Depressive symptoms post hip fracture in older adults are associated with phenotypic and functional alterations in T cells

Background

Ageing is accompanied by reduced immunity, termed immunesenescence. The immune system does not act in isolation and is sensitive to both psychological and physical stress. Hip fracture is a common physical stressor in older adults with a high incidence of new onset depression, which relates to poorer prognosis. We therefore set out to examine the possible synergistic effects of physical stress (hip fracture) and psychological stress (depressive symptoms) on the aged immune system.

Results

T cell phenotype and function was assessed in 101 hip fracture patients (81 female) 6 weeks after hip fracture and 43 healthy age-matched controls (26 female). 38 fracture patients had depressive symptoms at 6 weeks. T cell frequency (p?=?.01) and numbers (p?=?.003) were both lower in depressed hip fracture patients compared to healthy controls. The frequency of senescent CD28^-ve (p?=?.001), CD57^+ve (p?=?.001), KLRG1^+ve (p?=?.03) CD8 T cells, as well as senescent CD28^-ve CD4^+ve (p?=?.01) and CD57^+ve CD4^+ve (p?=?.003) T cells were higher in depressed hip fracture patients compared with healthy controls and the frequency of CD28^-ve CD8 T cells was also higher when compared to patients with hip fracture alone (p?=?.01). Additionally, activated CD69^+ve (p?=?.005) and HLADR^+ve (p?<?.001) CD8 T cells, were also higher in depressed hip fracture patients compared to healthy controls. On examining cytokine production by activated T cells, a significant increase in TNFα (p?=?.03) and IL6 (p?=?.04) production was observed in CD4 T cells from hip fracture patients with depressive symptoms compared to healthy controls.

Conclusions

As none of the patients in the study had a prior history of depression, our data suggest that the development of depressive symptoms in hip fracture patients is associated with altered T cell phenotype and increased pro-inflammatory function which is not seen in patients who do not develop depression after hip fracture. Treating depressive symptoms promptly in hip fracture patients may therefore improve immunity and outcomes in these patients.

     

CD77-dependent retrograde transport of CD19 to the nuclear membrane: Functional relationship between CD77 and CD19 during germinal center B-cell apoptosis

A region of the N-terminal extracellular domain of the B-cell restricted cell differentiation antigen, CD19, has high amino acid sequence similarity to the receptor binding subunit B of verotoxin 1 (VT), an Escherichia coli elaborated cytotoxin, which specifically binds to the cell surface glycolipid, globotriaosylceramide, also known as the germinal center (GC) B-cell differentiation antigen, CD77. We have previously provided evidence of the association of CD19 and CD77 on the cell surface and in CD19-mediated homotypic adhesion of the Daudi Burkitt Lymphoma cell line, one normal counterpart of which is a subset of GC B cells. Evidence for the role of CD77 in CD19-induced apoptosis is now presented. Initial cell surface distribution, antibody-induced redistribution, internalization, and intracellular routing of CD19 were studied by confocal microscopy, IF, and postembedding IEM in CD77^+ve and CD77^−ve cells to investigate the possible role of CD77 in CD19 internalization and signaling. Daudi Burkitt's lymphoma cells were used as CD77^+ve cells and as CD77^−ve cells, Daudi mutant VT500 cells, and Daudi cells treated with PPMP, an inhibitor of CD77 synthesis, were used. Antibody ligated CD19 surface redistribution, internalization, and subcellular distribution of internalized CD19 was found to be different in CD77^+ve and CD77^−ve cells. A delay in internalization of antibody-CD19 complex was observed in CD77^−ve cells. Internalized CD19 was targeted to the nuclear envelope in CD77^+ve cells in a manner similar to that reported for VT, but not in CD77^−ve cells. Internalization of CD77 by ligation with verotoxin prevented the internalization of ligated CD19. Induction of apoptosis following crosslinking of cell surface CD19 was greater in CD77^+ve cells than in CD77^−ve cells. The nuclear targeting of internalized CD19 and induction of apoptosis following CD19 crosslinking only in CD77^+ve cells indicates a role for CD77-dependent CD19 retrograde transport from the B cell surface via the ER to the nuclear envelope in CD19-mediated signal transduction for apoptosis. J. Cell. Physiol. 176:281–292, 1998. © 1998 Wiley-Liss, Inc.     

