Optimization of conditions for the production of lignocellulolytic enzymes by Sphingobacterium sp. ksn-11 utilizing agro-wastes under submerged condition.

The present work was aimed at studying the production of lignocellulolytic enzymes, namely cellulase, xylanase, pectinase, mannanase, and laccase by a newly isolated bacterium Sphingobacterium sp. ksn-11, utilizing various agro-residues as a substrate under submerged conditions. The production of lignocellulolytic enzymes was found to be maximum at the loading of 10%(w/v) agro-residues. The enzyme secretion was enhanced by two-fold at 2 mM CaCO3, optimum pH 7, and temperature 40°. The Field Emission Gun-Scanning Electron Microscope (FEG-SEM) results have shown the degradative effect of lignocellulases; cellulase, xylanase, mannanase, pectinase, and laccase on corn husk with 3.55 U/ml, 79.22 U/ml, 12.43 U/ml, 64.66 U/ml, and 21.12 U/ml of activity, respectively. The hydrolyzed corn husk found to be good adsorbent for polyphenols released during hydrolysis of corn husk providing suitable conditions for stability of lignocellulases. Sphingobacterium sp. ksn is proved to be a promising candidate for lignocellulolytic enzymes in view of demand for enzymes in the biofuel industry.

Mitigation of soil contaminated with diesel fuel using bioelectrokinetics.

This study investigated the effectiveness of bioelectrokinetics in rehabilitating a silty clayey sand contaminated with diesel fuel using three novel bacterial strains; Acinetobacter calcoaceticus, Sphingobacterium multivorum, and Sinorhizobium, isolated form agriculture land. Three electrokinetic bioremediation cells were used to conduct the tests and a novel electrode configuration technique was used to stabilize pH and water content in the soil specimen. Solar photovoltaic panels were used to generate sustainable energy for the process. The tests were carried out in outdoors for 55 days. Applied voltage, current passing through the electrokinetic cell, and the temperature of the soil specimen were recorded periodically during the test. The pH, water content, and diesel concentration were determined at the end of the tests. Over the test period, the voltage typically increased from zero before sunrise, remained relatively stabilized for about 4 h, and then started to decrease and dropped to zero by sunset. The temperatures in the cells were found to be 5-7 °C higher than the ambient temperature. The innovative electrode configuration succeeded in keeping the pH of soil to remain the same and thereby prevented the development of a pH gradient in the soil, an important development for survival of the bacteria. The diesel degradation in the soil after bioelectrokinetics were 20-30%, compared to 10-12% in the control test. The study was successful in developing environmentally friendly technology employing novel bacterial strains to degrade diesel fuel and utilizing solar panel to produce renewable energy for bioelectrokinetic during the winter season.

Seasonal assemblages and short-lived blooms in coastal north-west Atlantic Ocean bacterioplankton.

Temperate oceans are inhabited by diverse and temporally dynamic bacterioplankton communities. However, the role of the environment, resources and phytoplankton dynamics in shaping marine bacterioplankton communities at different time scales remains poorly constrained. Here, we combined time series observations (time scales of weeks to years) with molecular analysis of formalin-fixed samples from a coastal inlet of the north-west Atlantic Ocean to show that a combination of temperature, nitrate, small phytoplankton and Synechococcus abundances are best predictors for annual bacterioplankton community variability, explaining 38% of the variation. Using Bayesian mixed modelling, we identified assemblages of co-occurring bacteria associated with different seasonal periods, including the spring bloom (e.g. Polaribacter, Ulvibacter, Alteromonadales and ARCTIC96B-16) and the autumn bloom (e.g. OM42, OM25, OM38 and Arctic96A-1 clades of Alphaproteobacteria, and SAR86, OM60 and SAR92 clades of Gammaproteobacteria). Community variability over spring bloom development was best explained by silicate (32%)--an indication of rapid succession of bacterial taxa in response to diatom biomass--while nanophytoplankton as well as picophytoplankton abundance explained community variability (16-27%) over the transition into and out of the autumn bloom. Moreover, the seasonal structure was punctuated with short-lived blooms of rare bacteria including the KSA-1 clade of Sphingobacteria related to aromatic hydrocarbon-degrading bacteria.

Screening of a glucoside 3-dehydrogenase-producing strain, Sphingobacterium faecium, based on a high-throughput screening method and optimization of the culture conditions for enzyme production.

