Non-native liana, <Emphasis Type="Italic">Euonymus fortunei</Emphasis>, associated with increased soil nutrients,unique bacterial communities,and faster decomposition rate

Invasive plants have wide-ranging impacts on native systems including reducing native plant richness and altering soil chemistry, microbes, and nutrient cycling. Increasingly, these effects are found to linger long after removal of the invader. We examined how soil chemistry, bacterial communities, and litter decomposition varied with cover of Euonymus fortunei, an invasive evergreen liana, in two central Kentucky deciduous forests. In one forest, E. fortunei invaded in the late 1990s but invasion remained patchy and we paired invaded and uninvaded plots to examine the associations between E. fortunei cover and our response variables. In the second forest, E. fortunei had completely invaded the forest by 2005; areas where it had been selectively removed by 2010 were paired with an adjacent invaded plot. Where E. fortunei had patchily invaded, E. fortunei patches had up to 3.5× nitrogen, 2.7× carbon, and 1.9× more labile glomalin in soils than uninvaded plots, whereas there were no differences in soil characteristics between invaded and removal plots. In the patchily invaded forest, bacterial community composition varied among invaded and non-invaded plots, whereas bacterial communities did not vary among invaded and removal plots. Finally, E. fortunei leaf litter decomposed faster (k = 4.91 year^?1) than the native liana (k = 3.77 year^?1), Vitis vulpina; decomposition of both E. fortunei and V. vulpina was faster in invaded (k = 7.10 year^?1) than removal plots (k = 4.77 year^?1). Our findings suggest that E. fortunei invasion increases the rate of leaf litter decomposition via high-quality litter, alters the decomposition environment, and shifts in the soil biotic communities associated with a dense mat of wintercreeper. Land managers with limited resources should target the densest mats for the greatest restoration potential and remove wintercreeper patches before they establish dense mats.     

Short-term effects of crop rotation,residue management,and soil water on carbon mineralization in a tropical cropping system

The purpose of this study was to investigate the short-term effects of maize (Zea mays)-fallow rotation, residue management, and soil water on carbon mineralization in a tropical cropping system in Ghana. After 15 months of the trial, maize–legume rotation treatments had significantly (P?C ₀ (μg CO₂–C g^?1) than maize–elephant grass (Pennisetum purpureum) rotations. The C ₀ for maize–grass rotation treatments was significantly related to the biomass input (r?=?0.95; P?=?0.05), but that for the maize–legume rotation was not. The soil carbon mineralization rate constant, k (per day), was also significantly related to the rotation treatments (P?k values for maize–grass and maize–legume rotation treatments were 0.025 and 0.036 day^?1 respectively. The initial carbon mineralization rate, m ₀ (μg CO₂–C g^?1 day ^?1), was significantly (P?θ. The m ₀ ranged from 3.88 to 18.67 and from 2.30 to 15.35 μg CO₂–C g^?1 day^?1 for maize–legume and maize–grass rotation treatments, respectively, when the soil water varied from 28% to 95% field capacity (FC). A simple soil water content (θ)-based factor, f _w, formulated as: \(f_{\text{w}} = \left[ {\frac{{\theta - \theta _{\text{d}} }}{{\theta _{{\text{FC}}} - \theta _{\text{d}} }}} \right]\), where θ _d and θ _FC were the air-dry and field capacity soil water content, respectively, adequately described the variation of the m ₀ with respect to soil water (R ²?=?0.91; RMSE?=?1.6). Such a simple relationship could be useful for SOC modeling under variable soil water conditions.     

