Multifunctional Exopolysaccharides from Pseudomonas aeruginosa PF23 Involved in Plant Growth Stimulation,Biocontrol and Stress Amelioration in Sunflower Under Saline Conditions

Isolate PF23 selected from among 110 fluorescent pseudomonads, identified as Pseudomonas aeruginosa, displayed salinity tolerance and exopolysaccharides (EPS) production up to 2,000 mM NaCl concentration. EPS-defective mutant PF23^EPS? of the isolate showed 86 % reduction in EPS production in comparison with wild strain. Defect in EPS production brought loss in salt tolerance capability. Purified EPS obtained from PF23 displayed multiple roles. At low concentration EPS functioned as biocontrol agent, at high concentration EPS behaved as osmoprotective or stress ameliorating metabolite and when introduced in saline soil, served as a plant growth promotor along with seed biopriming agent. Both in planta and in vivo studies were performed taking sunflower as a test crop and it was observed that PF23 showed plant growth promotion and significant biocontrol potential against dreadful phytopathogen Macrophomina phaseolina (under saline conditions). The mutant PF23^EPS? was ineffective under saline conditions both in growth enhancement as well as in disease suppression. The study reports a potent strain, Pseudomonas aeruginosa PF23, capable of enhancing production of sunflower crop in semiarid regions and minimizing the incidence of charcoal rot disease in sunflower.     

Fungal treatment followed by FeCl3 treatment to enhance enzymatic hydrolysis of poplar wood for high sugar yields

Fungal treatment followed by FeCl₃ treatment was used to improve saccharification of wood from Populus tomentosa. Combined treatments accumulated lignin and slightly degraded cellulose, whereas almost all hemicelluloses were removed. The white rot fungus, Trametes orientalis, and the brown rot fungus, Fomitopsis palustris, both accompanied by FeCl₃ post-treatment resulted in 98.8 and 99.7 % of hemicelluloses loss at 180 °C, respectively, which were over twice than that of hot water pretreatment at the same level. In addition, the solid residue from the T. orientalis-assisted and F. palustris-assisted FeCl₃ treatment at 180 °C released 84.5 and 95.4 % of reducing sugars, respectively: 1.4- and 1.6-fold higher than that of FeCl₃ treatment alone at the same temperature. Combined treatments disrupted the intact cell structure and increased accessible surface area of cellulose therefore enhancing the enzymatic digestibility, as evidenced by XRD and SEM analysis data.     

A Review of: “Progress in Separation and Purification Vol. 3, E. S. Perry and C. J,van Oss,eds., Willy/Intersciencc,New York, 1970; xi + 316 pares; hard cover; 515.95”

The optimal culture conditions of exopolysaccharides (EPS) production in submerged culture medium by Pleurotus geesteranus 5 ^# were determined using an orthogonal matrix method. The optimal defined medium (per liter) was 60.0 g maltose, 5.0 g tryptone, 1 mM NaCl, 5 mM KH₂PO₄, and initial pH 6.0 at 28 °C. In the optimal culture medium, the maximum EPS production was 16.97 g/L in a shake flask. Two groups of EPSs (designated as Fr-I and Fr-II) were obtained from the culture filtrates by size exclusion chromatography (SEC), and their molecular characteristics were examined by a multiangle laser-light scattering (MALLS) and refractive index (RI) detector system. The approximate weight-average molar masses of the Fr-I and Fr-II of EPS were determined to be 3.263 × 10⁴ and 5.738 × 10³ g/mol, respectively. The low values of polydispersity ratio (1.176 and 1.124 for Fr-I and Fr-II, respectively) of EPSs mean that these EPS molecules exist much less dispersed in aqueous solution without forming large aggregates. Furthermore, the experiments in vitro indicated that P. geesteranus 5^# EPS exhibit high antitumor and antioxidative effects.     

