Purification and biochemical characterization of a mycelial glucose- and xylose-stimulated beta-glucosidase from the thermophilic fungus Humicola insolens

A mycelial beta-glucosidase from the thermophilic mold Humicola insolens was purified and biochemically characterized. The enzyme showed carbohydrate content of 21% and apparent molecular mass of 94 kDa, as estimated by gel filtration. Sodium dodecyl sulfate polyacrylamide gel electrophoresis analysis showed a single polypeptide band of 55 kDa, suggesting that the native enzyme was a homodimer. Mass spectrometry analysis showed amino acid sequence similarity with a P-glucosidase from Humicola grisea var. thermoidea, with about 22% coverage. Optima of temperature and pH were 60 degrees C and 6.0-6.5, respectively. The enzyme was stable up to I h at 50 degrees C and showed a half-life of approximately 44 min at 55 degrees C. The beta-glucosidase hydrolyzed cellobiose, lactose, p-nitrophenyl-beta-D-glucopyranoside, p-nitrophenyl-beta-D-fucopyranoside, p-nitrophenyl-beta-D-xylopyranoside, p-nitrophenyl-beta-D-galactopyranoside, o-nitrophenyl-beta-D-galactopyranoside, and salicin. Kinetic studies showed that p-nitrophenyl-beta-D-fucopyranoside and cellobiose were the best enzyme substrates. Enzyme activity was stimulated by glucose or xylose at concentrations up to 400 mM, with maximal stimulatory effect (about 2-fold) around 40 mM. The high catalytic efficiency for the natural substrate, good thermal stability, strong stimulation by glucose or xylose, and tolerance to elevated concentrations of these monosaccharides qualify this enzyme for application in the hydrolysis of cellulosic materials. (C) 2009 Elsevier Ltd. All rights reserved.

Differential responses of antioxidant enzymes in pioneer and late-successional tropical tree species grown under sun and shade conditions

To investigate the ability of pioneer and late-successional species to adapt to a strong light environment in a reforestation area, we examined the activities of antioxidant enzymes in relation to photosystem chlorophyll a fluorescence and photosynthetic pigment concentration for eight tropical tree species grown under 100% (sun) and 10% (shade) sunlight irradiation. The pioneer (early-succession) species (PS) were Cecropia pachystachya, Croton urucurana, Croton floribundus and Schinus terebinthifolius. The non-pioneer (late succession) species (LS) were Hymenaea courbaril L var. stilbocarpa, Esenbeckia leiocarpa, Cariniana legalis and Tabebuia roseo-alba. We observed a greater decline in the ratio of variable to maximum chlorophyll a fluorescence (F(v)/F(m)) under full sunlight irradiation in the late-successional species than in the pioneer species. The LS species most sensitive to high irradiance were C. legalis and H. courbaril. In LS species, chlorophyll a, chlorophyll b and total chlorophyll concentrations were higher in the shade-grown plants than in plants that developed under full sunlight, but in the PS species C. floribundus and C. pachystachya, we did not observe significant changes in chlorophyll content when grown in the two contrasting environments. The carotenoids/total chlorophyll ratio increased significantly when plants developed under high-sunlight irradiation, but this response was not observed in the PS species S. terebinthifolius and C. pachystachya. The improved performance of the pioneer species in high sunlight was accompanied by an increase in superoxide dismutase (SOD. EC 1.15.1.1) activity, though no light-dependent increase in the activity of ascorbate peroxidase (APX. EC 1.11.1.11) was observed. The activity of catalase (CAT, EC 1.11.1.6) was reduced by high irradiation in both pioneer and late-successional species. Our results show that pioneer species perform better under high-sunlight irradiation than late-successional species, as indicated by increased SOD activity and a higher F IF,, ratio. C. legalis was the LS species most susceptible to photoinhibition under full sunlight conditions. These results suggest that pioneer plants have more potential tolerance to photo-oxidative damage than late-successional species associated with the higher SOD activity found in pioneer species. Reduced photoinhibition in pioneer species probably results from their higher photosynthetic capacities, as has been observed in a previous survey carried out by our group. (C) 2010 Elsevier B.V. All rights reserved.

Monte Carlo evidence on cointegration and causation

The small sample performance of Granger causality tests under different model dimensions, degree of cointegration, direction of causality, and system stability are presented. Two tests based on maximum likelihood estimation of error-correction models (LR and WALD) are compared to a Wald test based on multivariate least squares estimation of a modified VAR (MWALD). In large samples all test statistics perform well in terms of size and power. For smaller samples, the LR and WALD tests perform better than the MWALD test. Overall, the LR test outperforms the other two in terms of size and power in small samples.

