Imaging techniques for the detection of stored product pests

Stored grains are subject to deterioration and losses through various factors, but mainly insects and fungi. Various techniques are employed to detect stored product pests; however, there is an urgent need for an industrial-scale on-line detection technique. Near-infrared hyperspectroscopic imaging and soft X-rays have shown the potential for real-time application. These techniques are particularly effective for detecting internal infestations of stored grains. The digital images of the scanned objects are analyzed for various spectral and image features using statistical techniques such as complex multivariate tools. Classification accuracies as high as 80–100 % have been achieved for various pest and grain combinations. Dual-energy X-rays have been shown to detect the concealed eggs of stored product insect pests. The main threats to stored cereals come from Aspergillus spp., Penicillium spp., and Fusarium spp., which may produce mycotoxins. These imaging techniques have shown good results in the detection of fungal infections of stored grain.     

Monitoring Cell-autonomous Circadian Clock Rhythms of Gene Expression Using Luciferase Bioluminescence Reporters

In mammals, many aspects of behavior and physiology such as sleep-wake cycles and liver metabolism are regulated by endogenous circadian clocks (reviewed^1,2). The circadian time-keeping system is a hierarchical multi-oscillator network, with the central clock located in the suprachiasmatic nucleus (SCN) synchronizing and coordinating extra-SCN and peripheral clocks elsewhere^1,2. Individual cells are the functional units for generation and maintenance of circadian rhythms^3,4, and these oscillators of different tissue types in the organism share a remarkably similar biochemical negative feedback mechanism. However, due to interactions at the neuronal network level in the SCN and through rhythmic, systemic cues at the organismal level, circadian rhythms at the organismal level are not necessarily cell-autonomous^5-7. Compared to traditional studies of locomotor activity in vivo and SCN explants ex vivo, cell-based in vitro assays allow for discovery of cell-autonomous circadian defects^5,8. Strategically, cell-based models are more experimentally tractable for phenotypic characterization and rapid discovery of basic clock mechanisms^5,8-13.Because circadian rhythms are dynamic, longitudinal measurements with high temporal resolution are needed to assess clock function. In recent years, real-time bioluminescence recording using firefly luciferase as a reporter has become a common technique for studying circadian rhythms in mammals^14,15, as it allows for examination of the persistence and dynamics of molecular rhythms. To monitor cell-autonomous circadian rhythms of gene expression, luciferase reporters can be introduced into cells via transient transfection^13,16,17 or stable transduction^5,10,18,19. Here we describe a stable transduction protocol using lentivirus-mediated gene delivery. The lentiviral vector system is superior to traditional methods such as transient transfection and germline transmission because of its efficiency and versatility: it permits efficient delivery and stable integration into the host genome of both dividing and non-dividing cells²⁰. Once a reporter cell line is established, the dynamics of clock function can be examined through bioluminescence recording. We first describe the generation of P(Per2)-dLuc reporter lines, and then present data from this and other circadian reporters. In these assays, 3T3 mouse fibroblasts and U2OS human osteosarcoma cells are used as cellular models. We also discuss various ways of using these clock models in circadian studies. Methods described here can be applied to a great variety of cell types to study the cellular and molecular basis of circadian clocks, and may prove useful in tackling problems in other biological systems.     

The potential therapeutic use of stem cells in cartilage repair

As our population demographics change, osteoarthritis and cartilage defects are becoming more prevalent. The discovery of stems cells and their ability for indefinite regeneration has revolutionised the way cartilage problems are viewed. Tissue engineering has been shown to be the ideal way of repairing articular cartilage lesions, i.e. back to native tissue. Cartilage is an ideal tissue engineering target as it is avascular, aneural and alymphatic. The two main types of stem cells being investigated in chondrogenesis are embryological and mesenchymal stem cells. Research into embryological stem cells has been surrounded by controversy because of ethical, religious and social concerns. We discuss the use of embryological and mesenchymal stem cells in cartilage repair and the various factors involved in the differentiation into chondrocytes. We also discuss commonly used mesenchymal stem cell markers and their limitations.     

