Nesting behaviour and foraging characteristics of Andrena cineraria (Hymenoptera: Andrenidae)

The current studies were carried out in the three experimental locations of Kashmir valley during 2013 to 2016. The species Andrena cineraria formed the dense nest aggregations in plan grounds, barren lands and hilly areas near the fruit orchards and other landscapes with clay loam soil type. The species start flying and foraging in the orchards from April till July. The nests were allodalous, 29–36 cm in depth, with cells located obliquely around the main barrow. The nests were dense with a maximum density of 11.09 nests/m² observed in landscapes of Budgam. The barrow diameters were found varying with depth from main entrance. The maximum barrow diameter recorded was 2.05 mm. At certain depth, the female constructs the first cell and the upper nest burrow is vertical and lower is oblique. The nest entrance is generally hidden under the tumulus. In the depth of average 30.48 cm, each cell directly opens to main burrow either alternately or unilaterally. The cell number, diameter, and length varied with depth. Foraging behaviour of A. cineraria on various fruit crops and other shrubs and social forestry trees were determined and the abundance, visitation rate, total visits and time spend per flower were found significant, especially on fruit crops. The significance of the studies is important for the melittologists, as it will help in the conservation of bee fauna. The study is also important in using this species for pollination purpose and would also help to detect and understand the possible pre-adaptation of species in temperate region of Kashmir valley.     

Effects of dietary selenium and vitamin E on immune response and biological blood parameters of broilers reared under thermoneutral or heat stress conditions

A study was conducted using 360 broiler chickens to evaluate the effects of dietary vitamin E (0, 125 and 250 mg/kg), selenium (Se, 0, 0.5 and 1 mg/kg), or their different combinations on immune response and blood biological parameters of broilers raised under either thermoneutral (TN, 23.9 °C constant) or heat stress (HS, 23.9 to 37 °C cycling) conditions. Humoral immunity was assessed by intravenous injection of 7 % sheep red blood cell (SRBC) followed by evaluation of serum for antibody titers in primary and secondary responses. Heterophil to lymphocyte (H/L) ratio also determined as an indicator of stress. Furthermore, at the end of the experiment, birds were bled for determination of some biological parameters. There was a significant reduction in body weight and feed intake, but the feed conversion ratio increased when the birds were exposed to HS (P?<?0.05). Body weight and feed intake were not influenced significantly by dietary vitamin E and Se (P?>?0.05), whereas feed conversion was improved significantly by 125 mg/kg vitamin E (P?<?0.05). The liver and lymphoid organ weights as well as IgM and IgG, antibody titers for primary and secondary antibody responses to SRBC were reduced significantly under HS (P?<?0.05). Heat stress also resulted in a significant increase in H/L ratio (P?<?0.05). Dietary vitamin E resulted in improvement of primary and secondary antibody responses both in TN and HS broilers (P?<?0.05). The HS birds also showed an improved antibody titer in secondary response with high concentration of Se (P?<?0.05). Vitamin E and Se had interactive effects on anti-SRBC titers; however, no consistent differences were found between dietary levels during the study. The H/L ratio decreased by feeding vitamin E at both levels either under HS or TN conditions (P?<?0.05). The serum concentrations of glucose, triglycerides, total cholesterol, and LDL-cholesterol were increased but serum HDL-cholesterol decreased in HS broilers (P?<?0.05).     

Effect of maternal nitrogen and drought stress on seed dormancy and germinability of Amaranthus retroflexus

Amaranthus retroflexus L. is an importunate annual weed in many cropping systems of different countries. The main aim of this study was to investigate the effects of maternal nitrogen and drought stress on the seed dormancy and germinability of A. retroflexus. Field experiment was carried out in a factorial based on randomized complete block design, with four potential levels of soil water (–2, ?6, ?8 and ?10 bar) and three levels of nitrogen (0, 100 and 200 kg/ha). The germination characteristics of the seeds were measured at three different times (1 month, 6 months and 1 year after harvesting). Results showed that drought stress had positive effects on breaking of A. retroflexus seed dormancy until 6 months after seed harvesting. Seeds that were developed under severe water stress exhibited the highest germination percentage and germination rate. The results obtained from this study revealed that application of 100 kg/ha nitrogen during seed development increases germinability of A. retroflexus, whereas application of 200 kg/ha nitrogen induced seed dormancy. Furthermore, 100 kg/ha nitrogen application in the field along with 200 ppm gibberellic‐acid treatment during seed after‐ripening showed the highest germination percentage and germination rate for seeds after 6 months harvesting. Results also indicated that after‐ripening significantly increased seed germination and germination rate of A. retroflexus. These findings indicate that long‐term management of the soil seed bank in this species requires more stringent control due to the changes in germination timing, as detected in this study.     

