Three new species of Lentinus from northern Thailand

There have been few studies on the taxonomy and biodiversity of the genus Lentinus in Thailand, which is a genus of edible mushrooms. Recently, collections from 17 sites in northern Thailand yielded 47 specimens of Lentinus sensu lato. Three were shown to be new species of Lentinus sensu stricto and Lentinus roseus, L. concentricus and L. megacystidiatus are introduced in this paper. The new species are described and illustrated with line drawings and are justified and compared with similar taxa. Furthermore, ITS sequence data do not match closely with any species presently lodged in GenBank.     

In vivo electroporation of morpholinos into the adult zebrafish retina

Many devastating inherited eye diseases result in progressive and irreversible blindness because humans cannot regenerate dying or diseased retinal neurons. In contrast, the adult zebrafish retina possesses the robust ability to spontaneously regenerate any neuronal class that is lost in a variety of different retinal damage models, including retinal puncture, chemical ablation, concentrated high temperature, and intense light treatment. Our lab extensively characterized regeneration of photoreceptors following constant intense light treatment and inner retinal neurons after intravitreal ouabain injection. In all cases, resident Müller glia re-enter the cell cycle to produce neuronal progenitors, which continue to proliferate and migrate to the proper retinal layer, where they differentiate into the deficient neurons. We characterized five different stages during regeneration of the light-damaged retina that were highlighted by specific cellular responses. We identified several differentially expressed genes at each stage of retinal regeneration by mRNA microarray analysis. Many of these genes are also critical for ocular development. To test the role of each candidate gene/protein during retinal regeneration, we needed to develop a method to conditionally limit the expression of a candidate protein only at times during regeneration of the adult retina. Morpholino oligos are widely used to study loss of function of specific proteins during the development of zebrafish, Xenopus, chick, mouse, and tumors in human xenografts. These modified oligos basepair with complementary RNA sequence to either block the splicing or translation of the target RNA. Morpholinos are stable in the cell and can eliminate or "knockdown" protein expression for three to five days. Here, we describe a method to efficiently knockdown target protein expression in the adult zebrafish retina. This method employs lissamine-tagged antisense morpholinos that are injected into the vitreous of the adult zebrafish eye. Using electrode forceps, the morpholino is then electroporated into all the cell types of the dorsal and central retina. Lissamine provides the charge on the morpholino for electroporation and can be visualized to assess the presence of the morpholino in the retinal cells. Conditional knockdown in the retina can be used to examine the role of specific proteins at different times during regeneration. Additionally, this approach can be used to study the role of specific proteins in the undamaged retina, in such processes as visual transduction and visual processing in second order neurons.     

Atomic layer deposition of titanium dioxide on cellulose acetate for enhanced hemostasis

TiO₂ films may be used to alter the wettability and hemocompatibility of cellulose materials. In this study, pure and stoichiometric TiO₂ films were grown using atomic layer deposition on both silicon and cellulose substrates. The films were grown with uniform thicknesses and with a growth rate in agreement with literature results. The TiO₂ films were shown to profoundly alter the water contact angle values of cellulose in a manner dependent upon processing characteristics. Higher amounts of protein adsorption indicated by blurry areas on images generated by scanning electron microscopy were noted on TiO₂-coated cellulose acetate than on uncoated cellulose acetate. These results suggest that atomic layer deposition is an appropriate method for improving the biological properties of hemostatic agents and other blood-contacting biomaterials.     

A novel form of oxytocin in New World monkeys

Oxytocin is widely believed to be present and structurally identical in all placental mammals. Here, we report that multiple species of New World monkeys possess a novel form of oxytocin, [P8] oxytocin. This mutation arises from a substitution of a leucine to a proline in amino acid position 8. Further analysis of this mutation in Saimiri sciureus (squirrel monkey) indicates that [P8] oxytocin is transcribed and translated properly. This mutation is specific to oxytocin, as the peptide sequence for arginine vasopressin, a structurally related nonapeptide, is unaltered. These findings dispel the notion that all placental mammals possess a 'universal' oxytocin sequence, and highlight the need for research on the functional significance of this novel nonapeptide in New World monkeys.     

