The Biosynthetic Origin of Irregular Monoterpenes in Lavandula: ISOLATION AND BIOCHEMICAL CHARACTERIZATION OF A NOVEL cis-PRENYL DIPHOSPHATE SYNTHASE GENE,LAVANDULYL DIPHOSPHATE SYNTHASE*

Lavender essential oils are constituted predominantly of regular monoterpenes, for example linalool, 1,8-cineole, and camphor. However, they also contain irregular monoterpenes including lavandulol and lavandulyl acetate. Although the majority of genes responsible for the production of regular monoterpenes in lavenders are now known, enzymes (including lavandulyl diphosphate synthase (LPPS)) catalyzing the biosynthesis of irregular monoterpenes in these plants have not been described. Here, we report the isolation and functional characterization of a novel cis-prenyl diphosphate synthase cDNA, termed Lavandula x intermedia lavandulyl diphosphate synthase (LiLPPS), through a homology-based cloning strategy. The LiLPPS ORF, encoding for a 305-amino acid long protein, was expressed in Escherichia coli, and the recombinant protein was purified by nickel-nitrilotriacetic acid affinity chromatography. The approximately 34.5-kDa bacterially produced protein specifically catalyzed the head-to-middle condensation of two dimethylallyl diphosphate units to LPP in vitro with apparent K_m and k_cat values of 208 ± 12 μm and 0.1 s⁻¹, respectively. LiLPPS is a homodimeric enzyme with a sigmoidal saturation curve and Hill coefficient of 2.7, suggesting a positive co-operative interaction among its catalytic sites. LiLPPS could be used to modulate the production of lavandulol and its derivatives in plants through metabolic engineering.     

Responses of grape berry anthocyanin and titratable acidity to the projected climate change across the Western Australian wine regions

More than a century of observations has established that climate influences grape berry composition. Accordingly, the projected global climate change is expected to impact on grape berry composition although the magnitude and direction of impact at regional and subregional scales are not fully known. The aim of this study was to assess potential impacts of climate change on levels of berry anthocyanin and titratable acidity (TA) of the major grapevine varieties grown across all of the Western Australian (WA) wine regions. Grape berry anthocyanin and TA responses across all WA wine regions were projected for 2030, 2050 and 2070 by utilising empirical models that link these berry attributes and climate data downscaled (to ～5 km resolution) from the csiro_mk3_5 and miroc3_2_medres global climate model outputs under IPCC SRES A2 emissions scenario. Due to the dependence of berry composition on maturity, climate impacts on anthocyanin and TA levels were assessed at a common maturity of 22 °Brix total soluble solids (TSS), which necessitated the determination of when this maturity will be reached for each variety, region and warming scenario, and future period. The results indicate that both anthocyanin and TA levels will be affected negatively by a warming climate, but the magnitude of the impacts will differ between varieties and wine regions. Compared to 1990 levels, median anthocyanins concentrations are projected to decrease, depending on global climate model, by up to 3–12 % and 9–33 % for the northern wine regions by 2030 and 2070, respectively while 2–18 % reductions are projected in the southern wine regions for the same time periods. Patterns of reductions in the median Shiraz berry anthocyanin concentrations are similar to that of Cabernet Sauvignon; however, the magnitude is lower (up to 9–18 % in southern and northern wine regions respectively by 2070). Similarly, uneven declines in TA levels are projected across the study regions. The largest reductions in median TA are likely to occur in the present day warmer wine regions, up to 40 % for Chardonnay followed by 15 % and 12 % for Shiraz and Cabernet Sauvignon, respectively, by 2070 under the high warming projection (csiro_mk3_5). It is concluded that, under existing management practices, some of the key grape attributes that are integral to premium wine production will be affected negatively by a warming climate, but the magnitudes of the impacts vary across the established wine regions, varieties, the magnitude of warming and future periods considered.     

