Enantioconvergent hydrolysis of racemic styrene oxide at high concentration by a pair of novel epoxide hydrolases into (<Emphasis Type="Italic">R</Emphasis>)-phenyl-1,2-ethanediol

Objectives

To prepare (R)-phenyl-1,2-ethanediol ((R)-PED) with high enantiomeric excess (ee _p) and yield from racemic styrene oxide (rac-SO) at high concentration by bi-enzymatic catalysis.

Results

The bi-enzymatic catalysis was designed for enantioconvergent hydrolysis of rac-SO by a pair of novel epoxide hydrolases (EHs), a Vigna radiata EH3 (VrEH3) and a variant (AuEH2^A250I) of Aspergillus usamii EH2. The simultaneous addition mode of VrEH3 and AuEH2^A250I, exhibiting the highest average turnover frequency (aTOF) of 0.12 g h^?1 g^?1, was selected, by which rac-SO (10 mM) was converted into (R)-PED with 92.6% ee _p and 96.3% yield. Under the optimized reaction conditions: dry weight ratio 14:1 of VrEH3-expressing E. coli/vreh3 to AuEH2^A250I-expressing E. coli/Aueh2 ^A250I and reaction at 20 °C, rac-SO (10 mM) was completely hydrolyzed in 2.3 h, affording (R)-PED with 98% ee _p. At the weight ratio 0.8:1 of rac-SO to two mixed dry cells, (R)-PED with 97.4% ee _p and 98.7% yield was produced from 200 mM (24 mg/ml) rac-SO in 10.5 h.

Conclusions

Enantioconvergent hydrolysis of rac-SO at high concentration catalyzed by both VrEH3 and AuEH2^A250I is an effective method for preparing (R)-PED with high ee _p and yield.

     

Effects of Sewage Sludge Application on Biomass Production and Concentrations of Cd,Pb and Zn in Shoots of <Emphasis Type="Italic">Salix</Emphasis> and <Emphasis Type="Italic">Populus</Emphasis> Clones: Improvement of Phytoremediation Efficiency in Contaminated Soils

Fast-growing clones of Salix and Populus species have been studied for phytoremediation of soils contaminated by risk elements (REs) using short-rotation coppice plantations. Biomass yield, accumulation and removal of RE (Cd, Pb and Zn) by highly productive willow (S1—(Salix schwerinii × Salix viminalis) × S. viminalis, S2—Salix × smithiana) and poplar (P1—Populus maximowiczii × Populus nigra, P2—P. nigra) clones were investigated with and without sewage sludge (SS) application. The precise field experiment was established in April 2008 on moderately Cd-, Pb- and Zn-contaminated soil. Initially, shoots were harvested after four seasons in February 2012 and then after two more seasons in February 2014. The application of SS limited plant growth during the first years of the experiment in the majority of treatments, mainly due to weed competition and higher concentrations of available soil nutrients causing lower yields than those of control (C) treatments. Well-developed roots were able to take advantage of SS applications, and shoot yield was mainly higher in SS treatments in the second harvest, reaching up to 15 t dry matter (DM)?ha^?1. Willows performed better than poplars. Application of SS reduced RE shoot concentrations compared to the C treatment. The removal of RE was significantly higher in the second harvest for all clones and elements (except the P2 clone), and the biomass yield was the major driving force for the amount of RE removed by shoots. Well-developed plantations of fast-growing trees showed better suitability for the phytoextraction of moderately contaminated soils for Cd and partly for Zn but not for Pb, which was less available to plants. From the four tested clones, S2 showed the best removal of Cd (up to 0.94 %) and Zn (up to 0.34 %) of the total soil element content, respectively, and this clone is a good candidate for phytoextraction. SS can be a suitable source of nutrients for Salix clones without any threat to the food chain in terms of biomass contamination, but its application to the soil can result in an increased incidence of some weeds during the first years of plantation.     

