Synthesis, characterization and kinetic study of silver and gold nanoparticles produced by the archaeon Haloferax volcanii.

AIMS: To evaluate the ability of the haloarchaeon Haloferax volcanii to produce Ag and Au nanoparticles (NPs) and to characterize the obtained material in order to find relevant properties for future potential applications. METHODS AND RESULTS: Nanoparticles were produced by incubating H. volcanii cells with the corresponding metal salt. In the presence of precursor salts, cultures evidenced a colour change associated to the formation of metallic nanostructures with plasmonic bands located in the visible range of the spectrum. X-ray fluorescence analysis confirmed the presence of Ag and Au in the NPs which were spherical, with average sizes of 25 nmol l-1 (Ag) and 10 nmol l-1 (Au), as determined by electronic microscopy. Fourier transformed infrared spectroscopy indicated that both types of NPs showed a stable protein capping. Ag NPs evidenced antibacterial activity and Au NPs improved the specificity of polymerase chain reaction reactions. Au and Ag NPs were able to reduce 4-nitrophenol when incubated with NaBH4 . CONCLUSIONS: Haloferax volcanii is able to synthesize metallic NPs with interesting properties for technological applications. SIGNIFICANCE AND IMPACT OF THE STUDY: Our data demonstrate the ability of H. volcanii to synthesize metal NPs and constitutes a solid starting point to deepen the study and explore novel applications.

Clinical profile and microbiological aetiology diagnosis in adult patients hospitalized with community-acquired pneumonia.

Early introduction of appropriate antibiotherapy is one of the major prognostic-modifying factors in community acquired pneumonia (CAP). Despite established guidelines for empirical therapy, several factors may influence etiology and, consequently, antibiotic choices. The aims of this study were to analyze the etiology of CAP in adults admitted to a northern Portugal University Hospital and evaluate the yield of the different methods used to reach an etiological diagnosis, as well as analyze of the impact of patient demographic and clinical features on CAP etiology. We retrospectively analyzed 1901 cases of CAP with hospitalization. The diagnostic performance increased significantly when blood and sputum cultures were combined with urinary antigen tests. The most frequent etiological agent was Streptococcus pneumoniae (45.7%), except in August, when it was overtaken by gram-negative bacilli (GNB) and Legionella pneumophila infections. Viral infections were almost exclusive to winter and spring. A negative microbiological result was associated with increasing age, non-smoking and lack of both blood/sputum cultures. Younger age was a predictor for S. pneumoniae, Influenza and L. pneumophila infections. Active smoking without any previously known respiratory disease was a risk factor for legionellosis. COPD was associated with Haemophilus influenzae cases, while dementia was typical in GNB and S. aureus patients. Diabetes mellitus (DM) and heart disease were negative predictors of S. pneumoniae and H. influenzae, respectively. P. aeruginosa was an independent risk factor for mortality (OR 13.02, 95% CI 2.94-57.7). This study highlights the importance of a comprehensive microbiological diagnostic workup and provides clues to predicting the most probable CAP causative agents, based on a patient's clinical profile. These may be taken into account when establishing first line antibiotherapy.

The Occurrence of Apparent Competition and Apparent Mutualism in a Modeled Greenhouse System with Two Non-competing Pests and a Shared Biocontrol Agent.

This work puts forward a dynamical population model to qualitatively reproduce the phenomena of apparent competition and apparent mutualism found in an experiment with two arthropods being attacked by a predator in a context of pest biological control in greenhouse crops. The two agricultural pests consist of one species of thrips (Frankliniella occidentalis (Pergande 1895)) and one species of whiteflies (Trialeurodes vaporariorum Westwood, 1956), and the shared predator is a predatory mite (Amblyseius swirskii Athias-Herriot, 1962). The predatory mite is the biocontrol agent employed in order to achieve the biological control. The proposed model successfully reproduces this density mediated indirect interactions between pests when their carrying capacities are increased. Moreover, the pests' final population levels may depend on their initial densities and those of their predator. With these results, the proposed model may have the potential to assess whether these indirect pest interactions disrupt or enhance biological control. Additionally, it can also be used as an ancillary tool to theoretically assess the effects of pest biocontrol strategies in the referred experimental setup.