Phylogenetic characterisation of Taenia tapeworms in spotted hyenas and reconsideration of the “Out of Africa” hypothesis of Taenia in humans

The African origin of hominins suggests that Taenia spp. in African carnivores are evolutionarily related to the human-infecting tapeworms Taenia solium, Taenia saginata and Taenia asiatica. Nevertheless, the hypothesis has not been verified through molecular phylogenetics of Taenia. This study aimed to perform phylogenetic comparisons between Taenia spp. from African hyenas and the congeneric human parasites. During 2010–2013, 233 adult specimens of Taenia spp. were collected from 11 spotted hyenas in Ethiopia. A screening based on short DNA sequences of the cytochrome c oxidase subunit 1 gene classified the samples into four mitochondrial lineages designated as I–IV. DNA profiles of nuclear genes for DNA polymerase delta (pold) and phosphoenolpyruvate carboxykinase (pepck) showed that lineages II and III can be assigned as two independent species. Common haplotypes of pold and pepck were frequently found in lineages I and IV, suggesting that they constitute a single species. Morphological observations suggested that lineage II is Taenia crocutae, but the other lineages were morphologically inconsistent with known species, suggesting the involvement of two new species. A phylogenetic tree of Taenia spp. was reconstructed by the maximum likelihood method using all protein-coding genes of their mitochondrial genomes. The tree clearly demonstrated that T. crocutae is sister to T. saginata and T. asiatica, whereas T. solium was confirmed to be sister to the brown bear tapeworm, Taenia arctos. The tree also suggested that T. solium and T. arctos are related to two species of Taenia in hyenas, corresponding to lineages I + IV and III. These results may partially support the African origin of human-infecting Taenia spp., but there remains a possibility that host switching of Taenia to hominins was not confined to Africa. Additional taxa from African carnivores are needed for further testing of the “Out of Africa” hypothesis of Taenia in humans.     

A systems approach to improving rural care in Ethiopia

Background

Multiple interventions have been launched to improve the quality, access, and utilization of primary health care in rural, low-income settings; however, the success of these interventions varies substantially, even within single studies where the measured impact of interventions differs across sites, centers, and regions. Accordingly, we sought to examine the variation in impact of a health systems strengthening intervention and understand factors that might explain the variation in impact across primary health care units.

Methodology/Principal Findings

We conducted a mixed methods positive deviance study of 20 Primary Health Care Units (PHCUs) in rural Ethiopia. Using longitudinal data from the Ethiopia Millennium Rural Initiative (EMRI), we identified PHCUs with consistently higher performance (n = 2), most improved performance (n = 3), or consistently lower performance (n = 2) in the provision of antenatal care, HIV testing in antenatal care, and skilled birth attendance rates. Using data from site visits and in-depth interviews (n = 51), we applied the constant comparative method of qualitative data analysis to identify key themes that distinguished PHCUs with different performance trajectories. Key themes that distinguished PHCUs were 1) managerial problem solving capacity, 2) relationship with the woreda (district) health office, and 3) community engagement. In higher performing PHCUs and those with the greatest improvement after the EMRI intervention, health center and health post staff were more able to solve day-to-day problems, staff had better relationships with the woreda health official, and PHCU communities'' leadership, particularly religious leadership, were strongly engaged with the health improvement effort. Distance from the nearest city, quality of roads and transportation, and cultural norms did not differ substantially among PHCUs.

Conclusions/Significance

Effective health strengthening efforts may require intensive development of managerial problem solving skills, strong relationships with government offices that oversee front-line providers, and committed community leadership to succeed.     

Caspase-3-truncated type 1 inositol 1,4,5-trisphosphate receptor enhances intracellular Ca2+ leak and disturbs Ca2+ signalling.

BACKGROUND INFORMATION: The IP(3)R (inositol 1,4,5-trisphosphate receptor) is a tetrameric channel that accounts for a large part of the intracellular Ca(2+) release in virtually all cell types. We have previously demonstrated that caspase-3-mediated cleavage of IP(3)R1 during cell death generates a C-terminal fragment of 95 kDa comprising the complete channel domain. Expression of this truncated IP(3)R increases the cellular sensitivity to apoptotic stimuli, and it was postulated to be a constitutively active channel. RESULTS: In the present study, we demonstrate that expression of the caspase-3-cleaved C-terminus of IP(3)R1 increased the rate of thapsigargin-mediated Ca(2+) leak and decreased the rate of Ca(2+) uptake into the ER (endoplasmic reticulum), although it was not sufficient by itself to deplete intracellular Ca(2+) stores. We detected the truncated IP(3)R1 in different cell types after a challenge with apoptotic stimuli, as well as in aged mouse oocytes. Injection of mRNA corresponding to the truncated IP(3)R1 blocked sperm factor-induced Ca(2+) oscillations and induced an apoptotic phenotype. CONCLUSIONS: In the present study, we show that caspase-3-mediated truncation of IP(3)R1 enhanced the Ca(2+) leak from the ER. We suggest a model in which, in normal conditions, the increased Ca(2+) leak is largely compensated by enhanced Ca(2+)-uptake activity, whereas in situations where the cellular metabolism is compromised, as occurring in aging oocytes, the Ca(2+) leak acts as a feed-forward mechanism to divert the cell into apoptosis.     