Co-inoculation with <Emphasis Type="Italic">Enterobacter</Emphasis> and Rhizobacteria on Yield and Nutrient Uptake by Wheat (<Emphasis Type="Italic">Triticum aestivum</Emphasis> L.) in the Alluvial Soil Under Indo-Gangetic Plain of India

The aim of this work was to evaluate the effects of co-inoculation with phosphate-solubilizing and nitrogen-fixing rhizobacteria on growth promotion, yield, and nutrient uptake by wheat. Out of twenty-five bacteria isolated from the rhizosphere soils of cereal, vegetable, and agro-forestry plants in eastern Uttar Pradesh, three superior most plant growth-promoting (PGP) isolates were characterized as Serratia marcescens, Microbacterium arborescens, and Enterobacter sp. based on their biochemical and 16S rDNA gene sequencing data and selected them for evaluating their PGP effects on growth and yield of wheat. Among them, Enterobacter sp. and M. arborescens fixed significantly higher amounts (9.32?±?0.57 and 8.89?±?0.58 mg Ng^?1 carbon oxidized, respectively) of atmospheric nitrogen and produced higher amounts (27.06?±?1.70 and 26.82?±?1.63 TP 100 µg mL^?1, respectively) of IAA in vitro compared to S. marcescens (8.32?±?0.39 mg Ng^?1 carbon oxidized and 21.29?±?0.99 TP 100 µg mL^?1). Although both M. arborescens and S. marcescens solubilized remarkable amounts of phosphate from tricalcium phosphate likely through production of organic acids, however, Enterobacter sp. was inactive. The effects of these three rhizobacteria were evaluated on wheat in alluvial soils of the Indo-Gangetic Plain by inoculation of plants with bacterial isolates either alone or in combinations in both pot and field conditions for two successive years. Rhizobacterial inoculation either alone or in consortium of varying combinations significantly (P?≤?0.05) increased growth and yield of wheat compared to mock inoculated controls. A consortium of two or three rhizobacterial isolates also significantly increased plant height, straw yield, grain yield, and test weight of wheat in both pot and field trials compared to single application of any of these isolates. Among the rhizobacterial treatment, co-inoculation of three rhizobacteria (Enterobacter, M. arborescens and S. marcescens) performed best in promotion of growth, yield, and nutrient (N, P, Cu, Zn, Mn, and Fe) uptake by wheat. Taken together, our results suggest that co-inoculation of Enterobacter with S. marcescens and M. arborescens could be used for preparation of an effective formulation of PGP consortium for eco-friendly and sustainable production of wheat.     

<Emphasis Type="Italic">Rhizophagus irregularis</Emphasis> MUCL 41833 can colonize and improve P uptake of <Emphasis Type="Italic">Plantago lanceolata</Emphasis> after exposure to ionizing gamma radiation in root organ culture

Long-lived radionuclides such as ⁹⁰Sr and ¹³⁷Cs can be naturally or accidentally deposited in the upper soil layers where they emit β/γ radiation. Previous studies have shown that arbuscular mycorrhizal fungi (AMF) can accumulate and transfer radionuclides from soil to plant, but there have been no studies on the direct impact of ionizing radiation on AMF. In this study, root organ cultures of the AMF Rhizophagus irregularis MUCL 41833 were exposed to 15.37, 30.35, and 113.03 Gy gamma radiation from a ¹³⁷Cs source. Exposed spores were subsequently inoculated to Plantago lanceolata seedlings in pots, and root colonization and P uptake evaluated. P. lanceolata seedlings inoculated with non-irradiated AMF spores or with spores irradiated with up to 30.35 Gy gamma radiation had similar levels of root colonization. Spores irradiated with 113.03 Gy gamma radiation failed to colonize P. lanceolata roots. P content of plants inoculated with non-irradiated spores or of plants inoculated with spores irradiated with up to 30.35 Gy gamma radiation was higher than in non-mycorrhizal plants or plants inoculated with spores irradiated with 113.03 Gy gamma radiation. These results demonstrate that spores of R. irregularis MUCL 41833 are tolerant to chronic ionizing radiation at high doses.     

<Emphasis Type="Italic">Pseudomonas</Emphasis> spp. increases root biomass and tropane alkaloid yields in transgenic hairy roots of <Emphasis Type="Italic">Datura</Emphasis> spp.