Sequencing and phylogenetic analyses of talaromyces amestolkiae from amazon: A producer of natural colorants

The population interest in health products is increasing day-by-day. Thus, the demand for natural products to be added in food and pharmaceutical commodity is also rising. Among these additives, colorants, which provides color to products, can be produced by microorganism through bioprocess. Looking for new source of natural colorants, fungi have been employed to this purpose producing novel and safer natural colorants. So, the main goal of this study was to describe a Talaromyces species able to produce natural colorants and investigate nutritional parameters of colorants production using statistical tool. The taxonomy classified the microorganism as Talaromyces amestolkiae. The statistical design evaluated pH and glucose, meat extract and meat peptone concentration as independent variables, and red colorants production as main response. Under the best condition (g/L: glucose 30, meat extract 1, meat peptone 10, and initial pH of 7.0) an increase of 229% in the red colorant production was achieved as compared with the initial media used. The dried fermented broth containing red colorants showed low cytotoxicity against fibroblasts cells (IC₅₀ > 187.5 g/L) and effective antimicrobial activity against S. aureus (MIC of 2.5 g/L). Thus, T. amestolkiae colorants can be attractive to food and pharmaceutical applications as it does not produce toxic compounds and can promote protection against microorganism contaminants. © 2018 American Institute of Chemical Engineers Biotechnol. Prog., 35: e2684, 2019     

Seizure induces activation of multiple subtypes of neural progenitors and growth factors in hippocampus with neuronal maturation confined to dentate gyrus

Adult hippocampal neurogenesis is altered in response to different physiological and pathological stimuli. GFAP^+ve/nestin^+ve radial glial like Type-1 progenitors are considered to be the resident stem cell population in adult hippocampus. During neurogenesis these Type-1 progenitors matures to GFAP^−ve/nestin^+ve Type-2 progenitors and then to Type-3 neuroblasts and finally differentiates into granule cell neurons. In our study, using pilocarpine-induced seizure model, we showed that seizure initiated activation of multiple progenitors in the entire hippocampal area such as DG, CA1 and CA3. Seizure induction resulted in activation of two subtypes of Type-1 progenitors, Type-1a (GFAP^+ve/nestin^+ve/BrdU^+ve) and Type-1b (GFAP^+ve/nestin^+ve/BrdU^−ve). We showed that majority of Type-1b progenitors were undergoing only a transition from a state of dormancy to activated form immediately after seizures rather than proliferating, whereas Type-1a showed maximum proliferation by 3 days post-seizure induction. Type-2 (GFAP^−ve/nestin^+ve/BrdU^+ve) progenitors were few compared to Type-1. Type-3 (DCX^+ve) progenitors showed increased expression of immature neurons only in DG region by 3 days after seizure induction indicating maturation of progenitors happens only in microenvironment of DG even though progenitors are activated in CA1 and CA3 regions of hippocampus. Also parallel increase in growth factors expression after seizure induction suggests that microenvironmental niche has a profound effect on stimulation of adult neural progenitors.     

Ozone pollution influences soil carbon and nitrogen sequestration and aggregate composition in paddy soils

Background and aims

Much attention has focused on the effects of tropospheric ozone (O₃) on terrestrial ecosystems and plant growth. Since O₃ pollution is currently an issue in China and many parts of the world, understanding the effects of elevated O₃ on soil carbon (C) and nitrogen (N) sequestration is essential for efforts to predict C and N cycles in terrestrial ecosystems under predicted increases in O₃. Thus the main objective of this study was to determine whether an increases in atmospheric O₃ concentration influenced soil organic C (SOC) and N sequestration.

Methods

A free-air O₃ enrichment (O₃-FACE) experiment was started in 2007 and used continuous O₃ exposure from March to November each year during crop growth stage in a rice (Oryza sativa L.)—wheat (Triticum aestivum L.) rotation field in the Jiangsu Province, China. We investigated differences in SOC and N and soil aggregate composition in both elevated and ambient O₃ conditions.

