Engineering levoglucosan metabolic pathway in <Emphasis Type="Italic">Rhodococcus jostii</Emphasis> RHA1 for lipid production

Oleaginous strains of Rhodococcus including R. jostii RHA1 have attracted considerable attention due to their ability to accumulate triacylglycerols (TAGs), robust growth properties and genetic tractability. In this study, a novel metabolic pathway was introduced into R. jostii by heterogenous expression of the well-characterized gene, lgk encoding levoglucosan kinase from Lipomyces starkeyi YZ-215. This enables the recombinant R. jostii RHA1 to produce TAGs from the anhydrous sugar, levoglucosan, which can be generated efficiently as the major molecule from the pyrolysis of cellulose. The recombinant R. jostii RHA1 could grow on levoglucosan as the sole carbon source, and the consumption rate of levoglucosan was determined. Furthermore, expression of one more copy of lgk increased the enzymatic activity of LGK in the recombinant. However, the growth performance of the recombinant bearing two copies of lgk on levoglucosan was not improved. Although expression of lgk in the recombinants was not repressed by the glucose present in the media, glucose in the sugar mixture still affected consumption of levoglucosan. Under nitrogen limiting conditions, lipid produced from levoglucosan by the recombinant bearing lgk was up to 43.54 % of the cell dry weight, which was comparable to the content of lipid accumulated from glucose. This work demonstrated the technical feasibility of producing lipid from levoglucosan, an anhydrosugar derived from the pyrolysis of lignocellulosic materials, by the genetically modified rhodococci strains.     

Glycosylphosphatidylinositol-induced cardiac myocyte death might contribute to the fatal outcome of Plasmodium falciparum malaria

Background  Glycosylphosphatidylinositol (GPI) purified from Plasmodium falciparum has been shown to play an important role as a toxin in the pathology of malaria. Previous studies demonstrated cardiac involvement in patients suffering from severe malaria due to P. falciparum. Therefore, we tested the hypothesis that GPI induces apoptosis in cardiomyocytes. Methods and results  By using TUNEL and caspase activity assays, we provided evidence for apoptosis induction in cardiomyocytes by P. falciparum GPI after 48 h of incubation. A similar result was obtained in heart cells of mice 48 h after in vivo injection of GPI. Gene expression analyses in GPI-treated cardiomyocytes showed an up-regulation of apoptotic genes (apaf-1, bax) and of a myocardial damage marker bnp (brain natriuretic peptide), while a down-regulation was observed for the anti-apoptotic gene bcl-2 and for the heat shock protein hsp70. In spite of inflammatory cytokine gene up-regulation by GPI, co-culture with peripheral mononuclear cells (PMNCs) did not change the results obtained with cardiomyocytes alone, indicating a direct effect of GPI on cardiac myocytes. Co-culture with non-myocytic cardiac cells (NMCCs) resulted in up-regulation of Hsp70 and Bcl-2 genes in GPI-treated cardiomyocytes but without repercussion on the apoptosis level. A malaria-infected patient, presenting fulminant heart failure showed typical signs of cardiac myocyte apoptosis demonstrating the clinical relevance of toxin induced heart damage for the lethality of malaria. Our studies performed in vitro and in mice suggest that the GPI could be responsible for cardiomyocyte apoptosis that occurred in this patient. Conclusion   Plasmodium falciparum GPI-induced apoptosis might participate in the lethality of malaria.

Predictive performance modelling of parallel component compositions

Large-scale scientific computing applications frequently make use of closely-coupled distributed parallel components. The performance of such applications is therefore dependent on the component parts and their interaction at run-time. This paper describes a methodology for predictive performance modelling and evaluation of parallel applications composed of multiple interacting components. In this paper, the fundamental steps and required operations involved in the modelling and evaluation process are identified—including component decomposition, component model combination, M×N communication modelling, dataflow analysis and overall performance evaluation. A case study is presented to illustrate the modelling process and the methodology is verified through experimental analysis.

