Effects of secondary tuber harvest on populations of devil's claw (Harpagophytum procumbens) in the Kalahari savannas of South Africa

Devil's claw ( Harpagophytum procumbens ) is an internationally traded species that is harvested for its secondary tubers. Root extracts are used to treat arthritis and other inflammatory diseases. This study examined population structure, density, growth, mortality, and seed and fruit production in harvested and unharvested populations in the Kalahari savannas of South Africa over 4 years. Plant density and population structure differed significantly between overgrazed and grass-dominated areas, suggesting that the differences may be due to competition for water and nutrients. Experimental removal of secondary tubers (harvest) was not a significant factor for mortality in any of the harvested size classes. Harvest also did not affect growth, although plants in the 3–4 cm size class grew more in both the harvested and unharvested populations. Fruit production was variable; fruits matured only after sufficient rains. Under the conditions of this study, the species appears to be resilient to harvest, with both harvested and unharvested plants surviving. After harvest, both groups recovered and grew (on average) at the same rate. Because of the spatially variable habitat and the plasticity of the plants themselves, a large number of plants over a wide area are required to better understand the species' life history.     

Strategies for Enhancing the Effectiveness of Metagenomic-based Enzyme Discovery in Lignocellulolytic Microbial Communities

Producing cellulosic biofuels from plant material has recently emerged as a key US Department of Energy goal. For this technology to be commercially viable on a large scale, it is critical to make production cost efficient by streamlining both the deconstruction of lignocellulosic biomass and fuel production. Many natural ecosystems efficiently degrade lignocellulosic biomass and harbor enzymes that, when identified, could be used to increase the efficiency of commercial biomass deconstruction. However, ecosystems most likely to yield relevant enzymes, such as tropical rain forest soil in Puerto Rico, are often too complex for enzyme discovery using current metagenomic sequencing technologies. One potential strategy to overcome this problem is to selectively cultivate the microbial communities from these complex ecosystems on biomass under defined conditions, generating less complex biomass-degrading microbial populations. To test this premise, we cultivated microbes from Puerto Rican soil or green waste compost under precisely defined conditions in the presence dried ground switchgrass (Panicum virgatum L.) or lignin, respectively, as the sole carbon source. Phylogenetic profiling of the two feedstock-adapted communities using SSU rRNA gene amplicon pyrosequencing or phylogenetic microarray analysis revealed that the adapted communities were significantly simplified compared to the natural communities from which they were derived. Several members of the lignin-adapted and switchgrass-adapted consortia are related to organisms previously characterized as biomass degraders, while others were from less well-characterized phyla. The decrease in complexity of these communities make them good candidates for metagenomic sequencing and will likely enable the reconstruction of a greater number of full-length genes, leading to the discovery of novel lignocellulose-degrading enzymes adapted to feedstocks and conditions of interest.     

Mutations in myosin light chain kinase cause familial aortic dissections

Mutations in smooth muscle cell (SMC)-specific isoforms of α-actin and β-myosin heavy chain, two major components of the SMC contractile unit, cause familial thoracic aortic aneurysms leading to acute aortic dissections (FTAAD). To investigate whether mutations in the kinase that controls SMC contractile function (myosin light chain kinase [MYLK]) cause FTAAD, we sequenced MYLK by using DNA from 193 affected probands from unrelated FTAAD families. One nonsense and four missense variants were identified in MYLK and were not present in matched controls. Two variants, p.R1480X (c.4438C>T) and p.S1759P (c.5275T>C), segregated with aortic dissections in two families with a maximum LOD score of 2.1, providing evidence of linkage of these rare variants to the disease (p = 0.0009). Both families demonstrated a similar phenotype characterized by presentation with an acute aortic dissection with little to no enlargement of the aorta. The p.R1480X mutation leads to a truncated protein lacking the kinase and calmodulin binding domains, and p.S1759P alters amino acids in the α-helix of the calmodulin binding sequence, which disrupts kinase binding to calmodulin and reduces kinase activity in vitro. Furthermore, mice with SMC-specific knockdown of Mylk demonstrate altered gene expression and pathology consistent with medial degeneration of the aorta. Thus, genetic and functional studies support the conclusion that heterozygous loss-of-function mutations in MYLK are associated with aortic dissections.     