The objective of this study was to screen glucoside 3-dehydrogenase (G3DH)-producing strain based on a high-throughput G3DH screening method. Optimization of culture conditions of the isolated strain was also applied in this study. This screening method employed electron transfer reaction in 96-well microtiter plates, α-methyl-D-glucoside, galactose, 2-deoxy-D-glucose, and 3-O-methyl-D-glucose were used as substrates. Using this screening method, one out of 78 strains isolated from different soil samples was obtained with high G3DH activity. The accuracy of the screening method was proved by alkaline treatment analysis of 3-keto sugars. The isolated strain was identified as Sphingobacterium faecium ZJF-D6 by phenotypic characterization and 16S rDNA sequence analysis. The culture conditions of S. faecium for G3DH production were optimized. Sucrose was found as the most suitable carbon source for the G3DH production. The highest G3DH production and cell growth were achieved using the medium at the initial pH of 7.0 at 25 °C for 36 h with activity of 8.03 × 10(-2) U/mL culture. This strain appears promising for potential application in the industry to produce 3-keto sugars. To our knowledge, this is the first report on S. faecium for G3DH production. The method described herein represents a useful tool for the high-throughput isolation of G3DH.

Transient response of microbial communities in a water well field to application of an impressed current.

Deterioration of water wells due to clogging and corrosion over time is a common problem where solutions may be costly and ineffective. Pilot studies have suggested that impressed current or cathodic protection may be used to reduce microbially-induced declines in water well performance. Two water wells in an alluvial aquifer close to the North Saskatchewan River were selected to study the response of subsurface microbial communities to the application of an impressed current as an anti-fouling technology. The treated well was exposed to an impressed current while the untreated well was used as a reference site. Biofilms grown on in situ coupons under the influence of the impressed current were significantly (p < 0.05) thicker (mean thickness = 67.3 µm) when compared to the biofilms (mean thickness = 19.3 µm) grown outside the electric field. Quantitative PCR analyses showed significantly (p < 0.05) higher numbers of total bacteria, iron- and nitrate-reducers in the electrified zone. Molecular analysis revealed that the predominant bacteria present in biofilms grown under the influence of the impressed current belonged to Rhodobacter spp., Sediminibacterium spp. and Geobacter spp. In addition to favouring the growth of biofilms, direct microscopic and ICP-AES analyses revealed that the impressed current also caused the deposition of iron and manganese on, and in the vicinity of, the well screen. Together, these factors contributed to rapid clogging leading to reduced specific pumping capacities of the treated well. The study revealed that the impressed current system was not effective as an anti-fouling technology but actually promoted both microbial growth and physical clogging in this aquifer.

Volatile methyl esters of medium chain length from the bacterium Chitinophaga Fx7914.

The analysis of the volatiles released by the novel bacterial isolate Chitinophaga Fx7914 revealed the presence of ca. 200 compounds including different methyl esters. These esters comprise monomethyl- and dimethyl-branched, saturated, and unsaturated fatty acid methyl esters that have not been described as bacterial volatiles before. More than 30 esters of medium C-chain length were identified, which belong to five main classes, methyl (S)-2-methylalkanoates (class A), methyl (S)-2,(ω-1)-dimethylalkanoates (class B), methyl 2,(ω-2)-dimethylalkanoates (class C), methyl (E)-2-methylalk-2-enoates (class D), and methyl (E)-2,(ω-1)-dimethylalk-2-enoates (class E). The structures of the compounds were verified by GC/MS analysis and synthesis of the target compounds as methyl (S)-2-methyloctanoate (28), methyl (S)-2,7-dimethyloctanoate ((S)-43), methyl 2,6-dimethyloctanoate (49), methyl (E)-2-methylnon-2-enoate (20a), and methyl (E)-2,7-dimethyloct-2-enoate (41a). Furthermore, the natural saturated 2-methyl-branched methyl esters showed (S)-configuration as confirmed by GC/MS experiments using chiral phases. Additionally, the biosynthetic pathway leading to the methyl esters was investigated by feeding experiments with labeled precursors. The Me group at C(2) is introduced by propanoate incorporation, while the methyl ester is formed from the respective carboxylic acid by a methyltransferase using S-adenosylmethionine (SAM).

Degradation characteristics and metabolic pathway of 17alpha-ethynylestradiol by Sphingobacterium sp. JCR5.