Cadmium induced oxalic acid secretion and its role in metal uptake and detoxification mechanisms in <Emphasis Type="Italic">Phanerochaete chrysosporium</Emphasis>

This study examines the role of oxalic acid in the uptake of Cd and participation in detoxification process in Phanerochaete chrysosporium. Cd-induced oxalic acid secretion was observed with growth inhibition and enzyme inactivation (LiP and MnP) of P. chrysosporium. The peak value of oxalic acid concentration was 16.6 mM at initial Cd concentration of 100 mg L^?1. During the short-term uptake experiments, the uptake of Cd was enhanced and accelerated in the presence of oxalic acid and resulted in alleviated growth and enzyme inhibition ratios. The formation of a metal-oxalate complex therefore may provide a detoxification mechanism via effect on metal bioavailability, whereby many fungi can survive and grow in environments containing high concentrations of toxic metals. The present findings will advance the understanding of fungal resistance to metal stress, which could show promise for a more useful application of microbial technology in the treatment of metal-polluted waste.     

Enhancement of exopolysaccharide production from Ganoderma lucidum using a novel submerged volatile co-culture system

Based on the impact of volatile organic compounds (VOCs) on secondary metabolite pathways, a novel submerged volatile co-culture system was constructed, and the effects of thirteen fungal and bacterial VOCs were investigated on Ganoderma lucidum exopolysaccharides production. The results demonstrated at least a 2.2-fold increase in exopolysaccharide (EPS) specific production yield in 6 days submerged volatile co-culture of G. lucidum with Pleurotus ostreatus. Therefore, P. ostreatus was selected as a variable culture, and the effects of agitation speed, inoculum size, initial pH, and co-culture volume on EPSs production were investigated using a Taguchi L₉ orthogonal array. Finally, the highest concentration of EPSs (3.35 ± 0.22 g L^?1) was obtained under optimized conditions; initial pH 5.0, inoculum size 10%, 150 rpm, and 3:1 volume ratio of variable culture to main culture.     

Effect of different media on exopolysaccharide and biomass production by the green microalga Botryococcus braunii

Botryococcus braunii is a colonial green microalga with recognized potential to synthesize lipids and hydrocarbons for biofuel production. Besides this ability, this microalga also produces exopolysaccharides (EPS). Nevertheless, there are few reports about their biotechnological aspects and industrial applications. In this study, the effect of the nutritional conditions was examined by using two different culture media (BG11 and D medium). To our knowledge, the latter has not been reported before for culturing B. braunii. After 49 days of incubation, the final production of EPS was found to be statistically higher (P < 0.05) in the D medium (0.549?±?0.044 g L^?1) than in BG11 (0.336?±?0.009 g L^?1). On the contrary, the biomass production was found to be higher in BG11 (1.019?±?0.051 g L^?1) than in the D medium (0.953?±?0.056 g L^?1). However, this difference was not statistically significant. The difference in salinity and nitrogen concentration between both media is suggested as the main factor involved in the EPS and biomass results. FTIR spectra of B. braunii EPS from both media revealed presence of uronic acids and absence of amino and sulfate groups. Despite the similarity between both spectra, there were some different signals (at 1,921.52 and 720.60 cm^?1) which may mean a difference in glycosyl composition.     

A novel exopolysaccharide from deep-sea bacterium Zunongwangia profunda SM-A87: low-cost fermentation,moisture retention,and antioxidant activities

Many marine microorganisms can secrete exopolysaccharides (EPSs) which have important applications in biotechnology. We have purified a novel EPS from deep-sea bacterium Zunongwangia profunda SM-A87, identified its glycosyl composition and linkage, and optimized its production to 8.9 g/l in previous studies. To reduce the fermentation cost, an economical fermentation medium containing 60.9 % whey, 10 g/l soybean meal, and 2.9 % NaCl was developed. The EPS yield of batch fermentation in this medium reached 12.1?±?0.3 g/l. Fed-batch fermentation was conducted and led to an EPS yield of 17.2?±?0.4 g/l, which represents the highest EPS yield ever reported for a marine bacterium. The EPS was extracted and it displayed good rheological properties, moisture-retention ability, and antioxidant activity. Particularly, its moisture-retention ability is superior to that of other marine bacterial EPSs reported to date. SM-A87 EPS also showed high antioxidant activity. These results suggest that SM-A87 EPS has promising potentials in biotechnology.     