Transgenic hairy roots of Datura spp., established using strain A4 of Agrobacterium rhizogenes, are genetically stable and produce high levels of tropane alkaloids. To increase biomass and tropane alkaloid content of this plant tissue, four Pseudomonas strains, Pseudomonas fluorescens P64, P66, C7R12, and Pseudomonas putida PP01 were assayed as biotic elicitors on transgenic hairy roots of Datura stramonium, Datura tatula, and Datura innoxia. Alkaloids were extracted from dried biomass, and hyoscyamine and scopolamine were quantified using liquid chromatography-tandem mass spectrometry analysis. D. stramonium and D. innoxia biomass production was stimulated by all Pseudomonas spp. strains after a 5-d treatment. All strains of P. fluorescens increased hyoscyamine yields compared to untreated cultures after both 5 and 10 d of treatment. Hyoscyamine yields were highest in D. tatula cultures exposed to a 5-d treatment with C7R12 (16.633 + 0.456 mg g^?1 dry weight, a 431% increase) although the highest yield increases compared to the control were observed in D. stramonium cultures exposed to strains P64 (511% increase) and C7R12 (583% increase) for 10 d. D. innoxia showed the highest scopolamine yields after elicitation with P. fluorescens strains P64 for 5 d (0.653 + 0.021 mg g^?1 dry weight, a 265% increase) and P66 for 5 and 10 d (5 d, 0.754 + 0.0.031 mg g^?1 dry weight, a 321% increase; 10 d 0.634 + 0.046 mg g^?1 dry weight, a 277% increase). These results show that the Pseudomonas strains studied here can positively and significantly affect biomass and the yields of hyoscyamine and scopolamine from transgenic roots of the three Datura species.     

Sap flow of the southern conifer, <Emphasis Type="Italic">Agathis australis</Emphasis> during wet and dry summers

Key message

Analysis of sap flux density during drought suggests that the large sapwood and rooting volumes of larger trees provide a buffer against drying soil.

Abstract

The southern conifer Agathis australis is amongst the largest and longest-lived trees in the world. We measured sap flux densities (F _d) in kauri trees with a DBH range of 20–176 cm to explore differences in responses of trees of different sizes to seasonal conditions and summer drought. F _d was consistently higher in larger trees than smaller trees. Peak F _d was 20 and 8 g m^?2 s^?1 for trees of diameters of 176 and 20 cm, respectively, during the wet summer. Multiple regression analysis revealed photosynthetically active radiation (PAR) and vapour pressure deficit (D) were the main drivers of F _d. During drought, larger trees were more responsive to D whilst smaller trees were more responsive to soil drying. Our largest tree had a sapwood area of 3,600 cm². Preliminary analysis suggests stem water storage provides a buffer against drying soil in larger trees. Furthermore, F _d of smaller trees had higher R ² values for soil moisture at 30 and 60 cm depth than soil moisture at 10 cm depth (R ² = 0.68–0.97 and 0.55–0.67, respectively) suggesting that deeper soil moisture is more important for these trees. Larger trees did not show a relationship between F _d and soil moisture, suggesting they were accessing soil water deeper than 60 cm. These results suggest that larger trees may be better prepared for increasing frequency and intensity of summer droughts due to deeper roots and/or larger stem water storage capacity.

     

Cloning,deletion, and overexpression of a glucose oxidase gene in <Emphasis Type="Italic">Aureobasidium</Emphasis> sp. P6 for Ca<Superscript>2+</Superscript>-gluconic acid overproduction

This study aimed to overexpress a glucose oxidase gene (GOD1) in Aureobasidium sp. P6 to achieve Ca²⁺-gluconic acid (GA) overproduction. The GOD1 gene was cloned, deleted, and overexpressed. A protein deduced from the GOD1 gene of Aureobasidium sp. P6 strain had 1824 bp that encoded a protein with 606 amino acids, with a conserved NADB-ROSSMAN domain and a GMC-oxred domain. Deleting the GOD1 gene made the disruptant GOK1 completely lose the ability to produce GA and GOD1 activity, whereas overexpressing the GOD1 gene rendered the transformant GOEX8 to produce considerably more Ca²⁺-GA (160.5?±?5.6 g/L) and higher GOD1 activity (1438.6?±?73.2 U/mg of protein) than its parent P6 strain (118.7?±?4.3 g/L of Ca²⁺-GA and 1100.0?±?23.6 U/mg of GOD1 protein). During a 10-L fermentation, the transformant GOEX8 grown in the medium containing 160.0 g/L of glucose produced 186.8?±?6.0 g/L of Ca²⁺-GA, the yield was 1.2 g/g of glucose, and the volumetric productivity was 1.7 g/L/h. Most of the produced GOD1 were located in the yeast cell wall. The purified product was identified to be a GA. The transformant GOEX8 overexpressing the GOD1 gene could produce considerably more Ca²⁺-GA (186.8?±?6.0 g/L) than its wild-type strain P6.     

