Aeroponic system for control of root-zone atmosphere

Control of rhizospheric gases can be critical to research. Culturing plants with roots in liquid is a common component of protocols described previously for controlling the root-zone atmosphere, and liquid culture is routine during assays of symbiotic N₂ fixation. However, spurious data may result from gases becoming trapped within the boundary layer of solution-cultured roots and nodules. Aeroponics is a viable alternative to other soilless culture systems for maintaining plants with a controlled root-zone atmosphere. We have designed an aeroponic gas-delivery system that allows statistical treatment of individual plants as discrete experimental units when root-zone-targeted treatments are imposed. Roots of intact plants were held in closed 1-L canning jars, with one plant per jar and the stem emerging from a hole drilled in the jar lid. Extra-fine fog nozzles controlled by a programmable timer sprayed roots periodically with nutrient solution. Gases were mixed and delivered to jars at a prescribed rate by using a system of capillary tubes. During a study of O₂ effects on formation of root nodules, 100% survival of plants and profuse nodulation resulted after 4 weeks. The pO₂ within each jar remained within ±1.0% of its prescribed value, and pre-and post-experimental O₂ measurements within individual jars did not differ (P < 0.0001). The system is easy to construct with inexpensive materials, has many applications for rhizosphere biology, and is unique among systems used previously.     

Immobilization of the hyperthermophilic hydrogenase from Aquifex aeolicus bacterium onto gold and carbon nanotube electrodes for efficient H2 oxidation

The electrochemistry of membrane-bound [NiFe] hydrogenase I ([NiFe]-hase I) from the hyperthermophilic bacterium Aquifex aeolicus was investigated at gold and graphite electrodes. Direct and mediated H₂ oxidation were proved to be efficient in a temperature range of 25–70 °C, describing a potential window for H₂ oxidation similar to that of O₂-tolerant hydrogenases. Search for enhancement of current densities and enzyme stability was achieved by the use of carbon nanotube coatings. We report high catalytic currents for H₂ oxidation up to 1 mA cm⁻², 10 times higher than at the bare electrode. Interestingly, high stability of the direct catalytic process was observed when encapsulating A. aeolicus [NiFe]-hase I into a carboxylic functionalized single walled carbon nanotube network. This suggests a peculiar interaction between the enzyme and the electrode material. The parameters that governed the orientation of the enzyme before electron transfer were thus investigated using self-assembled-monolayer gold electrodes. No control of the orientation by the charge or the hydrophobicity of the interface was demonstrated. This behavior was explained on the basis of a structural comparison between A. aeolicus [NiFe]-hase I and Desulfovibrio fructosovorans [NiFe] hydrogenase, which revealed the absence of acidic residues and an additional loop in the environment of the [4Fe–4S] distal cluster in A. aeolicus [NiFe]-hase I. Finally, the effect of inhibitors on the direct oxidation of H₂ by A. aeolicus [NiFe]-hase I encapsulated in a single walled carbon nanotube network was investigated. No inhibition by CO and tolerance toward O₂ were observed. Discussion of the reasons for such tolerance was undertaken on the basis of structural comparison with hydrogenases from aerobic bacteria.     

Culture of Amelanchier x grandiflora in a programmable micropropagation apparatus

A micropropagation system was developed to test concepts for automation and microenvironment alteration. Amelanchier x grandiflora Rehd. Princess Diana (serviceberry) shoot cultures were grown in seven-liter polycarbonate containers. Through computer control, the apparatus intermittently applied culture medium to the plant material according to a selected schedule. Shoot growth in the programmable system was compared with four micropropagation treatments: gelled and liquid medium in baby food jars and gelled and non-cycling liquid medium in a seven-liter vessel. Plants cultured in continuous contact with liquid medium became increasingly vitrified. Significantly greater shoot number, shoot length, shoot weight, and culture weight occurred in the programmable system than in jars with gelled medium. The combination of liquid medium, 7-liter vessel, and intermittent contact with medium caused a significantly higher proliferation rate than any combination of jar/vessel and gelled/liquid medium tested.     