A novel flow cytometric assay for measurement of In Vivo pulmonary neutrophil phagocytosis

Background  

Phagocytosis assays are traditionally performed in vitro using polymorphonuclear leukocytes (PMNs) isolated from peripheral blood or the peritoneum and heat-killed, pre-opsonized organisms. These assays may not adequately mimic the environment within the infected lung. Our laboratory therefore has developed a flow cytometric in vivo phagocytosis assay that enables quantification of PMN phagocytosis of viable bacteria within the lungs of rats. In these studies, rats are injected transtracheally with lipopolysaccharide (LPS) to recruit PMNs to their lungs. They are then infected with live 5(-and 6) carboxyfluorescein diacetate succinimidyl ester (CFDA/SE) labeled type 3 Streptococcus pneumoniae. Bronchoalveolar lavage is performed and resident alveolar macrophages and recruited PMNs are labeled with monoclonal antibodies specific for surface epitopes on each cell type. Three color flow cytometry is utilized to identify the cell types, quantify recruitment, and determine uptake of the labeled bacteria.     

A novel approach to identifying beta-secretase inhibitors: bis-statine peptide mimetics discovered using structure and spot synthesis

Beta-secretase 1 (BACE1) is an aspartic protease believed to play a critical role in Alzheimer's disease. Inhibitors of this enzyme have been designed by incorporating the non-cleavable hydroxyethylene and statine isosteres into peptides corresponding to BACE1 substrate sequences. We sought to develop new methods to quickly characterize and optimize inhibitors based on the statine core. Minimal sequence requirements for binding were first established using both crystallography and peptide spot synthesis. These shortened peptide inhibitors were then optimized by using spot synthesis to perform iterative cycles of substitution and deletion. The present study resulted in the identification of novel "bis-statine" inhibitors shown by crystallography to have a unique binding mode. Our results demonstrate the application of peptide spot synthesis as an effective method for enhancing peptidomimetic drug discovery.     

Semi-automated segmentation and visualisation of outer bone cortex from medical images

Good segmentation of the outer bone cortex from medical images is a prerequisite for applications in the field of finite element analysis, surgical planning environments and personalised, case dependent, bone reconstruction. However, current segmentation procedures are often unsatisfactory. This study presents an automated filter procedure to generate a set of adapted contours from which a surface mesh can be deduced directly. The degree of interaction is user determined. The bone contours are extracted from the patients CT data by quick grey value segmentation. An extended filter procedure then only retains contour information representing the outer cortex as more specific internal loops and shape irregularities are removed, tailoring the image for the above-mentioned applications. The developed medical image based design methodology to convert contour sets of multiple bone types, from tibia tumour to neurocranium, is reported and discussed.     

The resonance frequencies and mode shapes of dental implants: Rigid body behaviour versus bending behaviour. A numerical approach

The purpose of this study was to evaluate the modal behaviour of the bone-implant-transducer (Osstell) system by means of finite element analyses. The influence of different parameters was determined: (1) the type of implant anchorage being trabecular, cortical, uni-cortical, or bi-cortical, (2) the implant diameter, (3) the length of the implant embedded in the bone, and (4) the bone stiffness. The type of anchorage determines the resulting modal behaviour of the implant-transducer system. A rigid body behaviour was found for a uni-cortical anchoring and for a homogeneous anchoring with low bone stiffness (< or =1000 MPa), whereas a bending behaviour was found for a homogeneous anchoring with a high bone stiffness (> or =5000 MPa) and for a bi-cortical anchorage. The implant dimensions influence the values for the resonance frequencies. Generally, an increase in implant diameter or implant length (in bone) results in higher resonance frequencies. This study also showed that resonance frequencies in case of rigid body behaviour of the implant-transducer system are more sensitive to changes in bone stiffness than resonance frequencies in case of bending behaviour. In conclusion, it seems that the Osstell transducer is suited for the follow-up in time of the stability of an implant, but not for the quantitative comparison of the stability of implants.