Strategy for pH control and pH feedback-controlled substrate feeding for high-level production of l-tryptophan by Escherichia coli

Optimum production of l-tryptophan by Escherichia coli depends on pH. Here, we established conditions for optimizing the production of l-tryptophan. The optimum pH range was 6.5–7.2, and pH was controlled using a three-stage strategy [pH 6.5 (0–12 h), pH 6.8 (12–24 h), and pH 7.2 (24–38 h)]. Specifically, ammonium hydroxide was used to adjust pH during the initial 24 h, and potassium hydroxide and ammonium hydroxide (1:2, v/v) were used to adjust pH during 24–38 h. Under these conditions, NH₄ ⁺ and K⁺ concentrations were kept below the threshold for inhibiting l-tryptophan production. Optimization was also accomplished using ratios (v/v) of glucose to alkali solutions equal to 4:1 (5–24 h) and 6:1 (24–38 h). The concentration of glucose and the pH were controlled by adjusting the pH automatically. Applying a pH-feedback feeding method, the steady-state concentration of glucose was maintained at approximately 0.2 ± 0.02 g/l, and acetic acid accumulated to a concentration of 1.15 ± 0.03 g/l, and the plasmid stability was 98 ± 0.5 %. The final, optimized concentration of l-tryptophan was 43.65 ± 0.29 g/l from 52.43 ± 0.38 g/l dry cell weight.     

Eu3+ Sequestration by Biogenic Nano-Hydroxyapatite Synthesized at Neutral and Alkaline pH

Biogenic hydroxyapatite (bio-HA) has the potential for radionuclide capture and remediation of metal-contaminated environments. Biosynthesis of bio-HA was achieved via the phosphatase activity of a Serratia sp. supplemented with various concentrations of CaCl₂ and glycerol 2-phosphate (G2P) provided at pH 7.0 or 8.6. Presence of hydroxyapatite (HA) was confirmed in the samples by X-ray powder diffraction analysis. When provided with limiting (1 mM) G2P and excess (5 mM) Ca²⁺ at pH 8.6, monohydrocalcite was found. This, and bio-HA with less (1 mM) Ca²⁺ accumulated Eu(III) to ～31% and 20% of the biomineral mass, respectively, as compared to 50% of the mineral mass accumulated by commercial HA. Optimally, with bio-HA made at initial pH 7.0 from 2 mM Ca²⁺ and 5 mM G2P, Eu(III) accumulated to ～74% of the weight of bio-HA, which was equal to the mass of the HA mineral component of the biomaterial. The implications with respect to potential bio-HA-barrier development in situ or as a remediation strategy are discussed.     

Medium optimization, molecular characterization, and bioactivity of exopolysaccharides from Pleurotus eryngii

An optimal medium for exopolysaccharides (EPS) production was obtained through one-factor-at-a-time method and response surface methodology. Under optimal culture medium, the maximum EPS concentration in shake flask was 5.16 g/l. Two groups of EPSs (designated as Fr-I and Fr-II) were obtained from the culture filtrates by size exclusion chromatography/multiangle laser light scattering, and the weight average molar masses (M _w) of Fr-I and Fr-II were determined to be 4.098 × 10⁴ and 1.114 × 10⁴ g/mol, respectively. The molecular confirmation of Fr-I was revealed to be a rigid rod form in aqueous solution. Moreover, monosaccharide composition and characteristic groups were investigated by GC and Fourier transform infrared, respectively. Finally, pharmacology experiment in vitro indicated EPS Fr-II of Pleurotus eryngii exhibited higher antioxidant and antitumor abilities than Fr-I, which might be attributed to the different molecular weights and chemical compositions in the EPS fraction.     

