Assessing three industrially produced fungi for the bioremediation of diclofenac.

Diclofenac is an emerging pollutant: toxic, persistent, and bioaccumulative, present in several environmental niches in a concentration of parts per million. This pharmaceutical's biological removal was reported with various fungal species, showing promissory results. This work aimed at diclofenac removal by individually challenging the fungal species Pleurotus ostreatus, Aspergillus niger, and Penicillium roquefortii but triying to lower the biosorption nature of cell walls by NaCl addition. P. ostreatus removed 100% of the initial diclofenac concentration, whereas A. niger and P. roqueforti removed 74% and 32%, respectively. In all three cases, biosorption by polar interactions was negligible. We demonstrated that stressful environments, such as mineral media, force the fungus to take advantage of its metabolic tools to survive, hence showing higher removal capacity when limiting growth conditions. Bioremediation is an excellent alternative to give residual fungal biomass a secondary use.

A transfer learning approach for predictive modeling of bioprocesses using small data.

Predictive modeling of new biochemical systems with small data is a great challenge. To fill this gap, transfer learning, a subdomain of machine learning that serves to transfer knowledge from a generalized model to a more domain-specific model, provides a promising solution. While transfer learning has been used in natural language processing, image analysis, and chemical engineering fault detection, its application within biochemical engineering has not been systematically explored. In this study, we demonstrated the benefits of transfer learning when applied to predict dynamic behaviors of new biochemical processes. Two different case studies were presented to investigate the accuracy, reliability, and advantage of this innovative modeling approach. We thoroughly discussed the different transfer learning strategies and the effects of topology on transfer learning, comparing the performance of the transfer learning models against benchmark kinetic and data-driven models. Furthermore, strong connections between the underlying process mechanism and the transfer learning model's optimal structure were highlighted, suggesting the interpretability of transfer learning to enable more accurate prediction than a naive data-driven modeling approach. Therefore, this study shows a novel approach to effectively combining data from different resources for bioprocess simulation.

Kinetic and hybrid modeling for yeast astaxanthin production under uncertainty.

Astaxanthin is a high-value compound commercially synthesized through Xanthophyllomyces dendrorhous fermentation. Using mixed sugars decomposed from biowastes for yeast fermentation provides a promising option to improve process sustainability. However, little effort has been made to investigate the effects of multiple sugars on X. dendrorhous biomass growth and astaxanthin production. Furthermore, the construction of a high-fidelity model is challenging due to the system's variability, also known as batch-to-batch variation. Two innovations are proposed in this study to address these challenges. First, a kinetic model was developed to compare process kinetics between the single sugar (glucose) based and the mixed sugar (glucose and sucrose) based fermentation methods. Then, the kinetic model parameters were modeled themselves as Gaussian processes, a probabilistic machine learning technique, to improve the accuracy and robustness of model predictions. We conclude that although the presence of sucrose does not affect the biomass growth kinetics, it introduces a competitive inhibitory mechanism that enhances astaxanthin accumulation by inducing adverse environmental conditions such as osmotic gradients. Moreover, the hybrid model was able to greatly reduce model simulation error and was particularly robust to uncertainty propagation. This study suggests the advantage of mixed sugar-based fermentation and provides a novel approach for bioprocess dynamic modeling.

Increased removal of cadmium by Chlamydomonas reinhardtii modified with a synthetic gene for γ-glutamylcysteine synthetase.

Bioremediation with genetically modified microalgae is becoming an alternative to remove metalloids and metals such as cadmium, a contaminant produced in industrial processes and found in domestic waste. Its removal is important in several countries including Mexico, where the San Luis Potosi region has elevated levels of it. We generated a construct with a synthetic gene for γ-glutamylcysteine synthetase and employed it in the chloroplast transformation of Chlamydomonas reinhardtii. In dose-response kinetics with media containing from 1 to 20 mg/L of cadmium, both the transplastomic clone and the wild-type strain grew similarly, but the former removed up to 32% more cadmium. While the growth of both decreased with higher concentrations of cadmium, the transplastomic clone removed 20 ± 9% more than the wild-type strain. Compared to the wild-type strain, in the transplastomic clone the activity of glutathione S-transferase and the intracellular glutathione increased up to 2.1 and 1.9 times, respectively, in media with 2.5 and 10 mg/mL of cadmium. While 20 mg/L of cadmium inhibited the growth of both, the transplastomic clone gradually duplicated. These results confirm the expression of the synthetic gene gshA in the transformed strain as revealed in its increased removal uptake and metabolic response.