Dynamics of extinction in coupled populations of the flour beetle Tribolium castaneum.

In this study we analyzed the effect of migration on the persistence time of coupled local populations of Tribolium in different environments. Four treatments were set up to compare different levels of environmental heterogeneity. We established high, low, moderate, and no heterogeneity. These levels were estimated by the different amounts of food offered to each population. To investigate how risk spreading works, a stochastic model for two subpopulations was employed. The high heterogeneity treatment resulted in the longest persistence, even though survival analysis revealed no significant difference among treatments. The magnitude of differences in growth rates among subpopulations is probably associated with persistence.

Integrated Pest Management in a Predator-Prey System with Allee Effects.

A commonly used biocontrol strategy to control invasive pests with Allee effects consists of the deliberate introduction of natural enemies. To enhance the effectiveness of this strategy, several tactics of control of invasive species (e.g., mass-trapping, manual removal of individuals, and pesticide spraying) are combined so as to impair pest outbreaks. This combination of strategies to control pest species dynamics are usually named integrated pest management (IPM). In this work, we devise a predator-prey dynamical model in order to assess the influence of the intensity of chemical killing on the success of an IPM. The biological and mathematical framework presented in this study can also be analyzed in the light of species conservation and food web dynamics theory.

Action of metalloproteinases mutalysin I and II on several components of the hemostatic and fibrinolytic systems.

The zinc endopeptidases mutalysin I (100 kDa) and mutalysin II (22.5 kDa) have been previously isolated from bushmaster (Lachesis muta muta) snake venom. Hemorrhagic activity was observed with as little as 0.5 microg (2000 units/mg) and 17.8 microg (56.2 units/mg) for mutalysin I and II, respectively. Additionally, the proteases hydrolyse the Aalpha>Bbeta chain of fibrinogen without clot formation. The specific fibrinogenolytic activity was estimated as 5. 25 and 16.3 micromol fibrinogen/min/micromol protein for mutalysin I and II, respectively. In vitro, the enzymes act directly on fibrin and are not inhibited by serine proteinase inhibitors (SERPINS). Analysis by SDS-PAGE of fibrin hydrolysis by both enzymes showed that mutalysin II (0.22 microM) completely digested the alpha- and gamma-gamma chains and partially the beta-chain (in 120 min incubation). In contrast, mutalysin I (three fold higher concentration than mutalysin II) hydrolyzed selectively the alpha-chain of fibrin leaving the beta and gamma-gamma chains unaffected. Unlike with the plasminogen activator-based thrombolytic agents (e.g., streptokinase), mutalysins do not activate plasminogen. Neither enzyme had an effect on protein C activation. Mutalysin II does not inhibit platelet aggregation in human PRP induced by collagen or ADP. However, mutalysin I showed a selective inhibitory effect on collagen-induced aggregation of human PRP; it did not affect platelet aggregation with ADP as the agonist. The present investigation demonstrates that both native and EDTA-inactivated mutalysin I dose dependently blocked aggregation of human PRP elicited by 10 microg/mL of collagen with an IC(50) of 180 and 580 nM, respectively. These studies suggest that, in addition to the metalloprotease region of mutalysin I, the disintegrin-like domain also participates in the inhibitory effect. The proteolytic activity of mutalysin II against dimethylcasein and fibrin was completely abolished by alpha2-macroglobulin (alpha2-M). The stoichiometry of inhibition was 1.0 mol of enzyme per mol of alpha2-M. In contrast, the proteolytic effect of mutalysin I against the same substrates was not significantly inhibited by alpha2-M. Therefore, the data explain why mutalysin I contributes significantly not only to local but also to systemic bleeding associated with the observed pathological effects of the venom.

Molecular cloning of a gamma-phospholipase A2 inhibitor from Lachesis muta muta (the bushmaster snake).

