. ISTP10.

Identification of mVOCs from Andean rhizobacteria and field evaluation of bacterial and mycorrhizal inoculants on growth of potato in its center of origin.

Food security (a pressing issue for all nations) faces a threat due to population growth, land availability for growing crops, a changing climate (leading to increases in both abiotic and biotic stresses), heightened consumer awareness of the risks related to the use of agrichemicals, and also the reliance on depleting fossil fuel reserves for their production. Legislative changes in Europe mean that fewer agrichemicals will be available in the future for the control of crop pests and pathogens. The need for the implementation of a more sustainable agricultural system globally, incorporating an integrated approach to disease management, has never been more urgent. To that end, the Valorizing Andean Microbial Diversity (VALORAM) project (http://valoram.ucc.ie), funded under FP7, examined the role of microbial communities in crop production and protection to improve the sustainability, food security, environmental protection, and productivity for rural Andean farmers. During this work, microbial volatile organic compounds (mVOCs) of 27 rhizobacterial isolates were identified using gas chromatography/mass spectrometry (GC/MS), and their antifungal activity against Rhizoctonia solani was determined in vitro and compared to the activity of a selection of pure volatile compounds. Five of these isolates, Pseudomonas palleroniana R43631, Bacillus sp. R47065, R47131, Paenibacillus sp. B3a R49541, and Bacillus simplex M3-4 R49538 trialled in the field in their respective countries of origin, i.e., Bolivia, Peru, and Ecuador, showed significant increase in the yield of potato. The strategy followed in the VALORAM project may offer a template for the future isolation and determination of putative biocontrol and plant growth-promoting agents, useful as part of a low-input integrated pest management system.

Purification and characterization of β-agarase from Paenibacillus sp.

β-Agarase produced by Paenibacillus sp. WL (agarase WL) was purified using a combination of ammonium sulfate precipitation, DEAE-ion exchange, and gel-filtration chromatography. The purity of the agarase was increased by 11.9× with a recovery of 5.1% and a specific activity of 4,670.1 U/mg of protein. The molecular mass of the purified agarase was approximately 30 kDa (SDS-PAGE). The agarase was stable at temperature below 50°C and the favorable agar-hydrolysis activity was at 40°C. The agarase was active in the range of pH 5.0 to 8.0, and the optimal agar-hydrolysis pH value was approximately 6.0. Metal ions normally found in seawater (Na+, K+, Ca2+, Mg2+, and Al3+) could activate agarase WL. The Michaelis-Menten constant Km and maximal reaction velocity Vmax of purified agarase WL were 3.22 mg/mL and 41.5 μg/mL·min, respectively. The agarase WL was highly agar specific.

Biotransformation of the Principal Ginsenosides of Panax ginseng Into Minor Glycosides Through the Action of Bacterium Paenibacillus sp. BG134

The bacterium Paenibacillus sp. BG134 was capable of biotransforming the principal 20(S)-protopanaxadiol ginsenosides Rc, Rb2, Rd, and Rb1 into the corresponding minor glycosides C-Mc1, C-O, and F-2. The specificity of Paenibacillus sp. BG134 differed from that of several other microorganisms by cleaving only the terminal C-3 and C-20 β -D-glucose from their carbohydrate components.