Carbon Dioxide and Methane Fluxes From Tree Stems,Coarse Woody Debris,and Soils in an Upland Temperate Forest

Forest soils and canopies are major components of ecosystem CO₂ and CH₄ fluxes. In contrast, less is known about coarse woody debris and living tree stems, both of which function as active surfaces for CO₂ and CH₄ fluxes. We measured CO₂ and CH₄ fluxes from soils, coarse woody debris, and tree stems over the growing season in an upland temperate forest. Soils were CO₂ sources (4.58 ± 2.46 µmol m^?2 s^?1, mean ± 1 SD) and net sinks of CH₄ (?2.17 ± 1.60 nmol m^?2 s^?1). Coarse woody debris was a CO₂ source (4.23 ± 3.42 µmol m^?2 s^?1) and net CH₄ sink, but with large uncertainty (?0.27 ± 1.04 nmol m^?2 s^?1) and with substantial differences depending on wood decay status. Stems were CO₂ sources (1.93 ± 1.63 µmol m^?2 s^?1), but also net CH₄ sources (up to 0.98 nmol m^?2 s^?1), with a mean of 0.11 ± 0.21 nmol m^?2 s^?1 and significant differences depending on tree species. Stems of N. sylvatica, F. grandifolia, and L. tulipifera consistently emitted CH₄, whereas stems of A. rubrum, B. lenta, and Q. spp. were intermittent sources. Coarse woody debris and stems accounted for 35% of total measured CO₂ fluxes, whereas CH₄ emissions from living stems offset net soil and CWD CH₄ uptake by 3.5%. Our results demonstrate the importance of CH₄ emissions from living stems in upland forests and the need to consider multiple forest components to understand and interpret ecosystem CO₂ and CH₄ dynamics.     

Oxygen transfer as a tool for fine-tuning recombinant protein production by <Emphasis Type="Italic">Pichia pastoris</Emphasis> under glyceraldehyde-3-phosphate dehydrogenase promoter

Effects of oxygen transfer on recombinant protein production by Pichia pastoris under glyceraldehyde-3-phosphate dehydrogenase promoter were investigated. Recombinant glucose isomerase was chosen as the model protein. Two groups of oxygen transfer strategies were applied, one of which was based on constant oxygen transfer rate where aeration rate was Q _O/V = 3 and 10 vvm, and agitation rate was N = 900 min^?1; while the other one was based on constant dissolved oxygen concentrations, C _DO = 5, 10, 15, 20 and 40 % in the fermentation broth, by using predetermined exponential glucose feeding with μ _o = 0.15 h^?1. The highest cell concentration was obtained as 44 g L^?1 at t = 9 h of the glucose fed-batch phase at C _DO = 20 % operation while the highest volumetric and specific enzyme activities were obtained as 4440 U L^?1 and 126 U g^?1 cell, respectively at C _DO = 15 % operation. Investigation of specific enzyme activities revealed that keeping C _DO at 15 % was more advantageous with an expense of relatively higher by-product formation and lower specific cell growth rate. For this strategy, the highest oxygen transfer coefficient and oxygen uptake rate were K _L a = 0.045 s^?1 and OUR = 8.91 mmol m^?3 s^?1, respectively.     

Parasitism Rate of <Emphasis Type="Italic">Habrobracon hebetor</Emphasis> to <Emphasis Type="Italic">Ephestia kuehniella</Emphasis> Larvae: Residual Effect of Deltamethrin,Fenvalerate and Azadirachtin

Sublethal concentrations of chemical insecticides may cause changes in some behavioral characteristics of natural enemies such as functional responses. The residual effect of three synthetic insecticides including deltamethrin, fenvalerate and azadirachtin were studied on functional response of Habrobracon hebetor Say to Ephestia kuehniella Zeller larvae. Seven host densities (2, 4, 8, 16, 32, 64 and 96) were used during a 24 h period. The resulting data were appropriately fit to Type II functional response models in all treatments: (1) control (0.0916 h^?1; and T _h  = 0.2011 h); (2) deltamethrin (a = 0.0839 h^?1; and T _h  = 0.3560 h); (3) fenvalerate (a = 0.0808 h^?1 and T _h  = 0.3623 h); and (4) azadirachtin (a = 0.0900 h^?1 and T _h  = 0.2042 h). Maximum theoretical parasitism rate (T/T _h ) was 119.34 estimated for control wasps. There was no significant difference between the values of attack rates (a and a + D _a ) in all treatments while the handling time was statistically affected in female wasps treated with fenvalerate. Our findings will be useful in safe application of these insecticides in pest management programmes.     