Results

Elevated atmospheric O₃ (18–80 nmol mol^?1 or 50 % above the ambient) decreased the SOC and N concentration in the 0–20 cm soil layer after 5 years. Elevated O₃ significantly decreased the SOC concentration by 17 % and 5.6 % in the 0–3 cm and the 10–20 cm layers, respectively. Elevated O₃ significantly decreased the N concentration by 8.2–27.8 % in three layers at the 20 cm depth. In addition, elevated O₃ influenced the formation and transformation of soil aggregates and the distribution of SOC and N in the aggregates across soil layer classes. Elevated O₃ significantly decreased the macro-sized aggregate fraction (16.8 %) and associated C and N (0.5 g kg^?1 and 0.32 g kg^?1, respectively), and significantly increased the silt+ clay-sized aggregate fraction (61 %) and associated C (1.7 g kg^?1) in the 0–3 cm layer. Elevated O₃ significantly decreased the macro-sized aggregate fraction (9.6 %) and associated C and N (1.4 g kg^?1 and 0.35 g kg^?1, respectively), and significantly increased the silt+ clay-sized aggregate fraction (41.8 %) and decreased the corresponding associated N (0.14 g kg^?1) in the 3–10 cm layer. Elevated O₃ did not significantly effect the formation and transformation of aggregates in the 10–20 cm layer, yet it did significantly increase the C concentration in the macro-sized fraction (1 g kg^?1) and decrease the N concentration in the macro- and micro-sized fractions (0.24 g kg^?1 and 0.16 g kg^?1, respectively).

Conclusion

Long-term exposure to elevated atmospheric O₃ negatively affected the physical structure of the soil and impaired soil C and N sequestration.     

Thermostable phycocyanin from the red microalga <Emphasis Type="Italic">Cyanidioschyzon merolae</Emphasis>, a new natural blue food colorant

The demand for natural food colorants is growing as consumers question the use of artificial colorants more and more. The phycobiliprotein C-phycocyanin of Arthospira platensis is used as a natural blue colorant in certain food products. The thermoacidophilic red microalga Cyanidioschyzon merolae might provide an alternative source of phycocyanin. Cyanidioschyzon merolae belongs to the order Cyanidiophyceae of the phylum Rhodophyta. Its natural habitat are sulfuric hot springs and geysers found near volcanic areas in, e.g., Yellowstone National Park in the USA and in Java, Indonesia. It grows optimally at a pH between 0.5 and 3.0 and at temperatures up to 56 °C. The low pH at which C. merolae grows minimizes the risk of microbial contamination and could limit production loss. As C. merolae lacks a cell wall, phycocyanin with a high purity number of 9.9 could be extracted by an osmotic shock using a simple ultrapure water extraction followed by centrifugation. The denaturation midpoint at pH 5 was 83 °C, being considerably higher than the A. platensis phycocyanin (65 °C). The C. merolae phycocyanin was relatively stable at pH 4 and 5 up to 80 °C. The high thermostability at slightly acidic pH makes the C. merolae phycocyanin an interesting alternative to A. platensis phycocyanin as a natural blue food colorant.     

Modeling seed germination in Melisa officinalis L. in response to temperature and water potential

Seed germination is greatly influenced by both temperature (T) and water potential (ψ) and these factors largely determine germination rate (GR) in the field. Quantitative information about T and ψ effects on seed germination in lemon balm (Melisa officinalis L.) is scarce. The main objective of this study was to quantify seed germination responses of lemon balm to T and ψ, and to determine cardinal temperatures in a laboratory experiment. A segmented model was used to describe the effects of ψ (i.e., T) on GR and other germination parameters. The segmented model estimates were 7.2 °C for base (T _b), 28.9 °C for optimum (T _o), 40.1 °C for ceiling temperature (T _c) and 1.64 physiological days (f _o) (equivalent to a GR_max of 0.610 d^?1 and a thermal time of 35.6 °C days) to reach 50 % maximum germination in the control (0 MPa) treatment (R ² = 0.99, RMSE = 0.005 day^?1). The inherent maximum rate of germination (days) was calculated by the [GR_max = 1/f _o] model. ψ affected cardinal temperatures. From 0 to ?0.76 MPa, when ψ increased, T _b was a constant 7.2 °C to ?0.38 MPa and increased linearly to 20.1 °C as ψ decreased. T _o and f _o increased linearly from 28.9 to 30 °C, and from 1.64 to 5.4 day^?1, respectively as ψ decreased. However, there was no signification difference in T _o as ψ decreased nor did T _c decrease from 40.1 to 35 °C as ψ decreased. T _b, T _c and GR_max were the sole parameters affected by ψ and could be used to characterize differences between ψ treatments with respect to GR at various Ts. Therefore, the segmented model and its parameters can be used in lemon balm germination simulation models.     