Production,purification and titration of a lentivirus-based vector for gene delivery purposes

Viral vectors are valuable tools to deliver genetic materials into cells. Vectors derived from human immunodeficiency virus type 1 are being widely used for gene delivery, mainly because they are able to transduce both dividing and non-dividing cells which leads to stable and long term gene expression. In addition, these types of vectors are safe, with low toxicity, high stability and cell type specificity. Therefore, this work was aimed to produce lentivirus-based vector using a three-plasmid system. To produce this system, the eGFP marker gene was cloned into the plasmid pWPXLd. Subsequently, this vector plasmid, along with packaging plasmids, psPAX2 and envelope plasmid, pMD2.G, was co-transfected into packaging cell line (293T) using calcium phosphate method. 48 h post transfection, the constructed viral vector was harvested, purified and concentrated and stored at −80 °C for next experiments. The titration of the vector was carried out, using ELISA, flowcytometry, and fluorescent microscopy. Finally, transduction of HEK-293T, CHO, HepG2, MCF-7, MEFs and Jurkat cell lines was carried out with indicated cell numbers and multiplicities of infections of the vector in the presence of polybrene. Using this system, high titer lentivirus at titers of up to 2 × 10⁸ transducing units/ml (TU/ml) was successfully generated and its transduction efficacy was improved by seven to over 20-fold in various cell types. We demonstrate the applicability of this vector for the efficient transduction of dividing and non-dividing cells, including HEK-293T, CHO, HepG2, MCF-7, MEFs and Jurkat cell line. Transduction efficiency yielded titers of (6.3 ± 1.2) 10⁵ TU/ml. Furthermore, lentivirus transferred transgene was expressed at high level in the target cells and expression was followed until 90 days after transduction. Thus, the vector generated in this work, might be able to deliver the transgene into a wide range of mammalian cells.

Electronic supplementary material

The online version of this article (doi:10.1007/s10616-013-9652-5) contains supplementary material, which is available to authorized users.     

Disbudding treatments on pistachio trees cv. mateur: dry matter accumulation and distribution within fruiting and non-fruiting branches under dry climate

Key message

After applying disbudding treatments, removal of fifty percent of flower bud each year improves dry matter accumulation in fruiting and non-fruiting branches of pistachio trees, which could minimize alternate bearing.

Abstract

Dry matter accumulation and distribution within branches of pistachio trees were investigated during 2005-2008 to determine the effects of fruiting on shoot growth under rain-fed conditions in arid climate. Four treatments were applied: T ₀ normal alternation cycle, T ₁ trees disbudded for 1 year, T ₂ trees disbudded for two successive years, and T ₃ removal of 50 % of all floral buds for each year. Consecutive disbudded treatment (T ₂) allowed a higher growth potential of pistachio trees with reference to normal biennial cycle (T ₀). Individual current shoot of T ₂ accumulated 44 % as much dry matter cm^?1 as those of ‘On’ trees, which have the highest yield. Removal of 50 % of floral buds (T ₃) significantly increased the dry matter accumulation in fruiting branch to reach 57 g in postharvest compared to 42.6 g for the control T ₀. Trees disbudded for 2 years (T ₂) had increased dry matter accumulation in the non-fruiting branch from 3.3 to 16.3 g. Leaves, current shoot, 1-year-old wood and inflorescence buds represented, respectively, 87, 5.3, 5 and 2.7 % of the total dry matter of individual branch of T ₂. In fruiting branches, nuts consisted of 83–87 %, leaves 7–10 %, rachises 4 %, 1-year-old wood 1.6–2 % and current shoot 0.8–1.3 % of the total dry matter. One-year-old wood played a major role as sources and sinks for developing current year shoot, leaves, inflorescence buds and nuts. Removal of 50 % of flower bud (T ₃) improves the dry matter accumulation in fruiting and non-fruiting branches. Thus, under rain-fed conditions, annual pruning could be used to minimize alternate bearing of pistachio.     

The effect of polymers on the stability of microencapsulated formulations of Bacillus thuringiensis subsp. kurstaki (Bt-KD2) after exposure to Ultra Violet Radiation

We compared the protective effect of three polymers; starch, gelatin and sodium alginate (2, 3, 5%) as coating materials, on the stability of microencapsulated formulation of Bacillus thuringiensis after exposure to Ultra Violet (UV) R. Microencapsulated Spore Crystal Aggregate (SCA) formulations were prepared by the emulsion gelling method. The protective effect of polymers was evaluated by measuring spore viability. Bioassay and release tests were done on the microencapsulated formulations. Sodium alginate (5% w/w) showed the highest viabilities of 90 and 86% after exposure to Ultra Violet in long term (UVB 385 nm) and Ultra Violet in short term (UVC 254 nm) radiation, respectively, while viabilities of non-microencapsulated spores under these conditions were 40 and 50%, respectively. The crystal activity (mortality) of irradiated and non-irradiated free spore formulations on second-instar larvae of Ephestia kuehniella were 15 and 93%. However, the mortalities caused by irradiated and non-irradiated microencapsulated formulations were 70 and 80% on the 10th day of the experiment. The size range of the microcapsules was 7–20 µm while the microcapsulation efficiency was 86%. The release behaviour of microspheres conformed best to Korsmeyer–Peppas semi-empirical model with the correlation of R² = 0.98.     