Bifunctional recombinant cellulase–xylanase (rBhcell-xyl) from the polyextremophilic bacterium <Emphasis Type="Italic">Bacillus halodurans</Emphasis> TSLV1 and its utility in valorization of renewable agro-residues

The thermostable bifunctional CMCase and xylanase encoding gene (rBhcell-xyl) from Bacillus halodurans TSLV1 has been expressed in Escherichia coli. The recombinant E. coli produced rBhcell-xyl (CMCase 2272 and 910 U L^?1 xylanase). The rBhcell-xyl is a ~62-kDa monomeric protein with temperature and pH optima of 60 °C and 6.0 with T_1/2 of 7.0 and 3.5 h at 80 °C for CMCase and xylanase, respectively. The apparent K _m values (CMC and Birchwood xylan) are 3.8 and 3.2 mg mL^?1. The catalytic efficiency (k _cat/K _m ) values of xylanase and CMCase are 657 and 171 mL mg^?1 min^?1, respectively. End-product analysis confirmed that rBhcell-xyl is a unique endo-acting enzyme with exoglucanase activity. The rBhcell-xyl is a GH5 family enzyme possessing single catalytic module and carbohydrate binding module. The action of rBhcell-xyl on corn cobs and wheat bran liberated reducing sugars, which can be fermented to bioethanol and fine biochemicals.     

Single point mutations enhance activity of <Emphasis Type="Italic">cis</Emphasis>-epoxysuccinate hydrolase

Objectives

To enhance activity of cis-epoxysuccinate hydrolase from Klebsiella sp. BK-58 for converting cis-epoxysuccinate to tartrate.

Results

By semi-saturation mutagenesis, all the mutants of the six important conserved residues almost completely lost activity. Then random mutation by error-prone PCR and high throughput screening were further performed to screen higher activity enzyme. We obtained a positive mutant F10D after screening 6000 mutations. Saturation mutagenesis on residues Phe10 showed that most of mutants exhibited higher activity than the wild-type, and the highest mutant was F10Q with activity of 812 U mg^?1 (k _cat /K _m , 9.8 ± 0.1 mM^?1 s^?1), which was 230 % higher than that of wild-type enzyme 355 U mg^?1 (k _cat /K _m , 5.3 ± 0.1 mM^?1 s^?1). However, the thermostability of the mutant F10Q slightly decreased.

Conclusions

The catalytic activity of a cis-epoxysuccinate hydrolase was efficient improved by a single mutation F10Q and Phe10 might play an important role in the catalysis.

     

A survey of the aeromycota of Sydney and its correspondence with environmental conditions: grass as a component of urban forestry could be a major determinant

A comprehensive survey of airborne fungi has been lacking for the Sydney region. This study determined the diversity and abundance of outdoor airborne fungal concentrations in urban Sydney. Monthly air samples were taken from 11 sites in central Sydney, and culturable fungi identified and quantified. The genus Cladosporium was the most frequently isolated fungal genus, with a frequency of 78 % and a mean density of 335 CFU m^?3. The next most frequently encountered genus was Alternaria, occurring in 53 % of samples with a mean of 124 CFU m^?3. Other frequently identified fungi, in decreasing occurrence, were as follows: Penicillium, Fusarium, Epicoccum, Phoma, Acremonium and Aureobasidium. Additionally, seasonal and spatial trends of airborne fungi were assessed, with increases in total culturable fungal concentrations experienced in the summer months. The correspondence between a range of key environmental variables and the phenology of airborne fungal propagules was also examined, with temperature, wind speed and proximal greenspace having the largest influence on fungal propagule density. If the greenspace was comprised of grass, stronger associations with fungal behaviour were observed.     