Optimization of arylacetonitrilase production from Alcaligenes sp. MTCC 10675 and its application in mandelic acid synthesis

Alcaligenes sp. MTCC 10675 has been isolated from soil sample using enrichment method and has nitrilase catalytic system which is highly specific for the hydrolysis of arylaliphatic nitriles. Optimization of culture conditions using response surface methodology and inducer-mediated approach enhanced arylacetonitrilase production significantly (2.4-fold). Isobutyronitrile acted as an effective inducer for the induction of arylacetonitrilase, and it is highly specific for arylacetonitriles (phenyl acetonitrile and mandelonitrile). Arylacetonitrilase has no effect on its relative velocity (V _r) up to 20 mM substrate (mandelonitrile) concentration and at 30 mM mandelonitrile, 23.4 % degree of inhibition (I _d) was recorded. Half life of arylacetonitrilase of Alcaligenes sp. MTCC 10675 was 27.5 h at 25 °C. Hg²⁺, Ag⁺, Pb³⁺, and Co²⁺ were strong inhibitor of arylacetonitrilase activity which resulted into 100 %, 91 %, 84 %, and 83 % inhibition, respectively. Polar protic solvent (dichloromethane, dimethylsulphooxide, and n-butanol) reduce arylacetonitrilase activity up to 80–94 % at 10 % concentration. Alcaligenes sp. MTCC 10675 has higher biocatalytic activity, i.e., 3.9 gg^-1 dcw, which is highest in comparison to till reported organism. Arylacetonitrilase-mediated hydrolysis of racemic mandelonitrile resulted into R-(-) mandelic acid with 99.0 % enantiomeric excess (e.e.)     

Nitrous oxide emissions and nitrate leaching from a rain-fed wheat-maize rotation in the Sichuan Basin, China

Aims

A 3-year field experiment (October 2004–October 2007) was conducted to quantify N₂O fluxes and determine the regulating factors from rain-fed, N fertilized wheat-maize rotation in the Sichuan Basin, China.

Methods

Static chamber-GC techniques were used to measure soil N₂O fluxes in three treatments (three replicates per treatment): CK (no fertilizer); N150 (300 kg N fertilizer ha^?1 yr^?1 or 150 kg N?ha^?1 per crop); N250 (500 kg N fertilizer ha^?1 yr^?1 kg or 250 kg N?ha^?1 per crop). Nitrate (NO ₃ ^? ) leaching losses were measured at nearby sites using free-drained lysimeters.

Results

The annual N₂O fluxes from the N fertilized treatments were in the range of 1.9 to 6.7 kg N?ha^?1 yr^?1 corresponding to an N₂O emission factor ranging from 0.12 % to 1.06 % (mean value: 0.61 %). The relationship between monthly soil N₂O fluxes and NO ₃ ^- leaching losses can be described by a significant exponential decaying function.

Conclusions

The N₂O emission factor obtained in our study was somewhat lower than the current IPCC default emission factor (1 %). Nitrate leaching, through removal of topsoil NO ₃ ^? , is an underrated regulating factor of soil N₂O fluxes from cropland, especially in the regions where high NO ₃ ^- leaching losses occur.     

Isolation of natural red colorants from fermented broth using ionic liquid-based aqueous two-phase systems

There is a growing demand for natural colorants. This is prompting the search for new alternative and “benign” separation systems allowing higher recoveries, extraction yields, and selectivities. This work investigates the use of aqueous two-phase systems (ATPS) based on ionic liquids as extraction processes for the recovery of red colorants from the fermented broth of Penicillium purpurogenum DPUA 1275. Several ATPS based on quaternary ammonium and imidazolium were studied in this work aiming at separating the red colorants produced from the remaining colorants and contaminant proteins present in the fermented broth. The results suggest that the red colorants can be isolated by an appropriate manipulation of some of the process conditions, such as the use of quaternary ammonium with short alkyl chains, alkaline media, and short tie-line lengths (extraction point systems with lower concentrations of ionic liquid). These conditions allow large partition coefficients for the red colorants (K _red = 24.4 ± 2.3), high protein removal (60.7 ± 2.8 %) and selectivity parameters (S _red/prot = 10.05).     