Immunomodulatory compounds from Pestalotiopsis leucothës, an endophytic fungus from Tripterygium wilfordii

The immunomodulatory effects of three compounds designated BS, GS, and YS produced by Pestalotiopsis leucothës, an endophytic fungus isolated from Tripterygium wilfordii, were evaluated. The 50% inhibition concentration (IC₅₀) value of BS in the proliferative assay with various stimulating agents such as phytohemagglutinin-M (PHA-M), phorbol myristate acetate (PMA)/ionomycin, mixed lymphocyte reaction (MLR) and poke weed mitogen (PWM) was 0.35, 1.6, 0.8 and 5.4 μg/ml, respectively. In addition, BS significantly inhibited the production of cytokines such as interleukin (IL)-1β, IL-2, interferon (IFN)-γ and tumor necrosis factor (TNF)-α, by peripheral blood mononuclear cells (PBMNC) and soluble IL-2 receptor expression at concentrations greater than 1 μg/ml. Inhibition of PHA stimulated PBMNC proliferation and IL-2 and sIL-2R production by BS indicates that it is a T-cell specific immunosuppressant. However, BS also moderately inhibited immunoglobulin (Ig) G and M at concentrations greater than 1 μg/ml suggesting that it also has B cell immunosuppressive effects. YS was 10% less active than BS in all assay systems. In contrast, GS exhibited both suppression and enhancement of PBMNC proliferation in the presence of various stimulants. However, GS inhibited PWM stimulated PBMNC proliferation and IL-4 and IgG and IgM production at concentrations above 1 μg/ml. All three fungal compounds altered the percentage of T-lymphocyte subpopulations only at high concentrations. Cell viability was not affected at the immunosuppressive concentrations of these compounds. In conclusion, work from our laboratory has identified three potentially potent immunomodulatory compounds from P. leucothës. These compounds have variable effects on T- and B-cells and monocytes. They may partially explain the immunosuppressive activity of T. wilfordii. In addition, they may represent a new source of immunomodulatory compounds for the treatment of human immune mediated diseases.     

Phylogeny of Rosellinia capetribulensis sp. nov. and its allies (Xylariaceae)

A new Rosellinia species, R. capetribulensis isolated from Calamus sp. in Australia is described. R. capetribulensis is characterized by perithecia immersed within a carbonaceous stroma surrounded by subiculum-like hyphae, asci with large, barrel-shaped amyloid apical apparatus and large dark brown spores. Morphologically, R. capetribulensis appears to be similar to R. bunodes, R. markhamiae and R. megalospora. To gain further insights into the phylogeny of this new taxon we analyzed the ITS-5.8S rDNA using maximum parsimony and likelihood methods. In addition, a morphological dataset also was analyzed phylogenetically to investigate possible affinities. ITS rDNA based phylogenies reveal that R. capetribulensis is closely related to other Rosellinia species showing closest affinity to R. arcuata, RL necatrix and R. pepo. However, analysis of R. capetribulensis forms an unsupported branch sister to these taxa. Results from the morphological matrix indicate a close morphological affinity to members of Rosellinia subgenus Rosellinia. Despite that ITS rDNA and morphological analyses present difficulties in constructing a proper phylogenetic framework among Rosellinia and allied genera, there is sufficient evidence to support the establishment of the new taxon in the genus Rosellinia. The morphological similarities and differences between R. capetribulensis and allied genera such as Astrocystis and Entoleuca are also briefly discussed.     