Several endogenous phospholipase A(2) inhibitors (PLIs) have been purified from the blood plasma of a number of snake species and are classified into three classes (alpha, beta and gamma) according to their structure and specificity. In the present study we have cloned transcripts of a protein homologous to CNF, a gammaPLI present in Crotalus durissus terrificus plasma, that is encoded in the liver of Lachesis muta muta (the bushmaster snake), a species evolutionarily related to Crotalus. The cDNA sequences code for two isoforms of a 200-residue protein including a 19-residue signal peptide followed by 181 amino acid residues in the mature form and a putative N-linked carbohydrate site. The deduced primary structures and some properties of those new proteins were compared to those of CNF. Multiple alignment was performed with the aminoacid sequences of all the gammaPLIs described so far and this used in the construction of a phylogenetic tree.

Characterization of a hemorrhagic factor, LHF-I, isolated from the bushmaster snake (Lachesis muta muta) venom.

Hemorrhagic factor I (LHF-I) was previously purified from the venom of the bushmaster snake (Lachesis muta muta). In terms of biochemical and immunological properties, LHF-I is a glycoprotein (mol. wt 100,000, pI 4.7) consisting of two subunits; it loses its activity following mercaptoethanol treatment. LHF-I contains 0.7 g-atom zinc and 1.2 g-atom calcium per mole protein. The hemorrhagic and the proteinase activities are inhibited by EDTA; subsequent addition of Ca2+ or Mg2+ does not reverse the EDTA-induced inhibition of the hemorrhagic activity. The metalloenzyme does not hyrolyze arginine esters and is devoid of phospholipase A2 activity. It hydrolyzes the A alpha- > B beta-chain of fibrinogen without clot formation and hydrolyzes selectively the alpha-chain of fibrin, leaving the B beta- and tau-chains unaffected. Antibodies to the hemorrhagic factor in bushmaster venom were produced by immunizing rabbits with the purified protein. The antibody was purified by protein-A affinity chromatography. This antibody was also used to screen other Crotalinae venom samples for immunologically similar epitopes by ELISA assay. The purified antibody reacted only with LHF-I and two samples of bushmaster venom from different geographical locations.

Neutralizing potency of horse antibothropic Brazilian antivenom against Bothrops snake venoms from the Amazonian rain forest.

Neutralization of lethal toxicity (50% effective dose; ED(50)), hemorrhagic (minimum hemorrhagic dose; MHD) and hemolytic activity (PLA(2)) and levels of antibodies, measured by enzyme-linked immunosorbent assay (ELISA), were investigated to test the potency of horse antibothropic serum (ABS) against Bothrops venoms from the Amazonian rain forest. ABS neutralized the lethal activity with a potency (mg of venom neutralized per 1 ml of antivenom) of 5.5, 3.7, 1.6, 1.3 and 6.5, respectively, for B. jararaca (reference venom for assessing the ABS potency in Brazil), B. atrox, B. brazili, B. bilineatus smaragdinus and B. taeniatus venoms. The volume of antivenom (microl) that neutralized one MHD of B. jararaca, B. atrox, B. brazili, B. bilineatus smaragdinus and B. taeniatus venoms was 5, 7.71, 7.76, 8.3 and 5, respectively. ABS neutralized the PLA(2) activity with a potency of 6.2, 3.2, 1.4, 2.6 and 5 respectively, for B. jararaca, B. atrox, B. brazili, B. bilineatus smaragdinus and B. taeniatus venoms. ELISA reactivity of ABS against the separate venoms was found to be quite variable. The reactivity against B. jararaca venom was higher than against other Bothrops venoms. In conclusion, the assays described here suggest that Brazilian Bothrops polyspecific antivenom is not very efficient in neutralizing the effects of venom from some Amazonian Bothrops species.

Neutralization of the hemorrhagic activity of Bothrops and Lachesis snake venoms by a monoclonal antibody against mutalysin-II.

One mAb reactive with mutalysin-II, a hemorrhagic metalloproteinase isolated from Lachesis muta muta venom, was produced in mice immunized with L. m. muta venom. Indirect ELISA was employed to compare the antigenic cross-reactivity among the venoms from Bothrops snakes. The mAb anti-mutalysin-II efficiently neutralized the hemorrhagic effect of both mutalysin-II and L. m. muta crude venom. Furthermore, the mAb were cross-reactive with B. alternatus, B. atrox, B. itapetiningae, B. jararaca and B. neuwiedii and showed variable potencies in neutralizing the hemorrhagic activity of several bothropic venoms.