Field-based investigation on phytoremediation potentials of Lemna minor and Azolla filiculoides in tropical,semiarid regions: Case of Ethiopia

This study investigated the concurrent accumulation of eight heavy metals by two floating aquatic macrophytes (Lemna minor and Azolla filiculoides) cultivated in ambient media and blended wastewaters in the semiarid regions of Ethiopia. Both species accumulated heavy metals in varying degrees with a significant concentration gradient within the immediate water media. Highest bioconcentration factor (BCF) was determined for Mn and Fe in both plants. Results revealed that L. minor was high phytoaccumulator for Fe, Mn, Zn, and Co but moderate for Cd, Cu, Ni, and Cr. On the other hand, A. filiculoides was a high accumulator for Fe, Mn, Zn, and Cu, but its potency was moderate for Co, Cr, and Ni, but lower for Cd. Both species exhibited significant difference in accumulating Co, Zn, and Mn (p < 0.05). In general, the BCFs for both plants were comparable within the same treatment. In this study, stronger associations between the heavy metal concentrations in the plant tissues and in the grown water media were observed for A. filiculoides.     

Post-fire regeneration strategies and tree bark resistance to heating in frequently burning tropical savanna woodlands and grasslands in Ethiopia

Regeneration mechanisms of vegetation and the role of tree bark resistance to frequent fire were studied in savanna woodlands and grasslands in Gambella, Western Ethiopia. Data were collected from four sites, each with three replicate plots. The variation between sites in species composition and biomass correlated with the differences in fire intensity. Foliar cover was recorded for individual plant species regenerating by sprouting from older parts of plants that had survived fire or by seedlings; records were made during the dry season and at the beginning of the wet season. Data on bark thickness and tree diameters of 12 dominant tree species were also recorded. Both facultative and obligate sprouters significantly contributed to post‐fire recovery, comprising 98.5 % of total vegetation cover. The contribution of seedlings to cover and abundance immediately following fire was negligible, but seedling density increased in the beginning of the rainy season, 4 to 5 months after fire. The importance of the sprouting and seeding strategies varied between the different plant growth forms. The highest contribution to cover and frequency was made by the most abundant grass species, which reproduced in both ways. Facultative sprouters made up 67.3 % of the vegetation cover, out of which 54 % consisted of grasses. Broad‐leaved herbs and trees/shrubs regenerating mainly by sprouting made up 31.3 % of the vegetation cover. Adaptations to fire in tree species seemed to include the development of a thick bark, once the tree has passed seedling stages. Tree bark thickness and tree diameter at breast height were strongly correlated with the time taken for cambium to reach an assumed lethal temperature of 60°C when exposed to fire, which indicated that mature trees with thick barks might resist stronger fire better than, e.g., small or young trees and trees with thin bark. However, for a given bark thickness the cambium resistance to heat varied three‐fold among species. Hence, site differences in fire intensity seemed to influence the distribution of trees depending on their bark characteristics and resistance to fire.     

Production of macrolide antibiotics from a cytotoxic soil <Emphasis Type="Italic">Streptomyces</Emphasis> sp. strain ZDB

Crude extract from a culture of a soil Streptomyces sp. strain ZDB showed toxicity towards Artemia salina and antimicrobial activity against Escherichia coli, Bacillus subtilis, Staphylococcus aureus, Chlorella vulgaris, and Chlorella sorokiniana. Large scale fermentation of the strain led to the isolation of the macrolide antibiotics, bafilomycins A1 (1), B1 (2), and D (3) together with nonactic acid (4) and bostrycoidin-9-methyl ether (5). Structures of the antibiotics were determined based on spectral data analysis. We describe the isolation of the compounds and characterization of the producing strain.