Exercise Training Reduces Sympathetic Modulation on Cardiovascular System and Cardiac Oxidative Stress in Spontaneously Hypertensive Rats

BACKGROUND Spontaneously hypertensive rats (SHRs) show increased cardiac sympathetic activity, which could stimulate cardiomyocyte hypertrophy, cardiac damage, and apoptosis. Norepinephrine (NE)induced cardiac oxidative stress seems to be involved in SHR cardiac hypertrophy development. Because exercise training (ET) decreases sympathetic activation and oxidative stress, it may alter cardiac hypertrophy in SHR. The aim of this study was to determine, in vivo, whether ET alters cardiac sympathetic modulation on cardiovascular system and whether a correlation exists between cardiac oxidative stress and hypertrophy. METHODS Male SHRs (15-weeks old) were divided into sedentary hypertensive (SHR, n = 7) and exercise-trained hypertensive rats (SHR-T, n = 7). Moderate ET was performed on a treadmill (5 days/week, 60 min, 10 weeks). After ET, cardiopulmonary reflex responses were assessed by bolus injections of 5-HT. Autoregressive spectral estimation was performed for systolic arterial pressure (SAP) with oscillatory components quantified as low (LF: 0.2-0.75 Hz) and high (HF:0.75-4.0 Hz) frequency ranges. Cardiac NE concentration, lipid peroxidation, antioxidant enzymes activities, and total nitrates/nitrites were determined. RESULTS ET reduced mean arterial pressure, SAP variability (SAP var), LIF of SAP, and cardiac hypertrophy and increased cardiopulmonary reflex responses. Cardiac lipid peroxidation was decreased in trained SHRs and positively correlated with NE concentrations (r= 0.89, P < 0.01) and heart weight/body weight ratio (r= 0.72, P < 0.01), and inversely correlated with total nitrates/nitrites (r= -0.79, P < 0.01). Moreover, in trained SHR, cardiac total nitrates/nitrites were inversely correlated with NE concentrations (r= -0.82, P < 0.01). CONCLUSIONS ET attenuates cardiac sympathetic modulation and cardiac hypertrophy, which were associated with reduced oxidative stress and increased nitric oxide (NO) bioavailability. Am J Hypertens 2008;21:1138-1193 (C) 2008 American Journal of Hypertension, Ltd.

Bacterial and fungal pneumonias after lung transplantation

Objective. The aim of this study was to evaluate the epidemiology of bacterial and fungal pneumonia in lung transplant (LT) recipients and to assess donor-to-host transmission of these microorganisms. Materials and Methods. We retrospectively studied all positive cultures from bronchoalveolar lavage (BAL) of 49 lung transplant recipients and their donors from August 2003 to April 2007. Results. There were 108 episodes of pneumonia during a medium follow-up of 412 days (range, 1-1328 days). The most frequent microorganisms were: Pseudomonas aeruginosa (n = 36; 33.3%), Staphylococcus aureus (n = 29; 26.8%), and Aspergillus spp. (n = 18; 16%). Other fungal infections were due to Fusarium spp., Cryptococcus neoformans, and Paracoccidioides brasiliensis. Of the 31 donors with positive BAL, 15 had S. aureus. There were 21 pretransplant colonized recipients (43%) and 16 of them had suppurative underlying lung disease. P. aeruginosa was the most frequent colonizing organism (59% of pretransplant positive cultures). There were 11 episodes of bacteremia and lungs were the source in 5 cases. Sixteen deaths occurred and 6 (37.5%) were due to infection. Statistical analyses showed association between pretransplant colonizing microorganisms from suppurative lung disease patients and pneumonias after lung transplantation (RR = 4.76; P = .04; 95% CI = 1.02-22.10). No other analyzed factor was significant. Conclusions. Bacterial and fungal infections are frequent and contribute to higher mortality in lung transplant recipients. P. aeruginosa is the most frequent agent of respiratory infections. This study did not observe any impact of donor lung organisms on pneumonia after lung transplantation. Nevertheless, we demonstrated an association between pretransplant colonizing microorganisms and early pneumonias in suppurative lung transplant recipients.