Drug kinetics and drug resistance in optimal chemotherapy.

A system of differential equations for the control of tumor cells growth in a cycle nonspecific chemotherapy is presented. First-order drug kinetics and drug resistance are taken into account in a class of optimal control problems. The results show that the strategy corresponding to the maximum rate of drug injection is optimal for the Malthusian model of cell growth (which is a relatively good model for the initial phase of tumor growth). For more general models of cell growth, this strategy proved to be suboptimal under certain conditions.

Chemotherapeutic treatments involving drug resistance and level of normal cells as a criterion of toxicity.

A system of differential equations for the control of tumor cells growth in a cycle nonspecific chemotherapy is presented. Drug resistance and toxicity conveyed through the level of normal cells are taken into account in a class of optimal control problems. Alternative treatments for the exponential tumor growth are set forth for cases where optimal treatments are not available.

[Genetic and clinical diagnosis of infantile autism]. / Diagnóstico genético e clínico do autismo infantil.

The main objectives of this study were to characterize the selected variables for a better understanding and diagnosis of infantile autism such as clinical and image findings, diagnostic criteria, frequency of neuropsychiatric disorders in the subjects' families, familial recurrence and occurrence of consangunity between the subjects' parents and between other couples in the family. The sample was composed of 36 subjects of both sexes, in the age group from 1 though 20 years old, members of 35 distinctive families, all of which presenting clinical diagnosis for infantile autism. Mental retardation was clinically observed in all subjects of the sample and convulsion in 27.8%; neuropsychiatric disorders were referred in at least one family member of the subjects (97.14% of the families); recurrent autism in 11.42% grandparents and great-grand-parents (2.86%); abnormal findings in computed tomography scan were observed in three subjects. These results support the suggestion of the multifactorial heredity model with differential threshold fold sex in infantile autism. The clinical evaluation of all infantile autism cases should always appraise the neurological, psychiatric and genetic features.

[Mushroom poisoning in Portugal]. / Intoxicação por cogumelos em Portugal.

The renewed interest in mycology has been reflected in growing use of wild mushrooms in culinary, driven by its nutritional, organoleptic and commercial value. However, the international scientific literature describes several syndromes of poisoning by mushrooms. We live, therefore, a paradigm conducive to an increase of mycetism, whose diagnosis requires a high level of suspicion and knowledge of clinical profiles. In Portugal, the real dimension of this problem is unknown. Although some mycetisms, such as the hepatotoxic syndrome, have high morbidity and mortality, their relative incidences are unknown. Add up to the shortage of international scientific literature, often outdated and inappropriate to clinical practice. In this context, this article provides an updated epidemiological and clinical perspective emphasizing a narrative and descriptive information on the forms of presentation, differential diagnosis and therapeutic approach, with the ultimate goal of the elaboration of a national diagram-oriented approach to decision-making diagnosis. We analyzed all the clinical records of patients admitted into ten hospitals between 1990 and 2008, notified with the code 988.1 of GDH (acute poisoning by mushrooms). There were registered demographic data, way of presentation, time between ingestion and onset of symptoms, the annual distribution, clinical profile, clinical and analytical treatment performed and complications. We identified 93 cases of acute poisoning by mushrooms, with equal gender distribution and inclusion of individuals of all age groups (from 1 to 85 years), but with greater representation from 21 to 50 years. There was a bimodal seasonal pattern, with a higher peak between September and December and a second in the spring. The hepatotoxic profile presentation corresponded to 63.4% and 31.7% of the cases to gastroenteritis syndrome. The mortality in cases of hepatotoxicity was 11.8%. The developmental profile of the rate of prothrombin time (PT), aspartate aminotransferase (AST) and bilirubin, was an important setting for diagnosis and prognosis. TGO increases early, always within 48 hours, having an essential role in the diagnosis of hepatotoxicity. Despite the late elevation of bilirubin, the cases of death revealed that there was an earlier increase, reaching higher values, which seems to have a prognostic value, to be evaluated with further studies. Finally, we propose a diagram of diagnostic performance, considerating the generalized lack of mycological diagnosis in Portugal, which emphasizes the need for a careful history, focused on quantifying the latency period.