Growth and nitrogen fixation inAlnus glutinosa (L.) Gaertn. under carbon dioxide enrichment of the root atmosphere

The effects of aeration of the N-free rooting medium with elevated CO₂ on (a) acetylene reduction by perlite-grown plants and (b) N₂-fixation and long-term growth of nutrient solution-grown plants were determined for nodulatedAlnus glutinosa (L.) Gaertn. In the former experiments, roots of intact plants were incubated in acetylene in air in darkened glass jars for 3 hr, followed by a further 3 hr incubation period in air enriched with CO₂ (0–5%). During incubation, the CO₂ content of the jars increased by 0.17% per hour due to respiration of the root system, so that the CO₂ content at 3 hr was 0.5%. Additional enrichment of the rooting medium gas-phase with CO₂ equivalent to 1.1% and 1.75% CO₂ of the gas volume significantly increased nitrogenase activity (ethylene production) by 55% and 50% respectively, while enrichment with greater than 2.5% CO₂ decreased activity. In contrast, ethylene production by control plants, where CO₂ was not added to the assay jars, decreased by 8% over the assay period. In long-term growth experiments, nodulated roots of intactAlnus glutinosa plants were sealed into jars containing N-free nutrient solution (pH 6.3) and aerated with air, or air containing elevated levels of CO₂ (1.5% and 5%). Comparison of the appearance of CO₂-treated with air treated plants suggested that 1.5% CO₂ stimulated plant growth. However, at harvest after 5 or 6 weeks variability between plants masked the significance of differences in plant dry weight. A significant increase of 33% in total nitrogen of plants aerated with 1.5% CO₂, compared with air-treated plants, was demonstrated, broadly in line with the short-term increase in acetylene reducing activity observed following incubations with similar CO₂ concentrations. Shoot dry weight was not affected significantly by long-term exposure to 5% CO₂, the main effect on growth being a 20% reduction in dry weight of the root system, possibly through inhibition of root system respiration. However, in contrast to the inhibitory effects of high CO₂ on acetylene reduction there was no significant effect on the amounts of N₂ fixed.     

Unexpected Elevated Production of Aquifex aeolicus Cytochrome c 555 in Escherichia coli Cells Lacking Disulfide Oxidoreductases

Mutant strains of Escherichia coli lacking DsbA, DsbB, or DsbD (proteins required for disulfide bond formation in the periplasm) did not produce mitochondrial or chloroplast cytochromes c, as previously observed for bacterial ones. Unexpectedly, however, cytochrome c ₅₅₅ (AA c ₅₅₅) from a hyperthermophile, Aquifex aeolicus, was produced in the E. coli periplasm without Dsb proteins, three times more than with them. These results indicate that the Dsb proteins are not necessarily required for AA c ₅₅₅ production in E. coli, possibly because of hyperthermophilic origin compared with the others.     

Isolation and Characterization of a Hydrogen Sulfide-Removing Bacterium,Pseudomonas sp. Strain DO-1

Five microbial strains that removed hydrogen sulfide (H₂S) or methylmercaptan (CH₃SH) gas were newly isolated from soil samples. Strain DO-1, one of the isolates, was identified as a member of Pseudomonas sp., and it’s immobilized cells removed 1 or 10 ppm of H₂S gas within 2 hours. When strain DO-1 was cultured aerobically in a flask containing nutrient broth medium, the deodorizing activity increased, depending on the growth of the culture, and the maximum activity was obtained after 48 hours. Even though the immobilized cells were stored at 4 or 25°C in sealed bottles for 6 months, the deodorizing activity remained. Throughout this study, strain DO-1 removed H₂ S gas without preliminary feeding or exposure to sulfur com-pounds as growth substrates or inducers. These characteristics are advantageous for the deodorization of the malodorous gases surrounding us in daily life.     

Microplates as a microreactor platform for microalgae research

High‐throughput platforms for microalgae screening are not yet commercially available. In this study, the feasibility of 96‐well microplates was analyzed for microalgae research. Equivalence among wells, as culture microreactors, was investigated in controlled high CO₂ conditions. Specific growth rates of two microalgae species, Scenedesmus sp. UTEX1589 and an environmental isolate, were significantly higher in border wells than in internal positions. Furthermore, growth rate gradients analyzed as contours throughout the platform were observed for Scenedesmus sp. However, the output variable exhibited high precision associated with a low coefficient of variation (CV), between 6.8 and 7.8%. In a demonstrative experiment to determine the effect of culture media dilution on six microalgae species, treatments were randomized in the central subset of a microplate. Results were consistent and statistically sound (CV 9.4–12.9%), and showed that microalgae species could grow with no detrimental effect in 50% (v/v) dilution of the culture medium. Provided border wells exclusion and a randomized design, 96‐well microplates are a practical and statistical robust platform for microalgae research. © 2013 American Institute of Chemical Engineers Biotechnol. Prog., 29:638–644, 2013     