Impact of elevated CO<Subscript>2</Subscript> in <Emphasis Type="Italic">Casuarina equisetifolia</Emphasis> rooted stem cuttings inoculated with <Emphasis Type="Italic">Frankia</Emphasis>

Impact of different levels of elevated CO ₂ on the activity of Frankia (Nitrogen-fixing actinomycete) in Casuarina equisetifolia rooted stem cuttings has been studied to understand the relationship between C. equisetifolia, Frankia and CO₂. The stem cuttings of C. equietifolia were collected and treated with 2000 ppm of Indole Butyric Acid (IBA) for rooting. Thus vegetative propagated rooted stem cuttings of C. equisetifolia were inoculated with Frankia and placed in the Open top chambers (OTC) with elevated CO₂ facilities. These planting stocks were maintained in the OTC for 12 months under different levels of elevated CO₂ (ambient control, 600 ppm, 900 ppm). After 12 months, the nodule numbers, bio mass, growth, and photosynthesis of C. equisetifolia rooted stem cuttings inoculated with Frankia were improved under 600 ppm of CO₂. The rooted stem cuttings of C. equisetifolia inoculated with Frankia showed a higher number of nodules under 900 ppm of CO₂ and cuttings without Frankia inoculation exhibited poor growth. Tissue Nitrogen (N) content was also higher under 900 ppm of CO₂ than ambient control and 600 ppm levels. The photosynthetic rate was higher (17.8 μ mol CO₂ m^?2 s^?1) in 900 ppm of CO₂ than in 600 ppm (13.2 μ mol CO₂ m^?2 s^?1) and ambient control (8.3 μ mol CO₂ m^?2 s^?1). This study showed that Frankia can improve growth, N fixation and photosynthesis of C. equietifolia rooted stem cuttings under extreme elevated CO₂ level conditions (900 ppm).     

Nitrogen-induced variations in leaf gas exchange of spring triticale under field conditions

Nitrogen (N) is the key factor limiting photosynthetic processes and crop yield. Little is known about the response of leaf gas exchange of spring triticale (Triticosecale Wittm.) to N supply. The effect of N fertilizers on different gas exchange variables, i.e., photosynthetic rate (A), transpiration rate (E), stomatal conductance (g _s), instantaneous water use efficiency (WUE) and maximum quantum yield of photosystem II (PSII) (F _v/F _m), chlorophyll index (SPAD, soil–plant analysis development), and the relationship of these variables with yield were studied in spring triticale grown under field conditions. Six treatments of N—0, 90, 180, 90 + 30, 90 + 30 + 30 kg ha^?1 (applied as ammonium nitrate, AN) and one treatment of N 90 + 30 + 30 kg ha^?1 (applied as urea ammonium nitrate solution, UAN) were compared. The analysis of variance showed that throughout the triticale growing season, N fertilization had significant effects on A, WUE, g _s and SPAD. On average, N fertilizer application increased A values by 14–70%. E and F _v/F _m values were not influenced by N fertilization levels. The effect of growth stage and year on gas exchange variables and F _v/F _m and SPAD was found to be significant. At different growth stages, A values varied and maximum ones were reached at BBCH 31–33 (decimal code system of growth stages) and BBCH 59. With aging, values of A decreased independently of N fertilization level. The gas exchange variables were equally affected by both fertilizer forms. The interplay among grain yield, leaf gas exchange variables, F _v/F _m and SPAD of spring triticale was estimated. The statistical analysis showed that grain yield positively and significantly correlated with A and SPAD values throughout the growing season.     