Purification and properties of Sandercyanin, a blue protein secreted in the mucus of blue forms of walleye, Sander vitreus

A blue protein present in the mucus coating blue forms of walleye, Sander vitreus, was purified to homogeneity by a combination of ion exchange and hydrophobic interaction chromatography. The purified protein has a molecular mass of 87,850 and is a homotetramer with a subunit molecular mass of 21,836. Solutions of the protein are deep blue in color and show absorbance maxima at 383 and 633 nm, respectively. Acetone treatment of the protein releases a blue chromophore with the spectral characteristics of biliverdin IXα. The results suggest that the blue protein is a new biliprotein that we have termed Sandercyanin. The function of Sandercyanin is unknown, but limited amino acid sequence analysis suggests it is a lipocalin that may be involved in the transport of heme degradation products.

Two new species of <Emphasis Type="Italic">Mycena</Emphasis> from alpine sites in Norway

Mycena aphanes and Mycena exilis, belonging to section Filipedes and section Polyadelphia, are proposed as new species. They are described and illustrated and compared to other species in the two sections. Mycena aphanes is a strikingly characteristic Mycena with fulvous or yellowish brown colours of both pileus and stipe. Mycena exilis is characterized by its occurrence on fallen Salix leaves, a pale brown pileus, occasionally with a pale pink tinge, narrowly adnate lamellae, four-spored basidia, and the presence of clamp connections.

<Emphasis Type="Italic">Berlinia korupensis</Emphasis> (Leguminosae <Emphasis Type="Italic">–</Emphasis> Caesalpinioideae), a new tree species from Cameroon

Summary  The tree species Berlinia korupensis Mackinder & Burgt is described as new. The species is endemic to the southern part of Korup National Park in Cameroon. Seventeen trees have been found so far, the largest being 42 m high and having a trunk diam. of 88 cm. The new species is assessed as Critically Endangered (CR D) under the criteria of IUCN. Two distribution maps are included; one map indicating the single locality in Cameroon near the Nigerian border where the new species was found and another map of the permanent plot where 14 of the 17 trees were recorded. A line drawing is also included, along with colour photographs of the flowers and the trunk.

A single operon-encoded form of the acetyl-CoA decarbonylase/synthase multienzyme complex responsible for synthesis and cleavage of acetyl-CoA in <Emphasis Type="Italic">Methanosarcina thermophila</Emphasis>

Methanogens growing on C-1 substrates synthesize 2-carbon acetyl groups in the form of acetyl-CoA for carbon assimilation using the multienzyme complex acetyl-CoA decarbonylase/synthase (ACDS) which contains five different subunits encoded within an operon. In species growing on acetate ACDS also functions to cleave the acetate C-C bond for energy production by methanogenesis. A number of species of Methanosarcina that are capable of growth on either C-1 compounds or acetate contain two separate ACDS operons, and questions have been raised about whether or not these operons play separate roles in acetate synthesis and cleavage. Methanosarcina thermophila genomic DNA was analyzed for the presence of two ACDS operons by PCR amplifications with different primer pairs, restriction enzyme analyses, DNA sequencing and Southern blot analyses. A single ACDS operon was identified and characterized, with no evidence for more than one. MALDI mass spectrometric analyses were carried out on ACDS preparations from methanol- and acetate-grown cells. Peptide fragmentation patterns showed that the same ACDS subunits were present regardless of growth conditions. The evidence indicates that a single form of ACDS is used both for acetate cleavage during growth on acetate and for acetate synthesis during growth on C-1 substrates. Electronic Supplementary Material  Supplementary material is available for this article at

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Salal Harvester Local Ecological Knowledge, Harvest Practices and Understory Management on the Olympic Peninsula, Washington

Despite growing interest in traditional and local ecological knowledge for conservation and resource management, the role of migrant resource users is largely unexplored. Challenging many assumptions about what constitutes “local knowledge,” migrant and immigrant harvesters of non-timber forest products on the Olympic Peninsula, Washington possess useful ecological knowledge of overstory–understory relationships and how forestry practices affect understory biological and commercial production. Harvesters of salal (Gaultheria shallon), a shrub used in the multi-million dollar floral greens industry, were interviewed in Mason County, Washington in 2001–2003. Interviews revealed that harvesters possess different kinds of resource management knowledge depending on whether they are experienced harvesters or more recent newcomers to the area. These differences may also correlate with differences in their harvesting practices. Understanding how resource management knowledge differs between experienced and newcomer harvesters can inform forest managers in their efforts to develop effective management and permitting policies for floral greens and other non-timber forest resources in the Pacific Northwest of the United States.