Effects of macrophyte leachate on <Emphasis Type="Italic">Anabaena</Emphasis> sp. and <Emphasis Type="Italic">Chlamydomonas moewusii</Emphasis> growth in freshwater tropical ecosystems

This study reports on the effects of dissolved organic matter (DOM) derived from the aquatic macrophyte Pistia stratiotes (collected from a tropical reservoir) on the mixotrophic growth of two phytoplankton species (Chlamydomonas moewusii and Anabaena sp.). The DOM from P. stratiotes had a mainly aliphatic structure, low molecular weight, low cellulose and lignin content and high carbon content. The addition of DOM (5% v/v) significantly decreased the growth rate of Anabaena sp. but increased the chlorophyll a concentration of C. moewusii. Higher light intensity (100 versus 30 µmol m^?2 s^?1) was important for Anabaena sp., increasing its growth rate and chlorophyll content. The use of DOM from P. stratiotes to mitigate cyanobacterial blooms should be further explored in future studies.     

Toxin Gene Contents and Activity of <Emphasis Type="Italic">Bacillus thuringiensis</Emphasis> Strains Against Two Sugarcane Borer Species, <Emphasis Type="Italic">Diatraea saccharalis</Emphasis> (F.) and <Emphasis Type="Italic">D. flavipennella</Emphasis> (Box)

Bacillus thuringiensis (Berliner) bears essential characteristics in the control of insect pests, such as its unique mode of action, which confers specificity and selectivity. This study assessed cry gene contents from Bt strains and their entomotoxicity against Diatraea saccharalis (F.) and Diatraea flavipennella (Box) (Lepidoptera: Crambidae). Bioassays with Bt strains were performed against neonates to evaluate their lethal and sublethal activities and were further analyzed by PCR, using primers to identify toxin genes. For D. saccharalis and D. flavipennella, 16 and 18 strains showed over 30% larval mortality in the 7th day, respectively. The LC₅₀ values of strains for D. saccharalis varied from 0.08 × 10⁵ (LIIT-0105) to 4104 × 10⁵ (LIIT-2707) spores + crystals mL^?1. For D. flavipennella, the LC₅₀ values of strains varied from 0.40 × 10⁵ (LIIT-2707) to 542 × 10⁵ (LIIT-2109) spores + crystals mL^?1. For the LIIT-0105 strain, which was the most toxic to D. saccharalis, the genes cry1Aa, cry1Ab, cry1Ac, cry1B, cry1C, cry1D, cry1F, cry1I, cry2Aa, cry2Ab, cry8, and cry9C were detected, whereas for the strain LIIT-2707, which was the most toxic to D. flavipennella, detected genes were cry1Aa, cry1Ab, cry1Ac, cry1B, cry1D, cry1F, cry1I, cry2Aa, cry2Ab, and cry9. The toxicity data and toxin gene content in these strains of Bt suggest a great variability of activity with potential to be used in the development of novel biopesticides or as source of resistance genes that can be expressed in plants to control pests.     

Effects of light intensity on leaf photosynthetic characteristics,chloroplast structure,and alkaloid content of <Emphasis Type="Italic">Mahonia bodinieri</Emphasis> (Gagnep.) Laferr.