Production and Degradation of Oxalic Acid by Brown Rot Fungi

Our results show that all of the brown rot fungi tested produce oxalic acid in liquid as well as in semisolid cultures. Gloeophyllum trabeum, which accumulates the lowest amount of oxalic acid during decay of pine holocellulose, showed the highest polysaccharide-depolymerizing activity. Semisolid cultures inoculated with this fungus rapidly converted ¹⁴C-labeled oxalic acid to CO₂ during cellulose depolymerization. The other brown rot fungi also oxidized ¹⁴C-labeled oxalic acid, although less rapidly. In contrast, semisolid cultures inoculated with the white rot fungus Coriolus versicolor did not significantly catabolize the acid and did not depolymerize the holocellulose during decay. Semisolid cultures of G. trabeum amended with desferrioxamine, a specific iron-chelating agent, were unable to lower the degree of polymerization of cellulose or to oxidize ¹⁴C-labeled oxalic acid to the extent or at the rate that control cultures did. These results suggest that both iron and oxalic acid are involved in cellulose depolymerization by brown rot fungi.     

Culture and exopolysaccharide production from sugarcane molasses by Gordonia polyisoprenivorans CCT 7137, isolated from contaminated groundwater in Brazil

Gordonia polyisoprenivorans CCT 7137 was isolated from groundwater contaminated with leachate in an old controlled landfill (São Paulo, Brazil), and cultured in GYM medium at different concentrations of sugarcane molasses (2%, 6%, and 10%). The strain growth was analyzed by monitoring the viable cell counts (c.f.u. mL^?1) and optical density and EPS production was evaluated at the end of the exponential phase and 24 h after it. The analysis of the viable cell counts showed that the medium that most favored bacterial growth was not the one that favored EPS production. The control medium (GYM) was the one that most favored the strain growth, at the maximum specific growth rate of 0.232 h^?1. Differences in bacterial growth when cultured at three different concentrations of molasses were not observed. Production of EPS, in all culture media used, began during the exponential phase and continued during the growth stationary phase. The highest total EPS production, after 24 h of stationary phase, was observed in 6% molasses medium (172.86 g L^?1) and 10% (139.47 g L^?1) and the specific total EPS production was higher in 10% molasses medium (39.03 × 10^?11 g c.f.u.^?1). After the exponential phase, in 2%, 6%, and 10% molasses media, a higher percentage of free exopolysaccharides (EPS) was observed, representing 88.4%, 62.4%, and 64.2% of the total, respectively. A different result was observed in pattern medium, which presented EPS made up of higher percentage of capsular EPS (66.4% of the total). This work is the first study on EPS production by G. polyisoprenivorans strain in GYM medium and in medium utilizing sugarcane molasses as the sole nutrient source and suggests its potential use for EPS production by G. polyisoprenivorans CCT 7137 aiming at application in biotechnological processes.     

Correlation between oxalic acid production and tolerance of Tyromyces palustris strain TYP-6137 to N′,N-naphthaloylhydroxamine

Eleven strains of T. palustris were evaluated for mass loss and production of phosphate buffer soluble oxalic acid on pine wood blocks treated with 0.5% N′,N-naphthaloylhydroxamine (NHA) in a soil-block test. After 12 weeks higher percentage mass loss was observed in control groups for 10 strains, while TYP-6137 was shown to be tolerant with no difference between the untreated controls and 0.5% NHA-treated blocks. Strain TYP-6137 was evaluated at 6, 8, and 10 weeks for mass loss and production of oxalic acid in a colorimetric assay with increasing levels of NHA treatment (0.5–1%). Tyromyces palustris TYP-6137 was shown to be successful in degrading NHA-treated wood in the 0.5–0.8% treatment range, but was inhibited in the 0.9–1% treated groups. Although there is no simple linear correlation between mass loss and production of oxalic acid, there was an inverse relationship at the 6-week time period. Thus, it was concluded that production of oxalic acid in T. palustris TYP-6137 at 6 weeks may facilitate mass loss at 8–10 weeks in the 0.5–0.8% NHA-treated blocks. In the 0.9–1% treatment groups this response was not seen. The broad variation seen in oxalic acid production among the 11 T. palustris strains suggests that high oxalic acid production alone is not the sole mechanism of NHA tolerance. Oxalic acid appears to be non-specifically produced in response to the preservative treatment, yet its association with mass loss could not be clearly defined.     

