<Emphasis Type="Italic">Polyporus tenuiculus</Emphasis>: a new naturally occurring mushroom that can be industrially cultivated on agricultural waste

Polyporus tenuiculus is a naturally occurring species from Central and South America that is consumed by different ethnic groups in the region. To determine the optimal conditions for fruiting body production, two strains were assayed on wheat straw and sawdust with or without supplements. Sixty days of incubation at 25°C were needed to produce a solid block. The highest yield was obtained with strain ICFC 383/00 grown on supplemented willow sawdust. In a second experiment the strain ICFC 383/00 and different supplements were used to improve the biological efficiency (BE) and to determine the quality traits and its biodegradation capacity. The highest yields were obtained on sawdust with 25% of supplements reaching 82.7% of BE. Supplements raised the number of flushes, generally from four to five, contributing to increased yields. The type of substrate had a significant effect on fruiting body diameters of P. tenuiculus, and the largest mushrooms were harvested on supplemented substrate with the highest BE coinciding with the highest dry matter loss in substrates. P. tenuiculus showed a capacity to degrade sawdust, causing a decrease of 67.2–74.5% in cellulose, 80.4–85.7% in hemicellulose, and 60.6–66.2% in lignin content at the end of the cultivation cycle. The decrease in hemicellulose was relatively greater than that of cellulose and lignin on supplemented substrates. This is the first report of the cultivation of P. tenuiculus on lignocellulosic waste, and it is a promising species both for commercial production and for its potential use in the degradation of other biowastes.     

Effect of methotrexate and anti-TNF on Epstein-Barr virus T-cell response and viral load in patients with rheumatoid arthritis or spondylarthropathies

Introduction

There is a suspicion of increased risk of Epstein-Barr virus (EBV)-associated lymphoproliferations in patients with inflammatory arthritides receiving immunosuppressive drugs. We investigated the EBV load and EBV-specific T-cell response in patients treated with methotrexate (MTX) or anti-TNF therapy.

Methods

Data for patients with rheumatoid arthritis (RA) (n = 58) or spondylarthropathy (SpA) (n = 28) were analyzed at baseline in comparison with controls (n = 22) and after 3 months of MTX or anti-TNF therapy for EBV load and EBV-specific IFNγ-producing T cells in response to EBV latent-cycle and lytic-cycle peptides.

Results

The EBV load and the number of IFNγ-producing T-cells after peptide stimulation were not significantly different between groups at baseline (P = 0.61 and P = 0.89, respectively). The EBV load was not significantly modified by treatment, for RA with MTX (P = 0.74) or anti-TNF therapy (P = 0.94) or for SpA with anti-TNF therapy (P = 1.00). The number of EBV-specific T cells was not significantly modified by treatment, for RA with MTX (P = 0.58) or anti-TNF drugs (P = 0.19) or for SpA with anti-TNF therapy (P = 0.39). For all patients, the EBV load and EBV-specific T cells were significantly correlated (P = 0.017; R = 0.21). For most patients, short-term exposure (3 months) to MTX or anti-TNF did not alter the EBV load or EBV-specific T-cell response but two patients had discordant evolution.

Conclusions

These data are reassuring and suggest there is no short-term defect in EBV-immune surveillance in patients receiving MTX or anti-TNF drugs. However, in these patients, long term follow-up of EBV-specific T-cell response is necessary and the role of non-EBV-related mechanisms of lymphomagenesis is not excluded.     

Pyrenochaeta keratinophila sp. nov., isolated from an ocular infection in Spain

The new coelomycete Pyrenochaeta keratinophila, isolated from corneal scrapings of a case of keratitis in Spain, is described and illustrated. This fungus is morphologically characterized by grey-olivaceous to greenish olivaceous colonies, scarce pycnidial setae placed mainly near the ostiole and production of phialoconidia from the aerial mycelium. The latter feature is unknown in any other species of the genus Pyrenochaeta. Sequencing of the ITS rDNA region of this clinical strain confirmed this proposal and revealed its close genetic relationship with the Leptosphaeriaceae.     