Microbial lipid production: screening with yeasts grown on Brazilian molasses

Rhodotorula glutinis CCT 2182, Rhodosporidium toruloides CCT 0783, Rhodotorula minuta CCT 1751 and Lipomyces starkeyi DSM 70296 were evaluated for the conversion of sugars from Brazilian molasses into single-cell oil (SCO) feedstock for biodiesel. Pulsed fed-batch fermentations were performed in 1.65 l working volume bioreactors. The maximum specific growth rate (µ_max), lipid productivity (Pr) and cellular lipid content were, respectively, 0.23 h^?1, 0.41 g l^?1 h^?1, and 41 % for Rsp. toruloides; 0.20 h^?1, 0.27 g l^?1 h^?1, and 36 % for Rta. glutinis; 0.115 h^?1, 0.135 g l^?1 h^?1, and 27 % for Rta. minuta; and 0.11 h^?1, 0.13 g l^?1 h^?1, and 32 % for L. starkeyi. Based on their microbial lipid productivity, content, and profile, Rsp. toruloides and Rta. glutinis are promising candidates for biodiesel production from Brazilian molasses. All the oils from the yeasts were similar to the composition of plant oils (rapeseed and soybean) and could be used as raw material for biofuels, as well as in food and nutraceutical products.     

Opposite effects of litter and hemiparasites on a dominant grass under different water regimes and competition levels

Direct and indirect biotic interactions may affect plant growth and development, but the magnitude of these effects may vary depending on environmental conditions. In grassland ecosystems, competition is a strong structuring force. Nonetheless, if hemiparasitic plant species are introduced the competition intensity caused by the dominant species may be affected. However, the outcome of these interactions may change between wet or dry periods. In order to study this, we performed a pot experiment with different densities of the dominant species Schedonorus arundinaceus (1, 2 or 4 individuals) under constantly moist or intermittently dry conditions. The different Schenodorus densities were crossed with presence or absence of hemiparasites (either Rhinanthus minor or R. alectorolophus). Additionally, pots remained with bare ground or received a grass litter layer (400 g m^?2). We expected that indirect litter effects on vegetation (here Schedonorus or Rhinanthus) vary depending on soil moisture. We measured Schedonorus and Rhinanthus aboveground biomass and C stable isotope signature (δ¹³C) as response variables. Overall, Schedonorus attained similar biomass under moist conditions with Rhinanthus as in pots under dry conditions without Rhinanthus. Presence of Rhinanthus also increased δ¹³C in moist pots, indicating hemiparasite-induced water stress. Litter presence increased Schedonorus biomass and reduced δ¹³C, indicating improved water availability. Plants under dry conditions with litter showed similar biomass as under wet conditions without litter. Hemiparasites and litter had opposite effects: hemiparasites reduced Schedonorus biomass while litter presence facilitated grass growth. Contrary to our expectations, litter did not compensate Schedonorus biomass when Rhinanthus was present.     

Analysis of the <Emphasis Type="Italic">xynB5</Emphasis> gene encoding a multifunctional GH3-BglX β-glucosidase-β-xylosidase-α-arabinosidase member in <Emphasis Type="Italic">Caulobacter crescentus</Emphasis>

The Caulobacter crescentus (NA1000) xynB5 gene (CCNA_03149) encodes a predicted β-glucosidase-β-xylosidase enzyme that was amplified by polymerase chain reaction; the product was cloned into the blunt ends of the pJet1.2 plasmid. Analysis of the protein sequence indicated the presence of conserved glycosyl hydrolase 3 (GH3), β-glucosidase-related glycosidase (BglX) and fibronectin type III-like domains. After verifying its identity by DNA sequencing, the xynB5 gene was linked to an amino-terminal His-tag using the pTrcHisA vector. A recombinant protein (95 kDa) was successfully overexpressed from the xynB5 gene in E. coli Top 10 and purified using pre-packed nickel-Sepharose columns. The purified protein (BglX-V-Ara) demonstrated multifunctional activities in the presence of different substrates for β-glucosidase (pNPG: p-nitrophenyl-β-D-glucoside) β-xylosidase (pNPX: p-nitrophenyl-β-D-xyloside) and α-arabinosidase (pNPA: p-nitrophenyl-α-L-arabinosidase). BglX-V-Ara presented an optimal pH of 6 for all substrates and optimal temperature of 50 °C for β-glucosidase and α-l-arabinosidase and 60 °C for β-xylosidase. BglX-V-Ara predominantly presented β-glucosidase activity, with the highest affinity for its substrate and catalytic efficiency (K_m 0.24 ± 0.0005 mM, V_max 0.041 ± 0.002 µmol min^?1 mg^?1 and K_cat/K_m 0.27 mM^?1 s^?1), followed by β-xylosidase (K_m 0.64 ± 0.032 mM, V_max 0.055 ± 0.002 µmol min^?1 mg^?1 and K_cat/K_m 0.14 mM^?1s^?1) and finally α-l-arabinosidase (K_m 1.45 ± 0.05 mM, V_max 0.091 ± 0.0004 µmol min^?1 mg^?1 and K_cat/K_m 0.1 mM^?1 s^?1). To date, this is the first report to demonstrate the characterization of a GH3-BglX family member in C. crescentus that may have applications in biotechnological processes (i.e., the simultaneous saccharification process) because the multifunctional enzyme could play an important role in bacterial hemicellulose degradation.     