Growth conditions of clostridium perfringens type B for production of toxins used to obtain veterinary vaccines

The diseases caused for Clostridium perfringens are generically called enterotoxemias because toxins produced in the intestine may be absorbed into the general circulation. C. perfringens type B, grown in batch fermentation, produced toxins used to obtain veterinary vaccines. Glucose in concentrations of 1.4–111.1 mM was used to define the culture medium. The minimum concentration for a satisfactory production of vaccines against clostridial diseases was 55.6 mM. Best results were brought forth by meat and casein peptones, both in the concentration 5.0 g l^?1 in combination with glucose and a culture pH maintained at 6.5 throughout the fermentation process. The production of lactic, acetic and propionic organic acids was observed. Ethanol was the metabolite produced in the highest concentration when cultures maintained steady pH of 6.5 with exception of cultures with initial glucose concentration of 1.4 mM, where the highest production was of propionic acid. Maximal cell concentration and the highest toxin title concomitantly low yield coefficient to organic acids and ethanol were obtained using basal medium containing 111.1 mM glucose under a controlled pH culture (pH) 6.5 in batch fermentations of C. perfringens type B. These data contribute to improve process for industrial toxin production allowing better condition to produce a toxoid vaccine.     

Exopolysaccharide production in <Emphasis Type="Italic">Anabaena</Emphasis> sp. PCC 7120 under different CaCl<Subscript>2</Subscript> regimes

Influence of various levels of CaCl₂ (0, 1, 10 and 100 mM) on exopolysaccharide production has been investigated in the cyanobacterium Anabaena 7120. At the concentration of 1 mM CaCl₂, growth was found to be stimulatory while 100 mM was sub lethal for the cyanobacterial cells. Estimation of EPS content revealed that EPS production depends on the concentration of calcium ions in the immediate environment with maximum being at10 mM CaCl₂. A possible involvement of alr2882 gene in the process of EPS production was also revealed through qRT-PCR. Further, FTIR-spectra marked the presence of aliphatic alkyl-group, primary amine-group, and polysaccharides along with shift in major absorption peaks suggesting that calcium levels in the external environment regulate the composition of EPS produced by Anabaena 7120. Thus, both quantity and composition of EPS is affected under different calcium chloride concentrations presenting possibilities of EPS with novel unexplored features that may offer biotechnological applications.     

Production and characterisation of exopolysaccharide from Streptomyces carpaticus isolated from marine sediments in Egypt and its effect on breast and colon cell lines

Twenty streptomycete strains were isolated from marine sediment samples collected from Nabq area, Sharm El-Sheikh, Red Sea Coast, Egypt. Four of them produce exopolysaccharides (EPS) showing marked in vitro antitumor activities. Morphological and cultural characteristics of the most significant strain (No. 3) were shown. Moreover, the sequence of this strain showed similarity with Streptomyces carpaticus. The results reveal that EPS produced by Streptomyces carpaticus No. 3 had high cytotoxicity reaching 51.7% and 59.1% against human tumor cells of breast and colon lines respectively. A chemical analysis of EPS indicated that the composing monosaccharides were galactouronic acid, glucose, xylose, galactose, mannose, and fructose with relative ratio of 3:1:1:2:2:1 respectively, with an average molecular weight (Mw) 1.180 × 10⁵?g/mol and of a number average molecular weight (Mn) 1.052 × 10⁵?g/mol. Also the EPS contained uronic acid (0.5072%) and monosaccharide sulphates (21.753%).     