Regulation of Insulin Secretion by Phosphatidylinositol-4,5-Bisphosphate

The role of PIP₂ in pancreatic beta cell function was examined here using the beta cell line MIN6B1. Blocking PIP₂ with PH-PLC-GFP or PIP5KIγ RNAi did not impact on glucose-stimulated secretion although susceptibility to apoptosis was increased. Over-expression of PIP5KIγ improved cell survival and inhibited secretion with accumulation of endocytic vacuoles containing F-actin, PIP₂, transferrin receptor, caveolin 1, Arf6 and the insulin granule membrane protein phogrin but not insulin. Expression of constitutively active Arf6 Q67L also resulted in vacuole formation and inhibition of secretion, which was reversed by PH-PLC-GFP co-expression. PIP₂ co-localized with gelsolin and F-actin, and gelsolin co-expression partially reversed the secretory defect of PIP5KIγ-over-expressing cells. RhoA/ROCK inhibition increased actin depolymerization and secretion, which was prevented by over-expressing PIP5KIγ, while blocking PIP₂ reduced constitutively active RhoA V14-induced F-actin polymerization. In conclusion, although PIP₂ plays a pro-survival role in MIN6B1 cells, excessive PIP₂ production because of PIP5KIγ over-expression inhibits secretion because of both a defective Arf6/PIP5KIγ-dependent endocytic recycling of secretory membrane and secretory membrane components such as phogrin and the RhoA/ROCK/PIP5KIγ-dependent perturbation of F-actin cytoskeleton remodelling.     

Mucopolysaccharidosis I, II, and VI: Brief review and guidelines for treatment

Mucopolysaccharidoses (MPS) are rare genetic diseases caused by the deficiency of one of the lysosomal enzymes involved in the glycosaminoglycan (GAG) breakdown pathway. This metabolic block leads to the accumulation of GAG in various organs and tissues of the affected patients, resulting in a multisystemic clinical picture, sometimes including cognitive impairment. Until the beginning of the XXI century, treatment was mainly supportive. Bone marrow transplantation improved the natural course of the disease in some types of MPS, but the morbidity and mortality restricted its use to selected cases. The identification of the genes involved, the new molecular biology tools and the availability of animal models made it possible to develop specific enzyme replacement therapies (ERT) for these diseases. At present, a great number of Brazilian medical centers from all regions of the country have experience with ERT for MPS I, II, and VI, acquired not only through patient treatment but also in clinical trials. Taking the three types of MPS together, over 200 patients have been treated with ERT in our country. This document summarizes the experience of the professionals involved, along with the data available in the international literature, bringing together and harmonizing the information available on the management of these severe and progressive diseases, thus disclosing new prospects for Brazilian patients affected by these conditions.     

PKA-mediated effect of MAS receptor in counteracting angiotensin II-stimulated renal Na-ATPase

We showed previously that angiotensin-(1-7) [Ang-(1-7)] reversed stimulation of proximal tubule Na⁺-ATPase promoted by angiotensin II (Ang II) through a d-ala⁷-Ang-(1-7) (A779)-sensitive receptor. Here we investigated the signaling pathway coupled to this receptor. According to our data, Ang-(1-7) produces a MAS-mediated reversal of Ang II-stimulated Na⁺-ATPase by a Gs/PKA pathway because: (1) the Ang-(1-7) effect is reversed by GDPβS, an inhibitor of trimeric G protein and Gs polyclonal antibody. Cholera toxin, an activator of Gs protein, mimicked it; (2) in the presence of Ang II, Ang-(1-7) increased the PKA activity 10-fold; (3) the peptide inhibitor of PKA blocked the Ang-(1-7) effect on Ang II-stimulated Na⁺-ATPase; (4) Ang-(1-7) reverses the Ang II-stimulated PKC activity; (5) cAMP mimicked the Ang-(1-7) effect on the Ang II-stimulated Na⁺-ATPase. Our results provide new understanding about the signaling mechanisms coupled to MAS receptor-mediated renal Ang-(1-7) effects.     