Age validation and seasonal growth patterns of a subtropical marsh fish: The Gulf Killifish, <Emphasis Type="Italic">Fundulus grandis</Emphasis>

Fundulus grandis (Baird and Girard), the Gulf Killifish, is an abundant species throughout the marshes of the northern Gulf of Mexico. Its wide distribution and high site fidelity makes it an ideal indicator species for brackish and salt marshes, which experience a variety of anthropogenic disturbances. Despite the ecological, commercial, and scientific importance of F. grandis, age determination methods have not been validated and little is known of its growth pattern. By combining a tag-recapture study with a chemical marker to stain otoliths, we validated an ageing method for F. grandis adults (49–128 mm TL) using whole sagittal otoliths and determined growth rates of recaptured individuals in winter (n = 58) and summer (n = 36) in Louisiana. Mean somatic growth in length was significantly greater during the winter (0.085 mm d^?1) than summer (0.054 mm d^?1). In contrast, mean otolith growth was significantly greater in summer (1.37 μm d^?1) than winter (0.826 μm d^?1). The uncoupling of somatic and otolith growth may be primarily attributed to warm summer temperatures, which led to enhanced otolith growth while simultaneously reducing somatic growth. Fundulus grandis was aged to a maximum of 2.25 years. The parameters of the von Bertalanffy growth model were estimated as: L _∞  = 87.27 mm, k = 2.43 year^?1, and t ₀ = ?0.022. These findings reveal essential age and growth information for F. grandis and provide a benchmark to evaluate responses to environmental disturbances.     

Screening and identification of an <Emphasis Type="Italic">Enterobacter ludwigii</Emphasis> strain expressing an active <Emphasis Type="Italic">β</Emphasis>-xylosidase

Researchers have expressed increasing interest in the xylanolytic enzymes used in hemicellulose hydrolysis that convert wood and agricultural residues to second-generation biofuels. In our study, 32 isolates showed clear hydrolysis zones on agar plates containing xylan after Congo red staining. Among these isolates, strain LY-62 exhibited the highest β-xylosidase activity (1.29?±?0.05 U/mL). According to the phylogenetic analysis of the 16S rDNA, strain LY-62 belongs to the Enterobacter genus. Using a combination of electron microscopy, Gram-staining, and conventional physiological and biochemical examinations, the strain LY-62 was identified as Enterobacter ludwigii. The β-xylosidase gene from Enterobacter ludwigii LY-62 was cloned, and the full-length protein was expressed in Escherichia coli as an N-terminal or C-terminal His-tagged fusions protein. Optimal β-xylosidase activity was achieved at pH 7.0 and 40 °C. The Michaelis constant K_M values for His-Xyl62 and Xyl62-His were 1.55 and 2.8 mmol/L, respectively. The k_cat values for His-Xyl62 and Xyl62-His were 8.51 and 6.94 s^?1, respectively. The catalytic efficiencies of His-Xyl62 and Xyl62-His were 5.49 and 2.48 s^?1?×?mM^?1, respectively. Thus, Xyl62 is a functional β-xylosidase, and our study represents the first report of a β-xylosidase from Enterobacter ludwigii.     