Phylogenetics and evolution of nematode-trapping fungi (Orbiliales) estimated from nuclear and protein coding genes

The systematic classification of nematode-trapping fungi is redefined based on phylogenies inferred from sequence analyses of 28S rDNA, 5.8S rDNA and beta-tubulin genes. Molecular data were analyzed with maximum parsimony, maximum likelihood and Bayesian analysis. An emended generic concept of nematode-trapping fungi is provided. Arthrobotrys is characterized by adhesive networks, Dactylellina by adhesive knobs, and Drechslerella by constricting-rings. Phylogenetic placement of taxa characterized by stalked adhesive knobs and non-constricting rings also is confirmed in Dactylellina. Species that produce unstalked adhesive knobs that grow out to form loops are transferred from Gamsylella to Dactylellina, and those that produce unstalked adhesive knobs that grow out to form networks are transferred from Gamsylella to Arthrobotrys. Gamsylella as currently circumscribed cannot be treated as a valid genus. A hypothesis for the evolution of trapping-devices is presented based on multiple gene data and morphological studies. Predatory and nonpredatory fungi appear to have been derived from nonpredatory members of Orbilia. The adhesive knob is considered to be the ancestral type of trapping device from which constricting rings and networks were derived via two pathways. In the first pathway adhesive knobs retained their adhesive material forming simple two-dimension networks, eventually forming complex three-dimension networks. In the second pathway adhesive knobs lost their adhesive materials, with their ends meeting to form nonconstricting rings and they in turn formed constricting rings with three inflated-cells.     

Comparative folate metabolism in humans and malaria parasites (part II): activities as yet untargeted or specific to Plasmodium

The folate pathway represents a powerful target for combating rapidly dividing systems such as cancer cells, bacteria and malaria parasites. Whereas folate metabolism in mammalian cells and bacteria has been studied extensively, it is understood less well in malaria parasites. In two articles, we attempt to reconstitute the malaria folate pathway based on available information from mammalian and microbial systems, in addition to Plasmodium-genome-sequencing projects. In part I, we focused on folate enzymes that are already used clinically as anticancer drug targets or that are under development in drug-discovery programs. In this article, we discuss mammalian folate enzymes that have not yet been exploited as potential drug targets, and enzymes that function in the de novo folate-synthesis pathway of the parasite--a particularly attractive area of attack because of its absence from the mammalian host.     

Verification of a European corn borer (Lepidoptera: Crambidae) loss equation in the major corn production region of the northeastern United States

Field studies in Pennsylvania and Maryland were conducted during 2000, 2001, and 2002 to test the applicability of published yield loss relationships developed in central Pennsylvania for European corn borer, Ostrinia nubilalis (Hübner), management in warmer, longer season corn, Zea mays L., grain production regions of the northeastern United States. Both isoline hybrids and non-Bt lead hybrids were compared against Bacillus thuringiensis (Bt) hybrids to measure effects of the pest on yield. The European corn borer economic analysis model was used to make site-specific predictions of loss per European corn borer larva for comparison with measured yield loss per larva. Although the model did not predict loss per larva at a field level, it did predict loss at a regional level. The model predicted an overall percentage of yield loss per larva of 2.69+/-0.12% over the region, which was similar to the measured yield loss per larva of 2.66+/-0.59% for isoline hybrids and 3.08+/-0.51% for lead hybrids. The model, on average, provided a good prediction of percentage of yield loss per larva within the climatic zones of 1100-1700 degree-days (DD) (base threshold 12.5 degrees C). Our results suggest that the yield loss relationship developed in Central Pennsylvania, when matched to the timing of third instar second generation European corn borer stalk tunneling is adequate for major corn grain production zones of the northeast United States.     

Shortcomings of protein removal prior to high performance liquid chromatographic analysis-A case study using method development for BAY 11-7082

During the analytical method development for BAY 11-7082 ((E)-3-[4-methylphenylsulfonyl]-2-propenenitrile), using HPLC-MS-MS and HPLC-UV, we observed that the protein removal process (both ultrafiltration and precipitation method using organic solvents) prior to HPLC brought about a significant reduction in the concentration of this compound. The use of a structurally similar internal standard, BAY 11-7085 ((E)-3-[4-t-butylphenylsulfonyl]-2-propenenitrile), was not effective in compensating for the loss of analyte as the extent of reduction was different to that of the analyte. We present here a systematic investigation of this problem and a new validated method for the determination of BAY 11-7082.