The variation of light intensity has obvious effects on leaf external morphology, internal anatomy, and physiological characteristics; it even induces changes in secondary metabolite production. The effects of different irradiance levels on biomass, gas exchange parameters, and photosynthetic pigment contents in Mahonia bodinieri (Gagnep.) Laferr. were analyzed here. Combined analyses of physiology, cytology, and HPLC were used to study the differences in leaf morphology, structure, physiological characters, and alkaloid content in response to different irradiances. The results indicated that the highest foliar biomass was observed under I ₅₀ (50 % of full sunlight) followed by I ₃₀ (30 % of full sunlight), the highest net photosynthetic rate, stomatal conductance, transpiration rate values were observed under I ₃₀ followed by I ₅₀, and lower values occurred in I ₁₀ (10 % of full sunlight) and I ₁₀₀ (full sunlight). With increased light intensity, total leaf area and the contents of chlorophyll a (Chl a), chlorophyll b (Chl b), and chlorophyll (Chl a+b) per unit leaf area were clearly reduced, whereas leaf mass per area, carotenoid content, leaf thickness, thickness of palisade and spongy parenchyma, and stomatal density were all significantly increased. Electron microscopic observation revealed that the number of grana, stroma lamellae and the number of starch grains in chloroplasts were decreased, the number of plastoglobuli was increased when irradiance levels increased. The estimated total yield of alkaloids in a single plant was higher under I ₃₀ and I ₅₀ than under I ₁₀ or I ₁₀₀ as a result of the higher biomass of the plants. Therefore, I ₃₀ and I ₅₀ were not only beneficial to increase biomass, but also suitable for the synthesis and accumulation of the major secondary metabolites (alkaloids). Our findings provide valuable data for the determination and regulation of irradiance levels during artificial cultivation of M. bodinieri.     

M3 cholinoreceptors alter electrical activity of rat left atrium via suppression of L-type Ca<Superscript>2+</Superscript> current without affecting K<Superscript>+</Superscript> conductance

Electrophysiological effects produced by selective activation of M3 cholinoreceptors were studied in isolated left atrium preparations from rat using the standard sharp glass microelectrode technique. The stimulation of M3 receptors was obtained by application of muscarinic agonist pilocarpine (10^?5 M) in the presence of selective M2 antagonist methoctramine (10^?7 M). Stimulation of M3 receptors induced marked reduction of action potential duration by 14.4 ± 2.4% and 16.1 ± 2.5% of control duration measured at 50 and 90% of repolarization, respectively. This effect was completely abolished by selective M3 blocker 4-DAMP (10^?8 M). In isolated myocytes obtained from the rat left atrium, similar pharmacological stimulation of M3 receptors led to suppression of peak L-type calcium current by 13.9 ± 2.6% of control amplitude (measured at +10 mV), but failed to affect K⁺ currents I _to, I _Kur, and I _Kir. In the absence of M2 blocker methoctramine, pilocarpine (10^?5 M) produced stronger attenuation of I _CaL and induced an increase in I _Kir. This additive inward rectifier current could be abolished by highly selective blocker of K_ir3.1/3.4 channels tertiapin-Q (10^?6 M) and therefore was identified as I _KACh. Thus, in the rat atrial myocardium activation of M3 receptors leads to shortening of action potentials via suppression of I _CaL, but does not enhance the major potassium currents involved in repolarization. Joint stimulation of M2 and M3 receptors produces stronger action potential shortening due to M2-mediated activation of I _KACh.     

Effects of both substrate and nitrogen and phosphorus fertilizer on <Emphasis Type="Italic">Sphagnum palustre</Emphasis> growth in subtropical high-mountain regions and implications for peatland recovery

Human activities have recently caused severe destruction of Sphagnum wetlands in subtropical high-mountain regions, calling for urgent efforts to restore Sphagnum wetlands. Through a greenhouse experiment in western Hubei, China, we studied the effects of different substrate types (peat and mountain soil) and different levels of nitrogen (N) (0, 2, 4, 6, 10 g m^?2 year^?1) and phosphorus (P) (0, 0.2, 0.5, 1, 2 g m^?2 year^?1) on the growth of Sphagnum palustre, which was evaluated by four growth indicators: length growth, number of capitula, coverage change and biomass. We aimed to determine the optimal nutrient conditions for S. palustre growth, which would contribute to the rapid colonization and restoration of Sphagnum wetlands. The results showed that the different substrates significantly influenced S. palustre growth. Compared with those of peat, the acidic properties of the local yellow brown soil in the subtropical high-mountain regions were more favorable for S. palustre growth. As N addition increased, the four growth indicators responded inconsistently to the different substrates. While the number of capitula markedly increased, the other three indicators significantly decreased in the mountain soil or exhibited no definitive changes in the peat. The addition of P markedly promoted S. palustre growth in both substrates. However, a threshold for P fertilization existed; the highest productivity occurred at P additions of 0.2 and 0.5 g m^?2 year^?1 in the peat and mountain soil, respectively. The N and P contents in the capitula increased in parallel as the N and P fertilization rates increased, suggesting that these nutrients were absorbed proportionately and were used during the growth of S. palustre.     

