Selection for High Oridonin Yield in the Chinese Medicinal Plant Isodon (Lamiaceae) Using a Combined Phylogenetics and Population Genetics Approach

Oridonin is a diterpenoid with anti-cancer activity that occurs in the Chinese medicinal plant Isodon rubescens and some related species. While the bioactivity of oridonin has been well studied, the extent of natural variation in the production of this compound is poorly known. This study characterizes natural variation in oridonin production in order to guide selection of populations of Isodon with highest oridonin yield. Different populations of I. rubescens and related species were collected in China, and their offspring were grown in a greenhouse. Samples were examined for oridonin content, genotyped using 11 microsatellites, and representatives were sequenced for three phylogenetic markers (ITS, rps16, trnL-trnF). Oridonin production was mapped on a molecular phylogeny of the genus Isodon using samples from each population as well as previously published Genbank sequences. Oridonin has been reported in 12 out of 74 species of Isodon examined for diterpenoids, and the phylogeny indicates that oridonin production has arisen at least three times in the genus. Oridonin production was surprisingly consistent between wild-collected parents and greenhouse-grown offspring, despite evidence of gene flow between oridonin-producing and non-producing populations of Isodon. Additionally, microsatellite genetic distance between individuals was significantly correlated with chemical distance in both parents and offspring. Neither heritability nor correlation with genetic distance were significant when the comparison was restricted to only populations of I. rubescens, but this result should be corroborated using additional samples. Based on these results, future screening of Isodon populations for oridonin yield should initially prioritize a broad survey of all species known to produce oridonin, rather than focusing on multiple populations of one species, such as I. rubescens. Of the samples examined here, I. rubescens or I. japonicus from Henan province would provide the best source of oridonin.     

The effectiveness of constructed wetland for treatment of woodwaste leachate

Percolation of rainfall through woodwaste piles leaches natural chemicals from the wood residuals that can have adverse impacts on the environment. A study was conducted on a woodwaste storage site, adjacent to the Lower Fraser River, near Mission, BC, Canada. The objective of this research was to evaluate the effectiveness of constructed wetland for treatment of this woodwaste leachate. The leachate was characterized by high oxygen demand, tannin and lignin, and volatile fatty acids (VFAs), but low pH and nutrients. Diluted leachate passed through six pilot-scale wetland cells, four planted with cattail (Typha latifolia) and two unplanted controls, with a hydraulic retention time of 7 days and an average depth of 40 cm. Nutrient addition and pH adjustments were made to improve contaminant removal. Reductions in contaminants were consistently achieved, with average removals for BOD, COD, VFAs and tannin and lignin of 60, 50, 69 and 42%, respectively. Climatic conditions had an impact on the performance of the constructed wetland. Further operation of the system will help to elucidate the seasonal fluctuations. Aging of the constructed wetland system increased the treatment performance.     

Chronic exposure to inhaled anesthetics increases cholesterol content in Acholeplasma laidlawii

Acholeplasma laidlawii cells were grown in cholesterol-enriched medium and exposed continuously to either air (control), 4.0 vol.% halothane in air at 1 atm pressure (4% atm halothane), or 80% cyclopropane in oxygen for 24 h at 37°C. Cells grown in the presence of 4% atm halothane or 80% cyclopropane had approximately twice as much membrane cholesterol content/mg protein as the control cells. Cells grown in an anesthetic environment also tended to have a higher membrane cholesterol/phospholipid molar ratio compared to control cells. Membranes isolated from halothane-exposed cells grown in a cholesterol-enriched medium were more ordered at 37°C (measurements were made with no anesthetic present) than membranes from control cells grown in an identically enriched medium. This difference in membrane physical state between control and anesthetic-exposed cells decreased as the temperature decreased, and disappeared at approx. 23°C. Continuous exposure of A. laidlawii to 4% atm halothane or 80% cyclopropane for 24 h did not markedly affect membrane fatty acid composition, either in cells grown on an unsupplemented medium or in cells grown in a medium enriched in myristic, palmitic or stearic acids. These results further support the hypothesis that an increased membrane cholesterol content may play a role in the tolerance or dependence that develops after chronic exposure to anesthetic agents.     

A landscape of shifting-mosaic steady state in Lassen Volcanic National Park,California

It is generally perceived that landscape patterns or textures in a given protected area are spatially stationary. The findings of this study suggest that this common perception is only partially correct. Over the course of 52 years, equilibrium in landscape shifting was detected using digital data for the Lassen Volcanic National Park (USA). Vertical aerial photographs taken of the park in 1941 were geo-referenced with the digital orthophoto quarter quadrangle (DOQQ) images of the same area from 1993 to identify landscape compositions and to measure change. Spatial analysis was used to observe pattern changes over time. The results suggested that landscape development maintained equilibrium while patches were in various stages of a successional sequence. The total area of each landscape component held steady, although over time patches throughout the landscape changed—a shifting-mosaic steady state (SMSS). These findings reflect the limitations of contemporary environmental conservation theory. They also suggest the importance of considering landscape change in policies that currently govern park planning and management.     

Kinetic Basis for the Conjugation of Auxin by a GH3 Family Indole-acetic Acid-Amido Synthetase

The GH3 family of acyl-acid-amido synthetases catalyze the ATP-dependent formation of amino acid conjugates to modulate levels of active plant hormones, including auxins and jasmonates. Initial biochemical studies of various GH3s show that these enzymes group into three families based on sequence relationships and acyl-acid substrate preference (I, jasmonate-conjugating; II, auxin- and salicylic acid-conjugating; III, benzoate-conjugating); however, little is known about the kinetic and chemical mechanisms of these enzymes. Here we use GH3-8 from Oryza sativa (rice; OsGH3-8), which functions as an indole-acetic acid (IAA)-amido synthetase, for detailed mechanistic studies. Steady-state kinetic analysis shows that the OsGH3-8 requires either Mg²⁺ or Mn²⁺ for maximal activity and is specific for aspartate but accepts asparagine as a substrate with a 45-fold decrease in catalytic efficiency and accepts other auxin analogs, including phenyl-acetic acid, indole butyric acid, and naphthalene-acetic acid, as acyl-acid substrates with 1.4–9-fold reductions in k_cat/K_m relative to IAA. Initial velocity and product inhibition studies indicate that the enzyme uses a Bi Uni Uni Bi Ping Pong reaction sequence. In the first half-reaction, ATP binds first followed by IAA. Next, formation of an adenylated IAA intermediate results in release of pyrophosphate. The second half-reaction begins with binding of aspartate, which reacts with the adenylated intermediate to release IAA-Asp and AMP. Formation of a catalytically competent adenylated-IAA reaction intermediate was confirmed by mass spectrometry. These mechanistic studies provide insight on the reaction catalyzed by the GH3 family of enzymes to modulate plant hormone action.     

Factors associated with whole carcass condemnation rates in provincially-inspected abattoirs in Ontario 2001-2007: implications for food animal syndromic surveillance

Background  

Ontario provincial abattoirs have the potential to be important sources of syndromic surveillance data for emerging diseases of concern to animal health, public health and food safety. The objectives of this study were to: (1) describe provincially inspected abattoirs processing cattle in Ontario in terms of the number of abattoirs, the number of weeks abattoirs process cattle, geographical distribution, types of whole carcass condemnations reported, and the distance animals are shipped for slaughter; and (2) identify various seasonal, secular, disease and non-disease factors that might bias the results of quantitative methods, such as cluster detection methods, used for food animal syndromic surveillance.