Intersubunit Disulfide Interactions Play a Critical Role in Maintaining the Thermostability of Glucose-6-phosphate Dehydrogenase from the Hyperthermophilic Bacterium Aquifex aeolicus

Proteins from thermophilic microorganisms are stabilized by various mechanisms to preserve their native folded states at higher temperatures. A thermostable glucose-6-phosphate dehydrogenase (tG6PDH) from the hyperthermophilic bacterium Aquifex aeolicus was expressed as a recombinant protein in Escherichia coli. The A. aeolicus G6PDH is a homodimer exhibiting remarkable thermostability (t_1/2=24 hr at 90°C). Based on homology modeling and upon comparison of its structure with human G6PDH, it was predicted that cysteine 184 of one subunit could form a disulfide bond with cysteine 352 of the other subunit resulting in reinforced intersubunit interactions that hold the dimer together. Site-directed mutagenesis was performed on tG6PDH to convert C184 and C352 to serines. The tG6PDH double mutant exhibited a dramatic decrease in the half-life from 24 hr to 3 hr at 90°C. The same decrease in half-life was also found when either C184 or C352 was mutated to serine. The result indicates that C184 and C352 may play a crucial role in strengthening the dimer interface through disulfide bond formation, thereby contributing to the thermal stability of the enzyme.     

First characterisation of the active oligomer form of sulfur oxygenase reductase from the bacterium Aquifex aeolicus

Sulfur oxygenase reductase (SOR) enzyme is responsible for the initial oxidation step of elemental sulfur in archaea. Curiously, Aquifex aeolicus, a hyperthermophilic, chemolithoautotrophic and microaerophilic bacterium, has the SOR-encoding gene in its genome. We showed, for the first time the presence of the SOR enzyme in A. aeolicus, its gene was cloned and recombinantly expressed in Escherichia coli and the protein was purified and characterised. It is a 16 homo-oligomer of approximately 600 kDa that contains iron atoms indispensable for the enzyme activity. The optimal temperature of SOR activity is 80°C and it is inactive at 20°C. Studies of the factors involved in getting the fully active molecule at high temperature show clearly that (1) incubation at high temperature induces more homogeneous form of the enzyme, (2) conformational changes observed at high temperature are required to get the fully active molecule and (3) acquisition of an active conformation induced by the temperature seems to be more important than the subunit number. Differences between A. aeolicus SOR and the archaea SORs are described.     

Interactions of culture vessels, media volume, culture density, and carbon dioxide levels on lettuce and spearmint shoot growth in vitro

 The influence of culture chamber capacity, medium volume and culture density on the growth yields of lettuce (Lactuca sativa L.) and spearmint (Mentha spicata L.) shoots were determined in an environment containing either 350 or 10,000 μmol mol^–1 CO₂ after 8 weeks of incubation. High positive correlations occurred between the culture vessel capacity and spearmint fresh weight, leaf number, root number, and shoot number. Similarly, high positive correlations occurred between culture vessel capacity and lettuce fresh weight, leaf number, and root number. Higher fresh weights, leaf numbers, and root numbers were obtained from lettuce and spearmint shoots when cultured in 1-quart Mason jars containing 100- or 150-ml aliquots of medium compared to jars containing 25- or 50-ml aliquots of medium within an environment containing either 350 or 10,000 μmol mol^–1 CO₂. High culture density decreased growth yields, and this phenomenon could only be slightly off-set by the employment of an elevated CO₂ environment or larger culture vessels. Received: 22 December 1998 / Revision received: 2 July 1999 / Accepted: 12 July 1999     

Mixed culture hydrogenotrophic nitrate reduction in drinking water

Isolation and identification of the bacteria from a hydrogenotrophic reactor for the denitrification of drinking water revealed that several microorganisms are involved. Acinetobacter sp., Aeromonas sp., Pseudomonas sp. and Shewanella putrefaciens were repeatedly isolated from the hydrogenotrophic sludge and postulated to be of primary importance in the process. Nitrate reduction to nitrite appears to be a property of a diverse group of organisms. Nitrite reduction was found to be stimulated by the presence of organic growth factors. Thus, in a mixed culture, hydrogenotrophic denitrification reactor, NO _inf2 ^sup– formed by NO _inf3 ^sup– -reducers can be converted by true denitrifiers thriving on organic growth factors either present in the raw water, or excreted by the microbial community. Mixotrophic growth also contributes to NO _inf2 ^sup– reduction. Finally, chemolithotrophic bacteria participate in the nitrite to nitrogen gas conversion.Offprint requests to: W. Verstraete.     