A 17alpha-ethynylestradiol (EE2)-degrading bacterium was isolated from the activated sludge of the wastewater treatment plant (WWTP) of an oral contraceptives producing factory in Beijing, China. On the basis of its morphology, biochemical properties and the 16S rDNA sequence analysis, this strain was identified as Sphingobacterium sp. JCR5. This strain grew on EE2 as sole source of carbon and energy, and metabolized up to 87% of the substrate added (30 mgl-1) within 10 d at 30 degrees C. In addition to EE2 the strain could be cultivated on steroidal estrogens like estrone (E1), 17beta-estradiol (E2), estriol (E3) and mestranol (MeEE2), the intermediates of contraceptive medicine processing and on some aromatic compounds. Mass spectrum analysis of the EE2 degradation showed that in the first step it is oxygenized to E1, 2-hydroxy-2,4-dienevaleric acid and 2-hydroxy-2,4-diene-1,6-dioic acid, which are the main catabolic intermediates. The former was analogous to the pathway of a previously reported testosterone-degrading bacterium Comamonas testosteroni TA441 and the latter is a metabolite with a different cleavage position of 3-hydroxy-4,5-9,10-disecoestrane-1(10),2-diene-5,9,17-trione-4-oic acid from the former.

[Degradation pathway of 17alpha-ethynylestradiol by Sphingobacterium sp. JCR5].

A bacterial strain, JCR5, which degrades 17alpha-ethynylestradiol (EE2), was isolated from activated sludge of wastewater treatment plant treating wastewater from pharmacy factory mainly producing contraceptive medicine in Beijing, China. Based on its morphology, physiology biochemical properties and 16S rDNA sequence analysis, this strain was identified as Sphingobacteriumn sp. JCR5. Strain JCR5 can grow on EE2 as sole carbon and energy source. The degradation process for EE2 with initial concentration of 30 mg x L(-1) was studied, and the result indicated that the degradation rate of EE2 within 10 days was 87%. Experiments of different substrates showed that strain JCR5 can grow on many substrates other than EE2 such as several steroidal estrogens (estrone, 17beta-estradiol, estriol and mestranol), the intermediates in contraceptive medicine processing and some aromatic compounds. Mass spectrum analysis demonstrated that EE2 was oxygenized to estrone (E1) firstly, and 2-hydroxy-2,4-dienevaleric acid and 2-hydroxy-2,4-diene-1, 6-dioic acid were the main catabolic intermediates during EE2 degradation. The former adopted a pathway that was analogical to the pathway of the previously reported testosterone-degrading Comamnonas testosteroni TA441, and the latter was a metabolite with a different cleavage position of 3-hydroxy-4,5-9, 10-disecoestrane-1(10), 2-diene-5,9, 17- trione-4-oic acid from the former.

Effects of nutrient availability and Ochromonas sp. predation on size and composition of a simplified aquatic bacterial community.

Predation and competition are two main factors that determine the size and composition of aquatic bacterial populations. Using a simplified bacterial community, composed of three strains characterized by different responses to predation, a short-term laboratory experiment was performed to evaluate adaptations and relative success in communities with experimentally controlled levels of predation and nutrient availability. A strain with a short generation time (Pseudomonas putida), one with high plasticity in cell morphology (Flectobacillus sp. GC5), and one that develops microcolonies (Pseudomonas sp. CM10), were selected. The voracious flagellate Ochromonas sp. was chosen as a predator. To describe adaptations against grazing and starvation, abundance, biomass and relative heterogeneity of bacteria were measured. On the whole, the strains in the predation-free cultures exhibited unicellular growth, and P. putida represented the largest group. The presence of Ochromonas strongly reduced bacterial abundance, but not always the total biomass. The activity of grazers changed the morphological composition of the bacterial communities. Under grazing pressure the relative composition of the community depended on the substrate availability. In the presence of predators, P. putida abundance declined in both high and low nutrient treatments, and Pseudomonas CM10 developed colonies. Flectobacillus was only numerically codominant in the nutrient-rich environments.

Variations of bacterial community structure in flooded paddy soil contaminated with herbicide quinclorac.

The denaturing gradient gel electrophoresis (DGGE) method was applied to determine the relative genetic complexity of microbial communities in flooded paddy soil treated with herbicide quinclorac (3,7-dichloro-8-quinoline-carboylic acid). The results obtained showed a significant effect of quinclorac on the development of bacterial populations in soils contaminated with different concentrations of the herbicide at the early time after application. In general, however, the number of populations of the same soil sample treated with the same concentration of the quinclorac differed obviously with increasing incubation time within the early 8 weeks. The scale of differences in banding patterns-showed that the microbial community structures of the quinclorac-treated and non-quinclorac-treated soils were not significantly different after 21 weeks of incubation. Quantification, as demonstrated in this paper, was studied by establishing dose-response relationships. Significant pattern variations were quantified. Prominent DGGE bands were excised, cloned and sequenced to gain insight into the identities of predominant bacterial populations. The majority of DGGE band sequences were related to bacterial genera Clostridium, Sphingobacterium, Xanthomonas and Rhodococcus.