Inhibition of autophagy stimulate molecular iodine-induced apoptosis in hormone independent breast tumors

Estrogen receptor negative (ER^−ve) and p53 mutant breast tumors are highly aggressive and have fewer treatment options. Previously, we showed that molecular Iodine (I₂) induces apoptosis in hormone responsive MCF-7 breast cancer cells, and non-apoptotic cell death in ER^−ve–p53 mutant MDA-MB231 cells (Shrivastava, 2006). Here we show that I₂ (3 μM) treatment enhanced the features of autophagy in MDA-MB231 cells. Since autophagy is a cell survival response to most anti-cancer therapies, we used both in vitro and in vivo systems to determine whether ER^−ve mammary tumors could be sensitized to I₂-induced apoptosis by inhibiting autophagy. Autophagy inhibition with chloroquine (CQ) and inhibitors for PI3K (3MA, LY294002) and H+/ATPase (baflomycin) resulted in enhanced cell death in I₂ treated MDA-MB231 cells. Further, CQ (20 μM) in combination with I₂, showed apoptotic features such as increased sub-G1 fraction (∼5-fold), expression of cleaved caspase-9 and -3 compared to I₂ treatment alone. Flowcytometry of I₂ and CQ co-treated cells revealed increase in mitochondrial membrane permeability (p < 0.01) and translocation of cathepsin D activity to cytosol relative to I₂ treatment. For in vivo studies ICRC mice were transplanted subcutaneously with MMTV-induced mammary tumors. A significant reduction in tumor volumes, as measured by MRI, was found in I₂ and CQ co-treated mice relative to I₂ or vehicle treated mice. These data indicate that inhibition of autophagy renders ER^−ve breast tumor cells more sensitive to I₂ induced apoptosis. Thus, I₂ together with autophagy inhibitor could have a potential tumorostatic role in ER^−ve aggressive breast tumors that may be evaluated in future studies.     

A Counter-Current Acid Leaching Process for the Remediation of Contaminated Soils from a Small-Arms Shooting Range

The objective of this research was to use a counter-current leaching process (CCLP) with leachate treatment to develop a remediation process for contaminated soils at a small-arms shooting range (SASR). The soil contaminant concentrations were 245 mg Cu kg^?1, 3,368 mg Pb kg^?1, 73 mg Sb kg^?1, and 177 mg Zn kg^?1. The CCLP includes three acid leaching steps (1M H₂SO₄ + 4M NaCl, t = 1 h, T = 20°C, soil suspension = 100 g L^?1), followed by one water rinsing step (1 h). Seven counter-current remediation cycles were completed, and the average resulting metal removals were 93.2 ± 3.5% of Cu, 91.5 ± 5.7% of Pb, 82.2 ± 10.9% of Sb, and 30.0 ± 11.4% of Zn. The metal leaching performances decreased with the number of completed cycles. Soil treated with the CCLP with leachate treatment process met the USEPA threshold criteria of 5 mg Pb L^?1 in the TCLP leachate. The CCLP allows a decrease of the water use by 32.9 m³ t^?1 and the chemicals’ consumption by approximately 2,650 kg H₂SO₄, 6,014 kg NaCl, and 1,150 kg NaOH per ton of treated soil, in comparison to standard leaching processes. This corresponds to 78%, 69%, 83%, and 67% of reduction, respectively.     

An effective method for the detoxification of cyanide-rich wastewater by Bacillus sp. CN-22

The biodetoxification of cyanide-rich wastewater has become increasingly popular because of its cost-effectiveness and environmental friendliness. Therefore, we have developed an effective method, optimised by response surface methodology, for detoxifying cyanide-rich wastewater using Bacillus sp. CN-22, which was newly isolated from a cyanide-contaminated electroplating sludge and could tolerate a CN^? concentration of 700 mg L^?1. The concentration of CN^? in the treated wastewater decreased from 200 to 6.62 mg L^?1 after cultivation with 2.38 % inocula for 72 h on the medium, consisting of 0.05 % KH₂PO₄, 0.15 % K₂HPO₄, 1.0 mM MgCl₂, 1.0 mM FeCl₃, 0.1 % NH₄Cl, and 0.1 % glycerol. The CN^? degradability of 96.69 % is similar to the predicted value of 96.82 %. The optimal cultivation conditions were controlled as follows: initial pH, 10.3; temperature, 31 °C; and rotary speed, 193 rpm. The maintenance of higher pH in the overall treatment procedures may avoid the production of volatile HCN and the risk associated with cyanide detoxification. Additionally, the bacterial strain Bacillus sp. CN-22, with its potent cyanide-degrading activity at the initial CN^– concentration of 200 mg L^?1, may be employed to effectively treat cyanide-rich wastewater, especially electroplating effluent.     