Production of fully assembled and active Aquifex aeolicus F1FO ATP synthase in Escherichia coli

Background

F₁F_O ATP synthases catalyze the synthesis of ATP from ADP and inorganic phosphate driven by ion motive forces across the membrane. A number of ATP synthases have been characterized to date. The one from the hyperthermophilic bacterium Aquifex aeolicus presents unique features, i.e. a putative heterodimeric stalk. To complement previous work on the native form of this enzyme, we produced it heterologously in Escherichia coli.

Methods

We designed an artificial operon combining the nine genes of A. aeolicus ATP synthase, which are split into four clusters in the A. aeolicus genome. We expressed the genes and purified the enzyme complex by affinity and size-exclusion chromatography. We characterized the complex by native gel electrophoresis, Western blot, and mass spectrometry. We studied its activity by enzymatic assays and we visualized its structure by single-particle electron microscopy.

Results

We show that the heterologously produced complex has the same enzymatic activity and the same structure as the native ATP synthase complex extracted from A. aeolicus cells. We used our expression system to confirm that A. aeolicus ATP synthase possesses a heterodimeric peripheral stalk unique among non-photosynthetic bacterial F₁F_O ATP synthases.

Conclusions

Our system now allows performing previously impossible structural and functional studies on A. aeolicus F₁F_O ATP synthase.

General significance

More broadly, our work provides a valuable platform to characterize many other membrane protein complexes with complicated stoichiometry, i.e. other respiratory complexes, the nuclear pore complex, or transporter systems.     

Carbon metabolism inSulfobacillus thermosulfidooxidans subsp.asporogenes, strain 41

The activities of carbon metabolism enzymes were determined in cellular extracts of the moderately thermophilic, chemolithotrophic, acidophilic bacteriumSulfobacillus thermosulfidooxidans subsp.asporogenes, strain 41, grown either at an atmospheric content of CO₂ in the gas phase (autotrophically, heterotrophically, or mixotrophically) or autotrophically at a CO₂ content increased to 5–10%. Regardless of the growth conditions, all TCA cycle enzymes (except for 2-oxoglutarate dehydrogenase), one glyoxylate bypass enzyme (malate synthase), and some carboxylases (ribulose bisphosphate carboxylase, pyruvate carboxylase, and phosphoenolpyruvate carboxylase) were detected in the cell-free extracts of strain 411. During autotrophic cultivation of strains 41 and 1269, the increase in the CO2 content of the supplied air to 5–10% resulted in the activation of growth and iron oxidation, a 20–30% increase in the cellular content of protein, enhanced activity of the key TCA enzymes (citrate synthase and aconitase), and, in strain 41, a decrease in the activity of carboxylases.     

Comparison of Macroscopic Examination, Routine Gram Stains, and Routine Subcultures in the Initial Detection of Positive Blood Cultures

Blood was cultured in two vaccum bottles containing Columbia broth with sodium polyanethol sulfonate and CO₂. Filtered air was admitted to one bottle, and the bottles were incubated at 35 C until growth was detected or for a maximum of 7 days. Bottles were examined daily for macroscopic growth. Gram stains were made routinely on the 1st, 4th, and 7th days, and samples were routinely subcultured to sheep blood agar (incubated in GasPak jar) and chocolate agar (incubated in CO₂) on the 1st and 4th days of incubation. Of 1,127 positive blood cultures, 65% were first detected by macroscopic examination, 23% were first detected by Gram stain, and 12% were first detected only by subculture.     