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.     

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.     

Specific light uptake rates can enhance astaxanthin productivity in <Emphasis Type="Italic">Haematococcus lacustris</Emphasis>

Lumostatic operation was applied for efficient astaxanthin production in autotrophic Haematococcus lacustris cultures using 0.4-L bubble column photobioreactors. The lumostatic operation in this study was performed with three different specific light uptake rates (q _e) based on cell concentration, cell projection area, and fresh weight as one-, two- and three-dimensional characteristics values, respectively. The q _e value from the cell concentration (q _e1D) obtained was 13.5 × 10^?8 μE cell^?1 s^?1, and the maximum astaxanthin concentration was increased to 150 % compared to that of a control with constant light intensity. The other optimum q _e values by cell projection area (q _e2D) and fresh weight (q _e3D) were determined to be 195 μE m^?2 s^?1 and 10.5 μE g^?1 s^?1 for astaxanthin production, respectively. The maximum astaxanthin production from the lumostatic cultures using the parameters controlled by cell projection area (2D) and fresh weight (3D) also increased by 36 and 22 % over that of the controls, respectively. When comparing the optimal q _e values among the three different types, the lumostatic cultures using q _e based on fresh weight showed the highest astaxanthin productivity (22.8 mg L^?1 day^?1), which was a higher level than previously reported. The lumostatic operations reported here demonstrated that more efficient and effective astaxanthin production was obtained by H. lacustris than providing a constant light intensity, regardless of which parameter is used to calculate the specific light uptake rate.     

Promoter polymorphism MMP-1 (-1607 2G/1G) and MMP-3 (-1612 5A/6A) in development of HAND and modulation of pathogenesis of HAND

The pathogenesis of HIV-associated neurocognitive disorder (HAND) is modulated by host genetic susceptibility factors such as Matrix metalloproteinases (MMPs). Promoter polymorphism of MMP-1 and MMP-3 may modify the expression of the gene. Hence, we evaluated the association of MMP-1-16072G/1G and MMP-3-1612 5A/6A polymorphisms with development of HAND and the modulation of pathogenesis of HAND. We enrolled a total of 180 individuals, 50 HIV-infected individuals with HAND, 130 without HAND, and 150 healthy controls. Polymorphism of MMP-1 and MMP-3 were genotyped by PCR-RFLP. MMP-1-1607 2G1G, -16071G/2G-1G/1G genotypes and -1607 1G allele were associated with the development of HAND (OR = 1.64, P = 0.05; OR = 1.45, P = 0.04; OR = 1.69, P = 0.05). MMP-1-16071G1G, MMP-3-16125A5A genotypes increased the risk for the development of HAND (OR = 1.78, P = 0.25; OR = 2.39, P = 0.13). MMP-3-1612 5A5A, -1612 6A/5A-5A/5A genotypes and -1612 5A allele were associated with the reduced risk of HAND (OR = 0.40, P = 0.05; OR = 0.53, P = 0.04; OR = 0.40, P = 0.01). Haplotype 5A1G increased the risk of development of HAND (OR = 1.93, P = 0.05). As observed in advanced HIV disease stage, MMP-1-1607 1G1G genotype enhance the risk for advancement of HIV disease (OR = 1.69, P = 0.89). MMP-3-1612 6A5A genotype showed higher risk for development of HAND in alcohol users (0R = 1.65, P = 0.44). MMP-1 genotype may have an influence on development of HAND whereas MMP3-1612 5A5A genotype may reduce risk for pathogenesis of HAND.     