Aggregations of juvenile whale sharks (<Emphasis Type="Italic">Rhincodon typus</Emphasis>) in the Gulf of Tadjoura,Djibouti

A total of 23 whale sharks were identified over a 5 d period in the Arta Bay region of the Gulf of Tadjora, Djibouti. Most of the sharks aggregating in this area were small (<4 m TL) males. Individuals were identified using photographs of distinctive scars and spot and stripe patterns on the sides of the animals. Of these, 65% had scarring that was attributable to boat or propeller strikes. Most of the whale sharks we encountered were feeding on dense accumulations of plankton in shallow water just off (10–200 m) the shoreline. This food source may account for the aggregation of sharks in this area. One 3 m male shark was tagged with an ARGOS (Splash) satellite tag for 9 d. During this time the shark traversed to the shoreline on the opposite side of the Gulf (a distance of 14 km) and then returned to the Arta Bay area before retracing his path to the other shore. The shark spent most of the daylight hours at the surface, while at night dives were more frequent, deeper and for longer durations.

Genetic variability in relocated Père David’s deer (<Emphasis Type="Italic">Elaphurus davidianus</Emphasis>) populations—Implications to reintroduction program

Since 1985, China has established three breeding herds of Père David’s deer: the Beijing Père David’s Deer Park (39°07′N, 116°03′E), the Dafeng Père David’s Deer Nature Reserve (33°05′N, 120°49′E) and Shishou (Tianezhou) Père David’s Deer Nature Reserve (29°49′N, 112°33′E), through reintroductions of about 30–40 founders. Since establishment, all three populations have grown steadily. However, genetic backgrounds in those populations are still unknown. We studied the genetic diversity in Père David’s deer and genetic consequences of population relocations in China. We revealed that genetic diversity was extremely low in Père David’s deer populations in China. Only a single mtDNA D-loop haplotype was found in the deer, furthermore, only five polymorphic microsatellite loci were screened out from 84 pairs of species-transferred primers. Genetic makeup in the three Père David’s deer populations were significantly different (P < 0.01). H _E and allelic richness in the Tianezhou population were the highest (0.54, 2.60, n = 31), Beijing population (0.52, 2.4, n = 125) showed the second highest measures, while the Dafeng population (0.46, 2.39, n = 39) measured lowest. Our results suggest that effective management of a species of low genetic diversity like the Père David’s deer should consider the genetic background of each founder to make sure genetic variations are preserved in both source population and relocated population. Now, the Tianezhou population is the most appropriate source population in China when establishing new Père David deer populations in the wild.

Transfection of insect cell lines using polyethylenimine

Insect cell lines have been widely used in recombinant baculovirus expression systems and transient gene expression studies. Critical to these applications have been the transfection of foreign DNA. This has been frequently done using labor intensive and cytotoxic liposome-based transfection reagents. In the current study we have optimized a new kind of polyethylenimine-based DNA transfection reagent on the Spodoptera frugiperda Sf9 insect cell line. A plasmid vector that transiently expresses green fluorescent protein (GFP) was effectively delivered into Sf9 cells. A transfection efficiency of 54% and cell viability of 85–90% were obtained for Sf9 cells. The developed transfection protocol has now been successfully used to transfect eight insect cell lines derived from Bombyx mori, Trichoplusia ni, Helicoverpa zea, Heliothis virescens and S. frugiperda with GFP and GUS with transfection efficiencies of at least 45%. This method provides high heterologous protein expression levels, transfection efficacy and cell viability, and could be used for transient gene expression in other lepidopteran cell lines.

Influence of spinosad on immature and adult stages of <Emphasis Type="Italic">Chrysoperla carnea</Emphasis> (Stephens) (Neuroptera: Chrysopidae)

Toxicity of spinosad to immature stages of Chrysoperla carnea (Stephens) (Neuroptera: Chrysopidae) and its effect on the reproduction and survival of adult stages after direct spray and ingestion treatments were evaluated. Spinosad was harmless to C. carnea eggs and pupae irrespective of concentrations or method of treatments. Direct spray of spinosad to first instar caused significant reduction in rate of pupation and L1-adult survival, but did not affect the rate of adult emergence. Third-instar bioassay revealed significant difference in L3-adult survival. When C. carnea first instars were fed upon spinosad-treated-Brevicoryne brassicae (L.), significant differences in investigated parameters were not observed among different treatments. The weight of third instar and pupae were comparable regardless of tested spinosad concentrations. Also, feeding on spinosad treated-B. brassicae had no negative impact on fecundity and fertility. Furthermore, ingestion of spinosad contaminated-B. brassicae significantly prolonged larval and larval + pupal periods. The larval feeding capacity of C. carnea did not differ significantly between larvae fed on spinosad-treated and untreated aphids with the exception of first instar. When C. carnea adults were allowed to oviposit on spinosad treated-substrate, total number of eggs laid, percent of eggs laid on the treated substrate and egg hatching did not differ from those of control. However, when adults were fed on spinosad-treated artificial diet, negative effects on adult survival and fecundity were observed.