Linking geochemical processes with microbial community analysis: successional dynamics in an arsenic-rich, acid-sulphate-chloride geothermal spring

The source waters of acid‐sulphate‐chloride (ASC) geothermal springs located in Norris Geyser Basin, Yellowstone National Park contain several reduced chemical species, including H₂, H₂S, As(III), and Fe(II), which may serve as electron donors driving chemolithotrophic metabolism. Microorganisms thriving in these environments must also cope with high temperatures, low pH (～3), and high concentrations of sulphide, As(III), and boron. The goal of the current study was to correlate the temporal and spatial distribution of bacterial and archaeal populations with changes in temperature and geochemical energy gradients occurring throughout a newly formed (redirected) outflow channel of an ASC spring. A suite of complimentary analyses including aqueous geochemistry, microscopy, solid phase identification, and 16S rDNA sequence distribution were used to correlate the appearance of specific microbial populations with biogeochemical processes mediating S, Fe, and As cycling and subsequent biomineralization of As(V)‐rich hydrous ferric oxide (HFO) mats. Rapid As(III) oxidation (maximum first order rate constants ranged from 4 to 5 min^?1, t_1/2 = 0.17 ? 0.14 min) was correlated with the appearance of Hydrogenobaculum and Thiomonas–like populations, whereas the biogenesis of As(V)‐rich HFO microbial mats (mole ratios of As:Fe ～0.7) was correlated with the appearance of Metallosphaera, Acidimicrobium, and Thiomonas–like populations. Several 16S sequences detected near the source were closely related to sequences of chemolithotrophic hyperthermophilic populations including Stygiolobus and Hydrogenobaculum organisms that are known H₂ oxidizers. The use of H₂, reduced S(–II,0), Fe(II) and perhaps As(III) by different organisms represented throughout the outflow channel was supported by thermodynamic calculations, confirming highly exergonic redox couples with these electron donors. Results from this work demonstrated that chemical energy gradients play an important role in establishing distinct community structure as a function of distance from geothermal spring discharge.     

Chemolithotrophic growth of the phototrophic sulfur bacterium Thiocapsa roseopersicina

Chemotrophic growth capacities of the purple sulfur bacterium Thiocapsa roseopersicina strain M1 were studied in continuous culture under thiosulfate limitation.Pigment synthesis was completely inhibited upon a shift from anaerobic to semi-aerobic conditions (52 μM O₂) in the light, but no active breakdown occurred. During the transient state, the cells grew in a mixed photo- and chemolithotrophic mode; the specific respiration rate gradually increased with a concomitant drop in the bacteriochlorophyll a content. Photolithotrophically grown cells have the ability to respire. It was concluded that photosynthesis and respiration compete for electrons, but that photosynthesis is preferred under electron donor-limiting conditions, when the cells still contain large amounts of pigments. Eventually, a fully chemolithotrophic steady state was attained.The chemolithotropic growth of T. roseopersicina was studied in the dark under semiaerobic conditions at various dilution rates. The maximum specific growth rate was 68% of the maximum attainable growth rate under photolithotrophic conditions. The growth affinity for thiosulfate was high (K_m = 1.5 μM). The yield on thiosulfate under chemolithotrophic conditions exceeded that of thiobacilli. Oxygen uptake was studied in short-term experiments. It was shown that respiration in T. roseopersicina has a K_m of approx. 1 μM O₂. the ecological importance for T. roseopersicina of chemolithotrophic growth and pigment content is discussed with respect to the occurrence of T. roseopersicina in laminated microbial ecosystems and its possible competition with colorless sulfur bacteria.     

Physical environment in non-ventilated culture vessels affects <Emphasis Type="Italic">in vitro</Emphasis> growth and morphogenesis of several cultivars of <Emphasis Type="Italic">Dianthus Caryophyllus</Emphasis> L.

Summary Differences were shown in the environmental conditions inside four types of non-ventilated culture vessels (glass baby-food jars capped with metal steel or Magenta B-caps, jam glass jars capped with metal steel caps, and Magenta GA7 culture vessels) that were incubated in the same external growth room conditions. Vessel light transmittance varied from 83% to 53% and determined the availability of photosynthetic photon flux density (PPFD) for explants. The mean of medium desiccation in the culture period (30 d) was from 2.8 g in vessels with the lowest rate of gas exchange (0.02 h⁻¹) to 10.4 g in vessels with the highest (1.1 h⁻¹). In all cases, the temperature inside the culture vessel increased during the light period and decreased during the dark period, and relative humidity was close to 100%. Results of in vitro growth and morphogenesis of Dianthus caryophyllus L. evs. Scania, White Sim, Angeline, and Pink Calypso provided evidence that the environmental differences detected inside these four types of non-ventilated culture vessels was sufficient to affect micropropagation, mainly related with the specific sensitivity of each cultivar to the gas exchange and medium desiccation determined by the vessel type.     