Infectious diseases in paediatric pathology: experience from a developing country

Infectious and parasitic diseases have always challenged man. Although many of them are typically seen in some areas of the world and can be adequately managed by just improving socioeconomic status and sanitary conditions, they are still quite prevalent and may sometimes be seen outside their original geographical areas. Human migration due to different reasons, tourism, blood transfusion and solid organ transplantation has created new concerns for health professionals all over the world. If not for diagnostic purposes, at least these tropical and infectious diseases should be largely known because their epidemiology, pathogenesis, host/parasite interaction, inflammatory and reparative responses are quite interesting and teach us about human biology. Curiosity is inherent to pathology practice and so we are compelled to look for things other than tumours or degenerative diseases. This review focuses on infectious and parasitic diseases found in a developing country and brings up-to-date information on diseases caused by viruses (dengue, yellow fever), bacteria (typhoid fever, leprosy), parasites (Chagas` disease, cutaneous and visceral leishmaniasis, amoebiasis, Capillaria hepatica, schistosomiasis, cysticercosis) and caused by fungi (paracoccidioidomycosis, cryptococcosis, histoplasmosis) that may be useful for pathologists when facing somewhat strange cases from developing countries.

Role of Interleukin (IL)-18 in experimental paracoccidioidomycosis

Interleukin (IL)-18 has been regarded as a Th1 type cytokine involved in many fungal and parasitic infections. Since there have been no studies, as of yet, evaluating the role of this cytokine in paracoccidioidomycosis (PCM), we assessed the function of IL-18 by using an experimental PCM model. Our results showed that IL-18 knockout (IL-18-/-) BALB/c were more resistant to Paracoccidioides brasiliensis than their littermate controls (WT). In fact, mortality rate was higher in WT mice and in the first month of infection, the number of colony forming units of the etiologic agent recovered from the lungs was greater in WT mice. In histopathological analyses, well-formed granulomas were seen in both WT and IL-18-/- mice. However, substantial differences were observed at the second month of infection when epithelioid cells predominated in the lesions of IL-18-/- mice, which could infer that IL-18 postpones pulmonary healing. The levels of IL-10 were significantly higher in IL-18 sufficient mice at early stages of infection and therefore account for the delayed fungal clearance observed in WT mice. TNF- augmented later in the infection of WT mice, seemingly to compensate high levels of IL-10. Our results demonstrated that IL-18 has a critical role in protecting BALB/c mice against disseminated PCM.

Anti-inflammatory effects of red pepper (Capsicum baccatum) on carrageenan- and antigen-induced inflammation

Inflammation is a pivotal component of a variety of diseases, such as atherosclerosis and tumour progression. Various naturally occurring phytochemicals exhibit anti-inflammatory activity and are considered to be potential drug candidates against inflammation-related pathological processes. Capsicum baccatum L. var. pendulum (Willd.) Eshbaugh (Solanaceae) is the most consumed species in Brazil, and its compounds, such as capsaicinoids, have been found to inhibit the inflammatory process. However, the anti-inflammatory effects of C. baccatum have not been characterized. Thus, this study was designed to evaluate the effects of C. baccatum juice in animal models of acute inflammation induced by carrageenan and immune inflammation induced by methylated bovine serum albumin. Pretreatment (30 min) of rats with pepper juice (0.25-2.0 g kg(-1)) significantly decreased leucocyte and neutrophil migration, exudate volume and protein and LDH concentration in pleural exudates of a pleurisy model. This juice also inhibited neutrophil migration and reduced the vascular permeability on carrageenan-induced peritonitis in mice. C. baccatum juice also reduced neutrophil recruitment and exudate levels of pro-inflammatory cytokines TNF-alpha, and IL-1 beta in mouse inflammatory immune peritonitis. Furthermore, we demonstrated that the main constituent of C. baccatum juice, as extracted with chloroform, is capsaicin. In agreement with this, capsaicin was able to inhibit the neutrophil migration towards the inflammatory focus. To our knowledge, this is the first demonstration of the anti-inflammatory effect of C. baccatum juice and our data suggest that this effect may be induced by capsaicin. Moreover, the anti-inflammatory effect induced by red pepper may be by inhibition of pro-inflammatory cytokine production at the inflammatory site.

Antimicrobial and Cytotoxic Activity of Fruit Extract from Syzygium cumini (L.) Skeels

The aim of the present study was to evaluate the antimicrobial and cytotoxic activity of the ethanolic extract of S. cumini according to the Clinical and Laboratory Standards Institute reference method (with modifications), determining the minimal inhibitory and lethal concentration. Activity against Gram-positive (Staphylococcus aureus and S. epidermidis), Gram-negative (Pseudomonas aeruginosa) and yeast of Candida sp and Cryptococcus neoformans was evaluated. The effects of the fruit extract were examined in hamster cells ovaries in concentrations ranging from 1250.0 a 4.9 mu g/ml, measuring the reduction of the tetrazolium salt 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulphophenyl)-2H-tetrazolium. The extract showed both bactericidal and fungicidal activity among the various microorganisms tested and the MIC ranging from 7.8 to 250 mu g/ml. The MIC, MBC and MFC should values that were similar for all the microorganisms. Cytotoxicity index of the dried extract corresponded to the concentration of 400 mu g/ml. The extract could potentially be used in topical antimicrobial products. Thus, the activity of extract was potent to bacteria and mainly to non-albicans species and C. neoformans.