Assessment of nitrate leaching loss on a yield-scaled basis from maize and wheat cropping systems

Background and aims

It is so far a gap in knowledge to assess nitrate (NO₃ ^?) leaching loss linking with crop yield for a given cereal cropping system.

Methods

We conducted a meta-analysis on 32 published studies reporting both NO₃ ^? leaching losses and crop yields in the maize (N?=?20) and wheat (N?=?12) systems.

Results

On average, 22 % and 15 % of applied fertilizer N to wheat and maize systems worldwide are leached in the form of NO₃ ^?, respectively. The average area-scaled NO₃ ^- leaching loss for maize (57.4 kg N ha^?1) was approx. two times higher than for wheat (29.0 kg N ha^?1). While, if scaled to crop yields, the average yield-scaled NO₃ ^? losses were comparable between maize (5.40 kg N Mg^?1) and wheat (5.41 kg N Mg^?1) systems. Across all sites, the lowest yield-scaled NO₃ ^? leaching losses were observed at slightly suboptimal fertilization rates, corresponding to 90 % and 96 % of maximum maize or wheat yields, respectively.

Conclusions

Our findings suggest that small adjustments of agricultural N management practices can effectively reduce yield-scaled NO₃ ^? leaching losses. However, further targeted field experiments are still needed to identify at regional scale best agricultural management practices for reducing yield-scaled NO₃ ^? leaching losses in maize and wheat systems.     

Homocysteine-Induced Apoptosis in Endothelial Cells Coincides With Nuclear NOX2 and Peri-nuclear NOX4 Activity

Apoptosis of endothelial cells related to homocysteine (Hcy) has been reported in several studies. In this study, we evaluated whether reactive oxygen species (ROS)-producing signaling pathways contribute to Hcy-induced apoptosis induction, with specific emphasis on NADPH oxidases. Human umbilical vein endothelial cells were incubated with 0.01–2.5 mM Hcy. We determined the effect of Hcy on caspase-3 activity, annexin V positivity, intracellular NOX1, NOX2, NOX4, and p47^phox expression and localization, nuclear nitrotyrosine accumulation, and mitochondrial membrane potential (ΔΨ _m). Hcy induced caspase-3 activity and apoptosis; this effect was concentration dependent and maximal after 6-h exposure to 2.5 mM Hcy. It was accompanied by a significant increase in ΔΨ _m. Cysteine was inactive on these parameters excluding a reactive thiol group effect. Hcy induced an increase in cellular NOX2, p47^phox, and NOX4, but not that of NOX1. 3D digital imaging microscopy followed by image deconvolution analysis showed nuclear accumulation of NOX2 and p47^phox in endothelial cells exposed to Hcy, but not in control cells, which coincided with accumulation of nuclear nitrotyrosine residues. Furthermore, Hcy enhanced peri-nuclear localization of NOX4 coinciding with accumulation of peri-nuclear nitrotyrosine residues, a reflection of local ROS production. p47^phox was also increased in the peri-nuclear region. The Hcy-induced increase in caspase-3 activity was prevented by DPI and apocynin, suggesting involvement of NOX activity. The data presented in this article reveal accumulation of nuclear NOX2 and peri-nuclear NOX4 accumulation as potential source of ROS production in Hcy-induced apoptosis in endothelial cells.     

Comparison of ginsenoside composition in native roots and cultured callus cells of Panax quinquefolium L.

In order to compare the ginsenoside composition in native Panax quinquefolium and in suspension cultured cells derived from root callus, HPLC–ESI-MSⁿ analysis was performed. Under the present HPLC–ESI-MSⁿ conditions, ten ginsenosides from native root were acquired in the positive and negative ion modes, namely Rg₁, Re, Ro, malonyl-Rb₁, Rf, Rb₁, Rc, Rb₂, Rb₃ and Rd. Only four ginsenosides (Rg1, Re, Rf and Rb1) were identified from callus cells. Radical scavenging activity of P. quinquefolium callus cells with 250 mg l^?1 methanolic extract on 1,1-diphenyl-2-picrylhydrazyl (DPPH) was 55.72 %, while only 6.31 % DPPH inhibition was obtained in native root.     