Interaction of two new mixed ligand copper(II) complexes with DNA probed by thermodynamic and spectroscopic studies

The DNA binding behavior of [Cu(4,7-dmp)(phen-dione)Cl]Cl (1) and [Cu(2,9-dmp)(phen-dione)Cl]Cl (2) where dmp and phen-dion stand for dimethyl-1,10-phenanthroline and 1,10-phenanthroline-5,6-dion, respectively, was studied with a series of techniques including Viscometry, UV–Vis absorption, circular dichroism and fluorescence spectroscopy. Cytotoxicity effect was also investigated. Thermodynamic parameters, enthalpy and entropy changes were calculated according to Van’t Hoff equation, which indicated that both reactions are predominantly enthalpically driven. However, these two complexes show different behavior in fluorescence, circular dichroism and viscometry methods which indicate the Cu(II) complexes interact with calf-thymus DNA by different mode of binding. These have further been verified by competition studies using Hoechst as a distinct groove binder. All these results indicate that these two complexes (1) and (2) interact with CT-DNA via groove binding and partially intercalative mode, respectively and the binding affinity of the complex 1 is higher than that of complex 2. Finally, our findings suggest that the type of ligands and structure of complexes have marked effect on the binding affinity of complexes involving CT-DNA. Also, these new complexes showed excellent antitumor activity against human T lymphocyte carcinoma-Jurkat cell line.     

Study on the interaction of a copper(II) complex containing the artificial sweetener aspartame with human serum albumin

A copper(II) complex containing aspartame (APM) as ligand, Cu(APM)₂Cl₂·2H₂O, was synthesized and characterized. In vitro binding interaction of this complex with human serum albumin (HSA) was studied at physiological pH. Binding studies of this complex with HSA are useful for understanding the Cu(APM)₂Cl₂·2H₂O–HSA interaction mechanism and providing guidance for the application and design of new and more efficient artificial sweeteners drive. The interaction was investigated by spectrophotometric, spectrofluorometric, competition experiment and circular dichroism. Hyperchromicity observed in UV absorption band of Cu(APM)₂Cl₂·2H₂O. A strong fluorescence quenching reaction of HSA to Cu(APM)₂Cl₂·2H₂O was observed and the binding constant (K_f) and corresponding numbers of binding sites (n) were calculated at different temperatures. Thermodynamic parameters, enthalpy change (?H) and entropy change (?S) were calculated to be ?458.67 kJ mol^?1 and ?1,339 J mol^?1 K^?1 respectively. According to the van’t Hoff equation, the reaction is predominantly enthalpically driven. In conformity with experimental results, we suggest that Cu(APM)₂Cl₂·2H₂O interacts with HSA. In comparison with previous study, it is found that the Cu(II) complex binds stronger than aspartame.     

A Comparative Approach to Strategies for Cloning,Expression, and Purification of Mycobacterium tuberculosis Mycolyl Transferase 85B and Evaluation of Immune Responses in BALB/c Mice

Protein antigens have drawn a lot of attention from investigators working on tuberculosis vaccines. These proteins can be used to improve the immunogenicity of the new generation BCG vaccines or even replace them completely. Recombinant technology is used to insure the production of pure mycobacterial antigens in high quantities. Mycolyl transferase 85B (Ag85B) is a potent, mycobacterial antigen that significantly stimulates immune responses. Since Ag85B is an apolar protein, production of the water-soluble antigen is of interest. In this work, we report a systematic optimization strategy concerning cloning systems and purification methods, aiming at increasing the yield of recombinant Ag85B. Our optimized method resulted in a yield of 8 mg of recombinant Ag85B from 1 liter of induced culture (400 μg/ml) by using pET32a(+), Escherichia coli Rosseta-gami?(DE3) pLysS and a Ni–NTA agarose-based procedure and on-column re-solubilization. The purified recombinant Ag85B showed strong immunostimulating properties by inducing high levels of TNF-α, IFN-γ, IL-12, and IgG2a in immunized mice, therefore it can effectively be applied in TB vaccine researches.     

Label-free measurement of histone lysine methyltransferases activity by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry

Histone lysine methyltransferases (HKMTs) are enzymes that play an essential role in epigenetic regulation. Thus, identification of inhibitors specifically targeting these enzymes represents a challenge for the development of new antitumor therapeutics. Several methods for measuring HKMT activity are already available. Most of them use indirect measurement of the enzymatic reaction through radioactive labeling or antibody-recognized products or coupled enzymatic assays. Mass spectrometry (MS) represents an interesting alternative approach because it allows direct detection and quantification of enzymatic reactions and can be used to determine kinetics and to screen small molecules as potential inhibitors. Application of mass spectrometry to the study of HKMTs has not been fully explored yet. We describe here the development of a simple reliable label-free MALDI-TOF MS-based assay for the detection and quantification of peptide methylation, using SET7/9 as a model enzyme. Importantly, the use of expensive internal standard often required in mass spectrometry quantitative analysis is not necessary in this assay. This MS assay allowed us to determine enzyme kinetic parameters as well as IC₅₀ for a known inhibitor of this enzyme. Furthermore, a comparative study with an antibody-based immunosorbent assay showed that the MS assay is more reliable and suitable for the screening of inhibitors.