Pfaffia paniculata (Brazilian ginseng) roots decrease proliferation and increase apoptosis but do not affect cell communication in murine hepatocarcinogenesis

Pfaffia paniculata (Brazilian ginseng) roots and/or its extracts have shown anti-neoplastic, chemopreventive, and anti-angiogenic properties. The aim of this work was to investigate the chemopreventive mechanisms of this root in Mice Submitted to the infant model of hepatocarcinogenesis, evaluating the effects oil cellular proliferation, apoptosis. and intercellular communication. Fifteen-day-old BALB/c male mice were given, i.p., 10 mu g/g of the carcinogen N-nitrosodiethylamine (DEN). Animals were separated into three groups at weaning and were given different concentrations of powdered P. paniculata root (0%, 2%, or 10%) added to commercial food for 27 weeks. Control group (CT) was not exposed to the carcinogen and was given ration without the root. After euthanasia, the animals` liver and body weight were measured. Liver fragments were sampled to Study intercellular communication, molecular biology, and histopathological analysis. Cellular proliferation was evaluated by immunohistochemistry for PCNA, apoptosis was evaluated by apoptotic bodies count and alkaline cornet technique, and inter-cellular communication by diffusion of lucifer yellow dye, immunofluorescence, western blot and real-time PCR for connexins 26 and 32. Chronic treatment with powdered P. paniculata root reduced cellular proliferation and increased apoptosis in the 2%, group. Animals in the 10% group had an increase in apoptosis with chronic inflammatory process. Intercellular communication showed no alterations in any of the groups analyzed. These results Indicate that chemopreventive effects of P. paniculata are related to the control of cellular proliferation and apoptosis, but not to cell communication and/or connexin expression, and are directly Influenced by the root concentration. (C) 2009 Elsevier GmbH. All rights reserved.

Protective action of indole-3-acetic acid on induced hepatocarcinoma in mice

In this study, we report the protective effects of IAA on diethylnitrosamine (DEN)-induced hepatocarcinogenesis. BALB/c mice received daily IAA at 50 (T(50)), 250 (T(250)), and 500 (T(500)) mg K(-1) per body mass by gavage for 15 days. At day 15, animals were administered DEN and sacrificed 4 h later. Alanine aminotransferase (ALT) and aspartate aminotransferase (AST) were analyzed in sera. In addition., hepatomorphologic alterations, activity of superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx), and glutathione reductase (GR), gene expression of antioxidant enzymes and DNA integrity were evaluated in the liver. IAA administration did not show any alterations in any of the parameters available, except for a reduction of the gene expression for antioxidant enzyrries by 55, 56, 27, and 28% for SOD, CAT, GPx, and GR upon T(500). respectively compared with the control. Several hepatic alterations were observed by DEN exposure. Moreover, IAA administration at 3 doses was shown to provide a total prevention of the active reduction of CAT and GR induced by DEN exposure compared with the control. IAA at T5(00) was shown to give partial protection (87, 71, 57, and 90% for respectively SOD, CAT. GPx. and GR) on the down-regulation of the enzymes induced by DEN and this auxin showed a partial protection (50%) on DEN-induced DNA fragmentation for both parameters when compared to DEN alone. This work showed IAA hepatocarcinogenesis protection for the first time by means of a DEN-protective effect on CAT and GR activity. and by affecting antioxidant gene expression and DNA fragmentation. Copyright (C) 2008 John Wiley & Sons, Ltd.

The sequence of a 200 kb portion of a Plasmodium vivax chromosome reveals a high degree of conservation with Plasmodium falciparum chromosome 3

Within a 199 866 base pair (bp) portion of a Plasmodium vivax chromosome we identified a conserved linkage group consisting of at least 41 genes homologous to Plasmodium falciparum genes located on chromosome 3. There were no P. vivax homologues of the P. falciparum cytoadherence-linked asexual genes clag 3.2, clag 3.1 and a var C pseudogene found on the P. vivax chromosome. Within the conserved linkage group, the gene order and structure are identical to those of P. falciparum chromosome 3. This conserved linkage group may extend to as many as 190 genes. The subtelomeric regions are different in size and the P. vivax segment contains genes for which no P. falciparum homologues have been identified to date. The size difference of at least 900 kb between the homologous P. vivax chromosome and P. falciparum chromosome 3 is presumably due to a translocation. There is substantial sequence divergence with a much higher guanine + cytosine (G + C) content in the DNA and a preference for amino acids using GC-rich codons in the deduced proteins of P. vivax. This structural conservation of homologous genes and their products combined with sequence divergence at the nucleotide level makes the P. vivax genome a powerful tool for comparative analyses of Plasmodium genomes. (C) 2001 Elsevier Science B.V. All rights reserved.