Effect of soil application of humic acid on nutrients uptake,essential oil and chemical compositions of garden thyme (<Emphasis Type="Italic">Thymus vulgaris</Emphasis> L.) under greenhouse conditions

Humic acid is natural biological organic, which has a high effect on plant growth and quality. However, the mechanisms of the promoting effect of humic acid on the volatile composition were rarely reported. In this study, the effects of soil application of humic acid on the chemical composition and nutrients uptake of Thymus vulgaris were investigated. Treatments comprised 0, 50, 75 and 100 g m^?2. Essential oil was extracted by hydrodistillation and analyzed using GC–MS and GC–FID. Essential oil content was enhanced by increase of the humic acid level and its content ranged from 0.8% (control) to 2.0% (75 g m^?2). Thirty-two volatile compounds were identified and these compounds were considerably affected by humic acid. The highest percentage of thymol (74.15%), carvacrol (6.20%), p-cymene (4.24%), borneol (3.42%), trans-caryophyllene (1.70%) and cis-sabinene hydrate (1.35%) as major compounds were observed in T. vulgaris under 100 g m^?2 humic acid. There was a linear relationship (R² = 97%) between humic acid levels and thymol as a major compound. The oils were dominated by oxygenated monoterpenes followed by monoterpene hydrocarbons and sesquiterpene hydrocarbons. Based on the path coefficient analysis, the highest direct effects on essential oil content were observed in monoterpene esters (3.465) and oxygenated sesquiterpenes (3.146). The humic acid application also enhanced the uptake of N, P, K, Mg and Fe in garden thyme. The highest N (2.42%), P (0.75%), K (2.63%), Mg (0.23%) and Fe (1436.58 ppm) were observed in medium supplemented with 100 g m^?2 humic acid. In all, the utilization of humic acid could positively change nutrients uptake, essential oil content and its major constituents in T. vulgaris.     

Organogenesis from root explants of commercial populations of <Emphasis Type="Italic">Passiflora edulis</Emphasis> Sims and a wild passionfruit species, <Emphasis Type="Italic">P. cincinnata</Emphasis> Masters

Root explants of a wild passionfruit species (Passiflora cincinnata) and three P. edulis commercial populations (‘FB 100’, ‘FB 200’, and ‘FB 300’) were incubated on Murashige and Skoog (MS) medium supplemented with 4.44 μM 6-benzyladenine (BA) to induce shoot organogenesis. Shoots elongated in liquid medium with 2.89 μM gibberellic acid (GA₃) under agitation were rooted in coconut fiber and acclimatized followed by transfer to a greenhouse into pots containing mixture of coconut fiber and Plantmax^® (1:1). Explant samples were collected during organogenesis and submitted to light and scanning electron microscopy (SEM). Root explants of P. cincinnata responded earlier than those of P. edulis. However, on the third assessment, at 90 days, the genotype ‘FB 200’ showed shoot number significantly higher than ‘FB 100’ and ‘FB 300’, not differing from P. cincinnata. Organogenesis in P. cincinnata and P. edulis occurred via direct pathway, which was confirmed by anatomical studies and SEM. Flow cytometric analysis revealed no variation in DNA content of regenerated plantlets among all genotypes. Nuclear DNA (2C) values (pg) in regenerants of P. cincinnata (2.99 pg) and P. edulis (3.26–3.28 pg) were consistent with DNA amounts of seed-derived control plants.     

First-Rotation Performance of Five Short-Rotation Willow Cultivars on Different Soil Types and Along a Large Climate Gradient

The establishment of short-rotation willow coppice plantations for bioenergy production is proposed as a land management practice to reduce the use of fossil fuels. However, there is little information on the performance of different willow (Salix spp.) cultivars on various types of soils and over climate gradients. This study aimed to determine which set of soil, climatic conditions, and cultivars are conducive to greater growth in eastern Canada. The performance of five willow cultivars was followed over three growing seasons on eight sites representing a large hydroclimatic gradient. Both geographic location and cultivar had a significant effect on annual yields. Annual yields were on average greater in the southern part of the climatic gradient. Across all cultivars, annual yields were positively correlated to average annual temperature (r = 0.23), total annual precipitation (r = 0.77), average growing season temperature (r = 0.21), average growing season precipitation (r = 0.47), and degree days (r = 0.18), as well as soil pH (r = 0.37) and soil extractible P (r = 0.38), and negatively correlated to soil clay content (r = 0.33). Cultivars of the SX group (i.e., Salix miyabeana SX61, SX64, and SX67) showed greater yield than did cultivars of the SV group (i.e., Salix × dasyclados SV1 and Salix viminalis SV5027). These results indicate that at the landscape level, climate variables, especially climate-related variables, largely explain the yield of the selected willow cultivars. Nonetheless, soil pH, extractable P, and clay content likely play an important role in plantation yield.