Co-expression of <Emphasis Type="SmallCaps">l</Emphasis>-glutamate oxidase and catalase in <Emphasis Type="Italic">Escherichia coli</Emphasis> to produce α-ketoglutaric acid by whole-cell biocatalyst

Objectives

To improve the production of α-ketoglutaric acid (α-KG) from l-glutamate by whole-cell biocatalysis.

Results

A novel and highly active l-glutamate oxidase, SmlGOX, from Streptomyces mobaraensis was overexpressed and purified. The recombinant SmlGOX was approx. 64 kDa by SDS-PAGE. SmlGOX had a maximal activity of 125 ± 2.7 U mg^?1 at pH 6.0, 35 ^oC. The apparent K_m and V_max values of SmlGOX were 9.3 ± 0.5 mM and 159 ± 3 U mg^?1, respectively. Subsequently, a co-expression plasmid containing the SmlGOX and KatE genes was constructed to remove H₂O₂, and the protein levels of SmlGOX were improved by codon optimization. Finally, by optimizing the whole-cell transformation conditions, the production of α-KG reached 77.4 g l^?1 with a conversion rate from l-glutamate of 98.5% after 12 h.

Conclusions

An efficient method for the production of α-KG was established in the recombinant Escherichia coli, and it has a potential prospect in industrial application.

     

Improving EGSB reactor performance for simultaneous bioenergy and organic acid production from cheese whey via continuous biological H<Subscript>2</Subscript> production

Objectives

To evaluate the influence of hydraulic retention time (HRT) and cheese whey (CW) substrate concentration (15 and 25 g lactose l^?1) on the performance of EGSB reactors (R15 and R25, respectively) for H₂ production.

Results

A decrease in the HRT from 8 to 4 h favored the H₂ yield and H₂ production rate (HPR) in R15, with maximum values of 0.86 ± 0.11 mmol H₂ g COD^?1 and 0.23 ± 0.024 l H₂ h^?1 l^?1, respectively. H₂ production in R25 was also favored at a HRT of 4 h, with maximum yield and HPR values of 0.64 ± 0.023 mmol H₂ g COD^?1 and 0.31 ± 0.032 l H₂ h^?1 l^?1, respectively. The main metabolites produced were butyric, acetic and lactic acids.

Conclusions

The EGSB reactor was evaluated as a viable acidogenic step in the two-stage anaerobic treatment of CW for the increase of COD removal efficiency and biomethane production.

     

Cytokine Responses to Novel Antigens in an Indian Population Living in an Area Endemic for Visceral Leishmaniasis

Background

There are no effective vaccines for visceral leishmaniasis (VL), a neglected parasitic disease second only to malaria in global mortality. We previously identified 14 protective candidates in a screen of 100 Leishmania antigens as DNA vaccines in mice. Here we employ whole blood assays to evaluate human cytokine responses to 11 of these antigens, in comparison to known defined and crude antigen preparations.

Methods

Whole blood assays were employed to measure IFN-γ, TNF-α and IL-10 responses to peptide pools of the novel antigens R71, Q51, L37, N52, L302.06, J89, M18, J41, M22, M63, M57, as well as to recombinant proteins of tryparedoxin peroxidase (TRYP), Leishmania homolog of the receptor for activated C kinase (LACK) and to crude soluble Leishmania antigen (SLA), in Indian patients with active (n = 8) or cured (n = 16) VL, and in modified Quantiferon positive (EHC^+ve, n = 20) or modified Quantiferon negative (EHC^−ve, n = 9) endemic healthy controls (EHC).

Results

Active VL, cured VL and EHC^+ve groups showed elevated SLA-specific IFN-γ, but only active VL patients produced IL-10 and EHC^+ve did not make TNF-α. IFN-γ to IL-10 and TNF-α to IL-10 ratios in response to TRYP and LACK antigens were higher in cured VL and EHC^+ve exposed individuals compared to active VL. Five of the eleven novel candidates (R71, L37, N52, J41, and M22) elicited IFN-γ and TNF-α, but not IL-10, responses in cured VL (55–87.5% responders) and EHC^+ve (40–65% responders) subjects.

Conclusions

Our results are consistent with an important balance between pro-inflammatory IFNγ and TNFγ cytokine responses and anti-inflammatory IL-10 in determining outcome of VL in India, as highlighted by response to both crude and defined protein antigens. Importantly, cured VL patients and endemic Quantiferon positive individuals recognise 5 novel vaccine candidate antigens, confirming our recent data for L. chagasi in Brazil, and their potential as cross-species vaccine candidates.