Time of treatment influences the appearance of drug-resistant parasites in Plasmodium falciparum infections

A deterministic mathematical model which predicts the probability of developing a new drug-resistant parasite population within the human host is reported, The model incorporates the host's specific antibody response to PfEMP1, and also investigates the influence of chemotherapy on the probability of developing a viable drug-resistant parasite population within the host. Results indicate that early, treatment, and a high antibody threshold coupled with a long lag time between antibody stimulation and activity, are risk factors which increase the likelihood of developing a viable drug-resistant parasite population. High parasite mutation rates and fast PfEMP1 var gene switching are also identified as risk factors. The model output allows the relative importance of the various risk factors as well as the relationships between them to be established, thereby increasing the understanding of the conditions which favour the development of a new drug-resistant parasite population.

Transpecific microsatellites for hard pines M. Shepherd, M. Cross, T. Maguire, M. Dieters, C. Williams, R. Henry

Microsatellites are difficult to recover from large plant genomes so cross-specific utilisation is an important source of markers. Fifty micro satellites were tested for cross-specific amplification and polymorphism to two New World hard pine species, slash pine (Pinus elliottii var. elliottii) and Caribbean pine (R caribaea var. hondurensis). Twenty-nine (58%) markers amplified in both hard pine species, and 23 of these 29 were polymorphic. Soft pine (subgenus Strobus) microsatellite markers did amplify, but none were polymorphic. Pinus elliottii var. elliottii and R caribaea var. hondurensis showed mutational changes in the flanking regions and the repeat motif that were informative for Pinus spp. phylogenetic relationships. Most allele length variation could be attributed to variability in repeat unit number. There was no evidence for ascertainment bias.

Genetic variation in foliar carbon isotope composition in relation to tree growth and foliar nitrogen concentration in clones of the F-1 hybrid between slash pine and Caribbean pine

The objectives of this study were: (1) to quantify the genetic variation in foliar carbon isotope composition (delta(13)C) of 122 clones of ca. 4-year-old F-1 hybrids between slash pine (Pinus elliottii Engelm var. elliottii) and Caribbean pine (Pinus caribaea var. hondurensis Barr.,et Golf.) grown at two field experimental sites with different water and nitrogen availability in southeast Queensland, Australia, in relation to tree growth and foliar nitrogen concentration (N-mass); and (2) to assess the potential of using delta(13)C measurements, in the foliage materials collected from the clone hedges at nursery and the 4-year-old tree canopies in the field, as an indirect index of tree water use efficiency for selecting elite F-1 hybrid pine clones with improved tree growth. There were significant differences in foliar delta(13)C between the nursery hedges and the 4-year-old tree canopies in the field, between the summer and winter seasons, between the two experimental sites, and between the upper outer and lower outer canopy positions sampled. This indicates that delta(13)C measurements in the foliage materials are significantly influenced by the sampling techniques and environmental conditions. Significant differences in foliar delta(13)C, at the upper outer canopy in both field experiments in summer and winter, were detected between the clones, and between the female parents of the clones. Clone means of tree height at age ca. 3 years were positively related to those of the upper outer canopy delta(13)C at both experimental sites in winter, but only for the wetter site in summer. There were positive, linear relationships between clone means of canopy delta(13)C and those of canopy N-mass, indicating that canopy photosynthetic capacity might be an important factor regulating the clonal variation in canopy delta(13)C. Significant correlations were found between clone means of canopy delta(13)C at both experimental sites in summer and winter, and between those at the upper outer and lower outer canopy positions. Mean clone delta(13)C for the nursery hedges was only positively related to mean clone stem diameter at 1.3 m height at age 3 years on the wetter site. The clone by site interaction for foliar delta(13)C at the upper outer canopy was significant only in summer. Overall, the relatively high genetic variance components for foliar delta(13)C and significant, positive correlations between clone means of foliar delta(13)C and tree growth have highlighted the potential of using foliar delta(13)C measurements for assisting in selection of the elite F-1 hybrid pine clones with improved tree growth. (C) 2002 Elsevier Science B.V. All rights reserved.