Elimination of d-Lactate Synthesis Increases Poly(3-Hydroxybutyrate) and Ethanol Synthesis from Glycerol and Affects Cofactor Distribution in Recombinant Escherichia coli

The effect of eliminating d-lactate synthesis in poly(3-hydroxybutyrate) (PHB)-accumulating recombinant Escherichia coli (K24K) was analyzed using glycerol as a substrate. K24KL, an ldhA derivative, produced more biomass and had altered carbon partitioning among the metabolic products, probably due to the increased availability of carbon precursors and reducing power. This resulted in a significant increase of PHB and ethanol synthesis and a decrease in acetate production. Cofactor measurements revealed that cultures of K24K and K24KL had a high intracellular NADPH content and that the NADPH/NADP⁺ ratio was higher than the NADH/NAD⁺ ratio. The ldhA mutation affected cofactor distribution, resulting in a more reduced intracellular state, mainly due to a further increase in NADPH/NADP⁺. In 60-h fed-batch cultures, K24KL reached 41.9 g·liter⁻¹ biomass and accumulated PHB up to 63% ± 1% (wt/wt), with a PHB yield on glycerol of 0.41 ± 0.03 g·g⁻¹, the highest reported using this substrate.Poly(3-hydroxybutyrate) (PHB) is the best-known and most common polyhydroxyalkanoate (PHA). PHAs are polymers with thermoplastic properties that are totally biodegradable by microorganisms present in most environments and that can be produced from different renewable carbon sources (38). Accumulated as intracellular granules by many bacteria under unfavorable conditions (1, 21), PHAs are carbon and energy reserves and also act as electron sinks, enhancing the fitness and stress resistance of bacteria and contributing to redox balance (12, 30). Escherichia coli offers a well-defined physiological environment for the construction and manipulation of various metabolic pathways to produce different bioproducts, such as PHB, from cost-effective carbon sources.In recent years, a significant increase in the production of biodiesel has caused a sharp fall in the cost of glycerol, the main by-product of biodiesel synthesis. As a result, glycerol has become a very attractive substrate for bacterial fermentations (10), specially for reduced products, such as PHB (36). The E. coli strain used in this work, K24K, carries phaBAC, the structural genes responsible for PHB synthesis, from Azotobacter sp. strain FA8 (23) (Table ​(Table1).1). The pha genes in K24K are expressed from a chimeric promoter and consequently are not subject to the genetic regulatory systems present in natural PHA producers. Because of this, it can be assumed that regulation of PHA synthesis in the recombinants is restricted by enzyme activity levels, modulated principally by substrate availability. In most natural producers, and also in PHB-producing E. coli recombinants, PHB is synthesized through the condensation of two molecules of acetyl-coenzyme A (acetyl-CoA), catalyzed by an acetoacetyl-CoA transferase or 3-ketothiolase, resulting in acetoacetyl-CoA. This compound is subsequently reduced by an NAD(P)H-dependent acetoacetyl-CoA reductase to R-(−)-3-hydroxybutyryl-CoA, which is then polymerized by a specific PHA synthase (34).

TABLE 1.

E. coli strains, plasmids, and oligonucleotides used in this study

Seasonal climatic fluctuations in the Late Triassic tropics—High-resolution oxygen isotope records from aragonitic bivalve shells (Cassian Formation,Northern Italy)

Three megalodontoid bivalves from the Upper Triassic Cassian Formation (N Italy, Dolomites) were sampled for δ¹³C and δ¹⁸O sclerochronology (n = 270). With more than 1000 described invertebrate species, the Cassian Formation has one of the best records of an Early Mesozoic tropical fauna. In addition, the Cassian Formation is one of the very few Triassic occurrences with original aragonite preservation, with all studied shells consisting of pristine aragonite. The presence of aragonite and crossed lamellar as well as fibrous prismatic shell microstructures shown for the first time for Triassic megalodontoids suggests absence or minimal impact of diagenetic alteration. The δ¹³C values range from 3.6 to 5.8‰ and show a distinct cyclicity in two studied shells whereas a third shell shows no obvious cyclicity. In one bivalve specimen, the cycles are somewhat offset from δ¹⁸O cycles and in the other specimen δ¹⁸O and δ¹³C curves are inversely correlated. Seasonal variation in freshwater runoff including nutrient input and subsequent changes in plankton productivity during dry and wet seasons may explain δ¹³C cyclicity. δ¹⁸O values show a pronounced cyclicity within each of the studied shells varying from ? 3.6 to ? 1.4‰. The variations in δ¹⁸O suggest a pronounced seasonality in Late Triassic tropical shallow waters of the western Tethys, with inferred seasonal temperature changes ranging from 24 to 32 °C. Influx of fresh water during the rainy seasons (mega-monsoon) or upwelling might also explain part of the variation in δ¹⁸O. The presented data suggest that the diverse Cassian fauna lived under conditions characterized by warm sea-surface waters with a pronounced seasonality. Similar conditions are reported for some modern tropical settings.