Production of isokaurenoic and kaurenoic acids in double transformed cells of <Emphasis Type="Italic">Saccharomyces cerevisiae</Emphasis>

We report the bifunctional activity of the native ent-kaurene oxidase from Montanoa tomentosa (MtKO) and its N-terminal modified version (LMtKO) for producing both isokaurenoic acid and kaurenoic acid in Saccharomyces cerevisiae. The K_{m app} of MtKO showed more affinity for ent-kaurene (80.5 µM) than for isokaurene (96.4 µM). Interestingly, LMtKO exhibited an increase of the affinity for isokaurene (79.6 µM) but simultaneously showed an enhancement in the V_max for both substrates (32.6–38.9 μmol^?1 mg^?1 h^?1). Biotransformation assays using isokaurene and yeasts containing LMtKO, resulted in 70% more production of isokaurenoic acid, when compared with the yields from yeasts expressing MtKO. Likewise, biotransformation assays using geranylgeraniol and double transformed cells of S. cerevisiae containing an optimized version the ent-kaurene synthase from Phaeosphaeria sp. L487 (optKS) and the LMtKO, produced ~25% more kaurenoic acid than the yeasts containing optKS and MtKO. The isokaurenoic acid synthesized by transgenic yeasts was tested for its anti-acetylcholinesterase and antimicrobial properties. Isokaurenoic acid generated a non-competitive inhibition on acetylcholinesterase, decreasing the V_max from 0.0249 to 0.0104 mM min^?1 but not affecting the K_m (0.714 mM). The same diterpene showed antifungal activity against Fusarium oxysporum, Aspergillus niger and Phytophtora infestans with a minimum inhibitory concentration of 15.3, 18.3 and 19.2 µg mL^?1, respectively.     

Growth kinetics and scale-up of <Emphasis Type="Italic">Agrobacterium tumefaciens</Emphasis>

Production of recombinant proteins in plants through Agrobacterium-mediated transient expression is a promising method of producing human therapeutic proteins, vaccines, and commercial enzymes. This process has been shown to be viable at a large scale and involves growing large quantities of wild-type plants and infiltrating the leaf tissue with a suspension of Agrobacterium tumefaciens bearing the genes of interest. This study examined one of the steps in this process that had not yet been optimized: the scale-up of Agrobacterium production to sufficient volumes for large-scale plant infiltration. Production of Agrobacterium strain C58C1 pTFS40 was scaled up from shake flasks (50–100 mL) to benchtop (5 L) scale with three types of media: Lysogeny broth (LB), yeast extract peptone (YEP) media, and a sucrose-based defined media. The maximum specific growth rate (μ _max) of the strain in the three types of media was 0.46 ± 0.04 h^?1 in LB media, 0.43 ± 0.03 h^?1 in YEP media, and 0.27 ± 0.01 h^?1 in defined media. The maximum biomass concentration reached at this scale was 2.0 ± 0.1, 2.8 ± 0.1, and 2.6 ± 0.1 g dry cell weight (DCW)/L for the three media types. Production was successfully scaled up to a 100-L working volume reactor with YEP media, using k _L a as the scale-up parameter.     