Purification and characterization of levoglucosan kinase from <Emphasis Type="Italic">Lipomyces starkeyi</Emphasis> YZ-215

A protein named as levoglucosan kinase (EC 2.7.-.-)was purified to homogeneity from a wild isolated strain of Lipomyces starkeyi YZ-215. The protein was purified approximately 30-fold by conventional ammonium sulphate fractionation followed by Resource Q chromatography and two steps of Superdex 200 chromatography, and its physical and kinetic properties were investigated. The purified enzyme showed a molecular weight of 48 kDa by SDS-PAGE and 47.7 kDa by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS), respectively. The enzyme was stable at pH 7–10 and showed maximum activity at 30°C and pH 9.0. Kinetic constants (apparent K _m values) for levoglucosan and ATP were 68.6 ± 13.7 mM and 0.68 ± 0.06 mM, respectively. After in-gel digestion by trypsin, three peptides were sequenced and analyzed by electrospray ionization quadrupole time-of-flight tandem mass spectrometry (ESI-Q-TOF MS/MS). Data of the amino acid sequences indicated that this protein might be a novel kinase. The purification of levoglucosan kinase from L. starkeyi YZ-215 represented a fundamental step to provide insights into the efficient utilization of cellulosic pyrolysate by bioconversion.     

Influence des radiations lumineuses sur les processus de reproduction des champignons; hypotheses sur l'identite des photorecepteurs. Revue bibliographique

The present review describes the effects of light on reproductive processes in fungi, mainly when action spectra are available.The study of these has resulted in three kinds of photoresponses observable in fungi:

Binding free energy calculations of N-sulphonyl-glutamic acid inhibitors of MurD ligase

The increasing incidence of bacterial resistance to most available antibiotics has underlined the urgent need for the discovery of novel efficacious antibacterial agents. The biosynthesis of bacterial peptidoglycan, where the MurD enzyme is involved in the intracellular phase of UDP-MurNAc-pentapeptide formation, represents a collection of highly selective targets for novel antibacterial drug design. Structural studies of N-sulfonyl-glutamic acid inhibitors of MurD have made possible the examination of binding modes of this class of compounds, providing valuable information for the lead optimization phase of the drug discovery cycle. Binding free energies were calculated for a series of MurD N-sulphonyl-Glu inhibitors using the linear interaction energy (LIE) method. Analysis of interaction energy during the 20-ns MD trajectories revealed non-polar van der Waals interactions as the main driving force for the binding of these inhibitors, and excellent agreement with the experimental free energies was obtained. Calculations of binding free energies for selected moieties of compounds in this structural class substantiated even deeper insight into the source of inhibitory activity. These results constitute new valuable information to further assist the lead optimization process. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.

Personal Experience of Schizophrenia and the Role of <Emphasis Type="Italic">Danwei</Emphasis>: A Case Study in 1990s Beijing

Danwei as a cornerstone of Chinese urban society has received great research attention. The relationship between the Danwei and psychiatric patients, however, remains unclear. This article aims to shed light on the subject with an integrated micro–macro approach. It introduces a historical understanding of mental health in urban China under the “economic state in transition” framework. A detailed case study in clinical sociology is provided to reveal the many social factors affecting the experience of a schizophrenic patient and his significant others. A changing role of the Danwei is hypothesized and validated with qualitative data. The Danwei was shown to have changed significantly before the mid-1990s, yet it still played a major role in urban workers’ lives, including those of psychiatric patients, and even more so in people’s expectations. This lends support to the need for a more responsive public policy to address various social issues brought about by economic reform, with the learning of worldwide experiences including “community care,” “social support” and “social rehabilitation.” Implications for social research, policymaking and professional practice are discussed.