Responses of unsaturated Pseudomonas putida CZ1 biofilms to environmental stresses in relation to the EPS composition and surface morphology

The extracellular polymeric substance (EPS) and surface properties of unsaturated biofilms of a heavy metal-resistant rhizobacterium Pseudomonas putida CZ1, in response to aging, pH, temperature and osmotic stress, were studied by quantitative analysis of EPS and atomic force microscope. It was found that EPS production increased approximately linearly with culture time, cells in the air-biofilm interface enhanced EPS production and decreased cell volume to cope with nutrient depletion during aging. Low pH, high temperature and certain osmotic stress (120 mM NaCl) distinctly stimulated EPS production, and the main component enhanced was extracellular protein. In addition to the enhancement of EPS production in response to high osmotic (328 mM NaCl) stress, cells in the biofilm adhere tightly together to maintain a particular microenvironment. These results indicated the variation of EPS composition and the cooperation of cells in the biofilms is important for the survival of Pseudomonas putida CZ1 from environmental stresses in the unsaturated environments such as rhizosphere.     

Cryoprotective properties and preliminary characterization of exopolysaccharide (P-Arcpo 15) produced by the Arctic bacterium Pseudoalteromonas elyakovii Arcpo 15

Twenty-two bacterial strains that secrete exopolysaccharides (EPS) were isolated from marine samples obtained from the Chukchi Sea in the Arctic Ocean; of these, seven strains were found to be capable of producing cryoprotective EPS. The ArcPo 15 strain was isolated based on its ability to secrete large amounts of EPS, and was identified as Pseudoalteromonas elyakovii based on 16S rDNA analysis. The EPS, P-ArcPo 15, was purified by protease treatment and gel filtration chromatography. The purified EPS (P-ArcPo 15) had a molecular mass of 1.7 × 10⁷ Da, and its infrared spectrum showed absorption bands of hydroxyl and carboxyl groups. The principal sugar components of P-ArcPo 15 were determined to be mannose and galacturonic acid, in the ratio of 3.3:1.0. The cryoprotective properties of P-ArcPo 15 were characterized by an Escherichia coli viability test. In the presence of 0.5% (w/v) EPS, the survival percentage of E. coli cells was as high as 94.19 ± 7.81% over five repeated freeze–thaw cycles. These biochemical characteristics suggest that the EPS P-ArcPo 15 may be useful in the development of cryoprotectants for biotechnological purposes, and we therefore assessed the utility of this novel cryoprotective EPS.     

Aureobasidium pullulans batch cultivations based on a factorial design for improving the production and molecular weight of exopolysaccharides

Six factors: strain, carbon source, nitrogen source, nitrogen concentration, aeration, and initial pH, were investigated for their effects on exopolysaccharides (EPS) production in Erlenmeyer flasks by Aureobasidium pullulans using 2-level fractional factorial design. The concentration and molecular weight (MW) of EPS were optimized simultaneously. The effects of main factors together with possible two-factor interactions were detected, and the levels of the six factors were optimized using empirical models. Analysis of factor effects revealed that strain had the strongest influence on EPS concentration, and nitrogen source on MW of EPS. However, nitrogen concentration did not show significant influence on both parameters. The influences of factors on the production variability were also monitored for quality control purposes. At the optimum levels of control factors investigated, as high as 22.6 g/L EPS with a weight average MW of higher than 2 × 10⁶ was produced. The results of this work indicate that the production kinetics varies for different microorganism–medium–environment systems.     

Selection and Optimization of Culture Medium for Exopolysaccharide Production by Coriolus (Trametes) Versicolor

Summary Coriolus versicolor is a medicinal fungus producing exopolysaccharides (EPS). Five well-defined culture media were studied to select the medium that maximizes production of EPS by C. versicolor. Biomass, reducing sugars and EPS concentrations along with the rheological behaviour of the broth were followed during fermentations lasting 9 days. The yeast malt extract medium (YM) was shown to yield the highest production of EPS. Fermentation conditions with YM medium were further investigated to optimize EPS production by C. versicolor. An experimental design to do this was adopted, in which the effects of pH and initial substrate concentration were considered. The effects of initial glucose concentration (5, 15 and 25 g l⁻¹) and pH (4.0, 5.5 and 7.0) were evaluated. The initial glucose concentration was found to be the most important factor in EPS production and also cell growth.