Persistent but reversible coronary microvascular dysfunction after bypass grafting

The effect of coronary artery bypass grafting (CABG) on absolute myocardial blood flow (MBF) has not been investigated previously. MBF (ml. min(-1). g(-1)) was measured at rest and during hyperemia (0.56 mg/kg iv dipyridamole) using H(2)(15)O and positron emission tomography in eight patients with three-vessel disease before surgery and 1 and 6 mo after full revascularization. Baseline MBF was 0.87 +/- 0.12 preoperatively and 1.04 +/- 0.14 and 0.95 +/- 0.13 at 1 and 6 mo after CABG, respectively (P < 0.05, 6 mo vs. preoperatively). Hyperemic MBF was 1.36 +/- 0.28 preoperatively and increased to 1.98 +/- 0.50 and 2.45 +/- 0.64 at 1 and 6 mo after CABG, respectively (P < 0.01, 6 mo vs. preoperatively). Coronary vasodilator reserve (hyperemic/baseline MBF) increased from 1.59 +/- 0.40 preoperatively to 1.93 +/- 0.13 and 2.57 +/- 0.49 at 1 and 6 mo, respectively (P < 0.05, 6 mo vs. preoperatively). Minimal (dipyridamole) coronary resistance (mmHg. min. g(-1). ml(-1)) fell progressively from 59.37 +/- 14.56 before surgery to a nadir of 35. 76 +/- 10.12 at 6 mo after CABG (P < 0.01 vs. preoperatively). The results of the present study confirm that CABG improves coronary vasodilator reserve progressively as a result of reduction in minimal coronary resistance. These data suggest persistent microvascular dysfunction that recovers slowly after surgery.     

Interaction between PGPR and PGR for water conservation and plant growth attributes under drought condition

Drought is one of the key restraints to agricultural productivity worldwide and is expected to increase further. Drought stress accompanied by reduction in precipitation pose major challenges to future food safety. Strategies should be develop to enhance drought tolerance in crops like chickpea and wheat, in order to enhance their growth and yield. Drought tolerance strategies are costly and time consuming however, recent studies specify that plant growth promoting rhizobacteria (PGPR) and plant growth regulators (PGRs) can help plants to withstand under harsh environmental condition and enable plants to cope with drought stress. PGPR can act as biofertilizer and bioenhancer for different legumes and non-legumes. The use of PGPR and symbiotic microorganisms, may be valuable in developing strategies to assist water conservation in plants. The use of PGPR has been confirmed to be an ecologically sound way of enhancing crop yields by facilitating plant growth through direct or indirect mechanism. The mechanisms of PGPR for water conservation include secretion of exopolysaccharides, biofilm formation, alternation in phytohormone content, improvement in sugar concentration, enhancing availability of micro- and macronutrients and changes in plant functional traits. Similarly, plant growth regulators (PGRs) are specially noticed in actively growing tissues under stress conditions and have been associated in the control of cell division, embryogenesis, root formation, fruit development and ripening, and reactions to biotic and abiotic stresses and upholding water conservation status in plants. Previous studies also suggest that plant metabolites interact with plant physiology under stress condition and impart drought tolerance. Metabolites like, sugars, amino acids, organic acid and polyols play a key role in drought tolerance of crop plants grown under stress condition. It is concluded from the present study that PGRs in combination with PGPR consortium can be an effective formulation to promote plant growth and maintenance of plant turgidity under drought stress. This review is a compilation of the effect of drought stress on crop plants and described interactions between PGPR/PGRs and plant development, knowledge of water conservation and stress release strategies of PGPR and PGRs and the role of plant metabolites in drought tolerance of crop plants. This review also bridges the gaps that summarizes the mechanism of action of PGPR for drought tolerance of crop plants and sustainability of agriculture and applicability of these beneficial rhizobacteria in different agro-ecosystems under drought stress.     

Optimization of methods for peripheral blood mononuclear cells isolation and expansion of human gamma delta T cells

Human Vg9/Vδ2 T cells (γδ T cells) are immune surveillance cells both in innate and adaptive immunity and are a possible target for anticancer therapies, which can induce immune responses in a variety of cancers. Small non-peptide antigens such as zoledronate can do activation and expansion of T cells in vitro. It is evident that for adoptive cancer therapies, large numbers of functional cells are needed into cancer patients. Hence, optimization of methods needs to be carried out for the efficient expansion of these T cells. Standardization of peripheral blood mononuclear cells (PBMCs) isolation was devised. Cytokines (interleukin 2 (IL-2) and interleukin 15 (IL-15)) and zoledronate were also standardized for different concentrations. It was found that an increased number of PBMCs were recovered when washing was done at 1100 revolution per minute (rpm). Significantly high expansion fold was (2524 ± 787 expansion fold) achieved when stimulation of PBMCs was done with 1 µM of zoledronate and both cytokines IL-2 and IL-15 supported the expansion and survival of cells at the concentrations of 100 IU/ml and 10 ng/ml respectively. 14-day cultures showed highly pure (91.6 ± 5.1%) and live (96.5 ± 2.5%) expanded γδ T cells. This study aimed to standardize an easy to manipulate technique for the expansion of γδ T cells, giving a higher yield.