Bioconversion of isopropanol by a solvent tolerant Sphingobacterium mizutae strain.

The bioconversion of high concentration isopropanol (2-propanol, IPA) was investigated by a solvent tolerant strain of bacteria, which was identified as Sphingobacterium mizutae ST2 by partial 16S rDNA gene sequencing. This strain of bacteria exhibited the ability to utilise high concentration isopropanol as the sole carbon source, with mineralization occurring via an acetone intermediate into central metabolism. The biodegradative performance of this strain for IPA was examined over a 2-38 g l(-1) concentration range, using specific growth rate (mu) and conversion rate analysis. Maximum specific growth rates (mu(max)) of 0.0045 h(-1 )were routinely obtainable on IPA. In addition, the highest specific IPA degradation rate was obtained at a concentration of 7.5 g l(-1) with a corresponding value of 0.045 g IPA g cells(-1) h(-1). While the highest acetone yield reached its maximum value of 0.940 g acetone g IPA(-1) at 7.5 g IPA l(-1). This is the first report on bioconversion of isopropanol at such high concentration by this solvent tolerant strain of S. mizutae and may allow its application in novel biocatalytic processes for effective biological conversion in two-phase solvent systems.

Pedobacter sandarakinus sp. nov., isolated from soil.

A Gram-negative, non-motile, rod-shaped bacterial strain, designated DS-27(T), was isolated from a soil sample, and its taxonomic position was investigated by using a polyphasic approach. The organism grew optimally at 30 degrees C and in the presence of 0-0.5 % (w/v) NaCl. Strain DS-27(T) contained MK-7 as the predominant menaquinone and iso-C(15 : 0), C(16 : 1)omega7c and/or iso-C(15 : 0) 2-OH and iso-C(17 : 0) 3-OH as the major fatty acids. The DNA G + C content was 39.7 mol%. Phylogenetic analyses based on 16S rRNA gene sequences revealed that strain DS-27(T) is most closely related to the genus Pedobacter of the family Sphingobacteriaceae. Similarity values between the 16S rRNA gene sequences of strain DS-27(T) and the type strains of recognized Pedobacter species ranged from 90.6 to 95.5 %. Differential phenotypic properties, together with the phylogenetic distinctiveness, were sufficient to categorize strain DS-27(T) as representing a species that is separate from recognized Pedobacter species. On the basis of its phenotypic properties and phylogenetic distinctiveness, strain DS-27(T) (=KCTC 12559(T) = CIP 108922(T)) was classified in the genus Pedobacter as a member of a novel species, for which the name Pedobacter sandarakinus sp. nov. is proposed.

The effect of simulated microgravity on bacteria from the Mir space station.

The effects of simulated microgravity on two bacterial isolates, Sphingobacterium thalpophilium and Ralstonia pickettii (formerly Burkholderia pickettii), originally recovered from water systems aboard the Mir space station were examined. These bacteria were inoculated into water, high and low concentrations of nutrient broth and subjected to simulated microgravity conditions. S. thalpophilium (which was motile and had flagella) showed no significant differences between simulated microgravity and the normal gravity control regardless of the method of enumeration and medium. In contrast, for R. pickettii (that was non-motile and lacked flagella), there were significantly higher numbers in high nutrient broth under simulated microgravity compared to normal gravity. Conversely, when R. pikkettii was inoculated into water (i.e., starvation conditions) significantly lower numbers were found under simulated microgravity compared to normal gravity. Responses to microgravity depended on the strain used (e.g., the motile strain exhibited no response to microgravity, while the non-motile strain did), the method of enumeration, and the nutrient concentration of the medium. Under oligotrophic conditions, non-motile cells may remain in geostationary orbit and deplete nutrients in their vicinity, while in high nutrient medium, resources surrounding the cell may be sufficient so that high growth is observed until nutrients becoming limiting.

Bacteremia caused by a novel species of Sphingobacterium.

A Gram-negative rod was isolated from the blood cultures of an 84-year-old man with foot cellulitis. The bacterium was first identified as Sphingobacterium spiritivorum on the basis of standard assimilation tests. However, sequencing analysis of its 16S rRNA genes and whole genome hybridization studies with other related bacteria showed that this isolate belongs to a so far undescribed species of Sphingobacterium, close to S. mizutae. This bacterium was susceptible to most of the antibiotics tested, including glycopeptides, but was resistant to aminoglycosides and polymyxins. Treatment with amoxicillin-clavulanate cured the infection.