AQUALINDERELLA FERMENTANS GEN. ET SP. N., A PHYCOMYCETE ADAPTED TO STAGNANT WATERS. II. ISOLATION,CULTURAL CHARACTERISTICS,AND GAS RELATIONS

This newly described member of the Leptomitales was incapable of growth in air on complex media routinely used to culture its nearest relatives. All attempts to isolate it in pure culture were unsuccessful until it was held in an atmosphere established by burning a candle in a sealed jar. Under these conditions it grew vigorously on various complex media and could be readily propagated in the laboratory in pure as well as in unifungal gross cultures. Like a number of other aquatic phycomycetes, A qualinderella fermentans proved to be a strong acid producer, its cultures requiring frequent neutralization during active growth. Work with pure cultures revealed a combination of gas relationships not described before among the fungi. A high level of atmospheric CO₂ is required, with good growth occurring between 5 and 20%. Growth also takes place under 99% CO₂. Supplemental CO₂ is partially replaceable by succinate. A. fermentans exhibits undiminished growth in an atmosphere of CO₂-supplemented hydrogen, essentially devoid of oxygen (leuco-methylene blue). These features are accompanied by an obligately fermentative energy metabolism. Ecologically, A. fermentans is adapted to an environment poor in oxygen and rich both in carbon dioxide and in fermentable organic matter. Such environments are likely to prevail in the warm, stagnant pools where this water mold has been found growing on submerged fruits. In many aspects of its cultural behavior and metabolism A. fermentans resembles another highly fermentative water mold, Blastocladia (Blastocladiales, Chytridiomycetes). The suggestion is made that in both instances parallel regressive evolution has occurred.     

Ethylene and in vitro rooting of rose shoots

Effects of the ethylene precursor 1-aminocyclopropane-1-carboxylic acid (ACC), ethylene biosynthesis inhibitor, (CoCl₂), and inhibitor of ethylene binding to receptors, 1-methylcyclopropene (1-MCP), on ethylene production and rooting in shoot culture of Rosa hybrida L. cv. Alba meidiland were studied. Additionally, effect of ethylene removal by KMnO₄ and HgClO₄ on rooting was tested. ACC increased ethylene production and delayed root formation, decreased the number of roots per shoot and inhibited root growth. In contrast, inhibition of ethylene production by CoCl₂ accelerated root emergence, and increased the number of roots per shoot. Likewise, removing ethylene from the ambient atmosphere improved root emergence and, increased root number of per shoot and markedly inhibited root growth. Blocking the ethylene receptors by 1-MCP increased ethylene level in the ambient atmosphere and increased both emergence and root formation. Both ethylene biosynthesis and action are involved in the control of rooting. Ethylene concentration in glass jars was too high for root emergence and formation, but was appropriate for root growth. CoCl₂ or 1-MPC can be recommended for regulation of rooting in rose shoot culture, since both emergence and number of roots were improved but root growth was not inhibited.     

Optimization of culture conditions and medium components for the production of mycelial biomass and exo-polysaccharides with Cordyceps militaris in liquid culture

Both crude exo-biopolymers and mycelial biomass, produced by liquid culture of Cordyceps species, are believed to possess several potential health benefits. As a result of its known biological activities, Cordyceps militaris has been extensively characterized in regards to potential medicinal applications. However, optimized liquid culture conditions for enhanced polysaccharide productivity have yet to be developed, which is a necessary step for industrial applications. Therefore, in this study, the liquid culture conditions were optimized for maximal production of mycelial biomass and exo-polysaccharide (EPS) by C. militaris. The effects of medium composition, environmental factors, and C/N ratio were investigated. Among these variables 80 g, glucose; 10 g, yeast extract; 0.5 g, MgSO₄·7H₂O; and 0.5 g, KH₂PO₄ in 1 L distilled water were found to be the most suitable carbon, nitrogen, and mineral sources, respectively. The optimal temperature, initial pH, agitation, and aeration were determined to be 24°C, uncontrolled pH, 200 rpm, and 1.5 vvm, respectively. Under these optimal conditions, mycelial growth in shake flask cultures and 5 L jar bioreactors was 29.43 and 40.60 g/L, respectively, and polysaccharide production in shake flask cultures and 5 L jar bioreactors was 2.53 and 6.74 g/L, respectively.