Species and Genotype Effects of Bioenergy Crops on Root Production,Carbon and Nitrogen in Temperate Agricultural Soil

Bioenergy crops have a secondary benefit if they increase soil organic C (SOC) stocks through capture and allocation below-ground. The effects of four genotypes of short-rotation coppice willow (Salix spp., ‘Terra Nova’ and ‘Tora’) and Miscanthus (M.?×?giganteus (‘Giganteus’) and M. sinensis (‘Sinensis’)) on roots, SOC and total nitrogen (TN) were quantified to test whether below-ground biomass controls SOC and TN dynamics. Soil cores were collected under (‘plant’) and between plants (‘gap’) in a field experiment on a temperate agricultural silty clay loam after 4 and 6 years’ management. Root density was greater under Miscanthus for plant (up to 15.5 kg m^?3) compared with gap (up to 2.7 kg m^?3), whereas willow had lower densities (up to 3.7 kg m^?3). Over 2 years, SOC increased below 0.2 m depth from 7.1 to 8.5 kg m^?3 and was greatest under Sinensis at 0–0.1 m depth (24.8 kg m^?3). Miscanthus-derived SOC, based on stable isotope analysis, was greater under plant (11.6 kg m^?3) than gap (3.1 kg m^?3) for Sinensis. Estimated SOC stock change rates over the 2-year period to 1-m depth were 6.4 for Terra Nova, 7.4 for Tora, 3.1 for Giganteus and 8.8 Mg ha^?1 year^?1 for Sinensis. Rates of change of TN were much less. That SOC matched root mass down the profile, particularly under Miscanthus, indicated that perennial root systems are an important contributor. Willow and Miscanthus offer both biomass production and C sequestration when planted in arable soil.     

Bioturbation by the sand dollar <Emphasis Type="Italic">Encope emarginata</Emphasis> (Echinoidea,Mellitidae) changes the composition and size structure of microphytobenthic assemblages

The goal of this study is to examine the suitability of three plants, Typha spp., Phragmites spp. and Iris pseudacorus, in a free-water surface constructed wetland created to treat eutrophic water from Lake Albufera (Valencia, Spain), a wetland of international importance. The growth, coverage and nutrient content of the three plants were studied, and chemical analyses were performed according to standard methods. The maximum standing crops measured for each plant were 1.9, 18.2 and 3.3 kg m^?2, respectively, and their average nutrient concentrations were 2.1, 1.2 and 1.7 g P kg^?1 and 12.1, 11.7 and 10.1 g N kg^?1, respectively. A multiple harvest of Iris pseudacorus revealed that the removal of nutrients could be increased up to 50% for N and 100% for P compared with a single harvest. Biomass decomposition assays showed high values for five-day biochemical oxygen demand (115–207 mg O₂ g^?1, depending on the plant and its age) and a substantial release of phosphorus, up to 100% of that contained in the biomass, highlighting the need to remove the litter fall. This study provides key aspects for vegetation selection and management (planting and harvesting) in a novel application of constructed wetlands to enhance water quality and biodiversity.     

A saponin isolated from <Emphasis Type="Italic">Agapanthus africanus</Emphasis> differentially induces apoplastic peroxidase activity in wheat and displays fungicidal properties

A spirostane with an attached trisaccharide, (25R)-5α-spirostane-2α,3β,5α-triol 3-O-(O-α-l-rhamnopyranosyl-(1 → 2)-O-(β-d-galactopyranosyl-(1 → 3))-β-d-glucopyranoside), was isolated and identified from the aerial parts of Agapanthus africanus by activity-guided fractionation. Fungicidal properties of the crude extract, semi-purified fractions as well as the purified active saponin from A. africanus were screened in vitro against Fusarium oxysporum. At a concentration of 1 mg mL^?1, the crude extract and semi-purified ethyl acetate and dichloromethane fractions showed significant antifungal activity. The purified saponin inhibited the in vitro mycelial growth of F. oxysporum completely (100 %) at a concentration of 125 µg mL^?1. Furthermore, to verify previously observed induced resistance by crude extracts of A. africanus towards leaf rust, intercellular PR-protein activity was determined in wheat seedlings following foliar application of the purified saponin at 100 µg mL^?1. In vitro peroxidase enzyme activity increased significantly (60 %) in wheat seedlings 48 h after treatment with the purified saponin, demonstrating its role as an elicitor to activate a defence reaction in wheat.