Population dynamics, life stage and ecological modeling in Chrysomya albiceps (Wiedemann) (Diptera: Calliphoridae).

In this study we investigated the population dynamics of Chrysomya albiceps (Wiedemann) with laboratory experiments, employing survival analysis and stage structure mathematical models, emphasizing survival among life stages. The study also assessed the theoretical influence of density dependence and cannibalism during immature stages, on the population dynamics of the species. The survival curves were similar, indicating that populations of C. albiceps exhibit the same pattern of survival among life stages. A strong nonlinear trend was observed, suggesting density dependence, acting during the first life stages of C. albiceps. The time-series simulations produced chaotic oscillations for all life stages, and the cannibalism did not produce qualitative changes in the dynamic behavior. The bifurcation analysis shows that for low values for survival, the population reaches a stable equilibrium, but the cannibalism results in chaotic oscillations practically over all the parametric space. The implications of the patterns of dynamic behavior observed are discussed.

Catalytic Reduction of Noble Metal Salts by Sodium Hypophosphite Promoted by the Film Poly-(p-Allyl Ether Benzenesulfonic Acid).

Glassy carbon electrodes were coated with the film poly-(p-allyl ether benzenesulfonic acid) by an anodic procedure. Nickel, platinum, and palladium ions were introduced into the film by ion exchange of H(+) with the corresponding salts. These ions were catalytically reduced to their corresponding metals using the known electroless reducing agent sodium hypophosphite. Scanning electron microcopy and energy dispersive X-ray spectroscopy were carried out to demonstrate the occurrence of the catalytic process. To compare this method with another one carried out in our laboratory, the electrocatalytic reduction of H(+) was studied using the same modified electrodes. A suggested mechanism for the catalysis is proposed.

A correct formulation for a spatially implicit predator-prey metacommunity model.

In order to mitigate the problem of increasing model complexity with increasing number of occupation states in spatially implicit metacommunity models, the assumption of independency among species distributions is often required. In the present paper, we show that this approach only works correctly if set relations among patch occupancy states are considered adequately. This is illustrated by means of a well-known, although incorrectly formulated, predator-prey metacommunity model devised by Bascompte and Solé[1]. We demonstrate that this model shows anomalous dynamical behavior caused by inconsistence between the model formulation and its assumptions. In order to formalize our finding we develop a corrected model formulation that accounts for the principles of set theory so that the sum of the system compartments change rate is nulled. Applying this method successfully rules out the occurrence of anomalous dynamical behavior found in the original model. Finally we discuss the implications of our findings for the accuracy of model predictions.

The interplay among prey preference, nutrient enrichment and stability in an omnivory system.

Food webs usually display an intricate mix of trophic interactions where multiple prey are common. In this context omnivory has been the subject of intensive analysis regarding food web stability and structure. In a three species omnivory setting it is shown that the modeling of prey preference by the top predator may exert a strong influence on the short as well as on the long term dynamics of the respective food web. Clearly, this has implications concerning the stability and the structure of omnivory systems under disturbances such as nutrient enrichment.

Conflicting objectives in chemotherapy with drug resistance.

A system of differential equations for the control of tumor cells growth in a cycle nonspecific chemotherapy is presented. Spontaneously acquired drug resistance is accounted for, as well as the evolution in time of normal cells. In addition, optimization of conflicting objectives forms the aim of the chemotherapeutic treatment. For general cell growth, some results are given, whereas for the special case of Malthusian (exponential) growth of tumor cells and rather general growth rate for normal cells, the optimal strategy is worked out. The latter, from the clinical standpoint, corresponds to maximum drug concentration throughout the treatment.

Therapy burden, drug resistance, and optimal treatment regimen for cancer chemotherapy.

Three nonlinear models of tumour cell growth under continuous delivery of cycle nonspecific anticancer agents are studied. A dynamical optimization problem with the objective of minimizing the final level of tumour cells is posed for these mathematical setups. The simplest setup does not possess toxicity constraints, whereas the other setups contain a dynamical equation describing the therapy burden as a toxicity criterion. In addition, the third setting contains the dynamics of drug resistant cells. A discussion concerning the optimal strategies of the respective models is performed.