Designing Microflowreactors for Photocatalysis Using Sonochemistry: A Systematic Review Article.

Use of sonication for designing and fabricating reactors, especially the deposition of catalysts inside a microreactor, is a modern approach. There are many reports that prove that a microreactor is a better setup compared with batch reactors for carrying out catalytic reactions. Microreactors have better energy efficiency, reaction rate, safety, a much finer degree of process control, better molecular diffusion, and heat-transfer properties compared with the conventional batch reactor. The use of microreactors for photocatalytic reactions is also being considered to be the appropriate reactor configuration because of its improved irradiation profile, better light penetration through the entire reactor depth, and higher spatial illumination homogeneity. Ultrasound has been used efficiently for the synthesis of materials, degradation of organic compounds, and fuel production, among other applications. The recent increase in energy demands, as well as the stringent environmental stress due to pollution, have resulted in the need to develop green chemistry-based processes to generate and remove contaminants in a more environmentally friendly and cost-effective manner. It is possible to carry out the synthesis and deposition of catalysts inside the reactor using the ultrasound-promoted method in the microfluidic system. In addition, the synergistic effect generated by photocatalysis and sonochemistry in a microreactor can be used for the production of different chemicals, which have high value in the pharmaceutical and chemical industries. The current review highlights the use of both photocatalysis and sonochemistry for developing microreactors and their applications.

CTLA4Fcε, a novel soluble fusion protein that binds B7 molecules and the IgE receptors, and reduces human in vitro soluble CD23 production and lymphocyte proliferation.

Immunoglobulin E-mediated allergy and certain autoimmune diseases are characterized by the presence of a T helper type 2 (Th2) immune response and allergen-specific or self-reactive IgE. Soluble CD23 (sCD23) is a B-cell factor that fosters IgE class-switching and synthesis, suggesting that sCD23 may be a therapeutic target for these pathologies. We produced a recombinant protein, CTLA4Fcε, by fusing the ectodomain of the immunoregulatory molecule cytotoxic T-lymphocyte antigen 4 (CTLA-4) with a fragment of the IgE H-chain constant region. In SDS-PAGE/inmunoblot analyses, CTLA4Fcε appeared as a 70,000 MW polypeptide that forms homodimers. Flow cytometry showed that CTLA4Fcε binds to IgE receptors FcεRI and FcεRII/CD23, as well as to CTLA-4 counter-receptors CD80 and CD86. Binding of CTLA4Fcε to FcεRII/CD23 appeared stronger than that of IgE. Since the cells used to study CD23 binding express CD80 and CD86, simultaneous binding of CTLA4Fcε to CD23 and CD80/CD86 seems to occur and would explain this difference. As measured by a human CD23-specific ELISA, CTLA4Fcε - but not IgE - induced a concentration-dependent reduction of sCD23 in culture supernatants of RPMI-8866 cells. Our results suggest that the simultaneous binding of CTLA4Fcɛ to CD23-CD80/CD86 may cause the formation of multi-molecular complexes that are either internalized or pose a steric hindrance to enzymatic proteolysis, so blocking sCD23 generation. CTLA4Fcε caused a concentration-dependent reduction of lymphocyte proliferation in human peripheral blood mononuclear cell samples stimulated in vitro with concanavalin A. The ability to bind IgE receptors on effector cells, to regulate the production of sCD23 and to inhibit lymphocyte proliferation suggests that CTLA4Fcɛ has immunomodulatory properties on human Th2 responses.

Removal and accumulation of As, Cd and Cr by Typha latifolia.

The removal from the solution and the accumulation of As, Cd and Cr by Typha latifolia was studied. Small plants of T. latifolia, collected from a non-contaminated site, were exposed to individual concentrations of As, Cd and Cr for 10 days. The ability of T. latifolia for the removal of toxic elements ranged from 23% to 54% for As, 43%-55% for Cd and 28%-73% for Cr. The accumulation of toxic elements in T. latifolia occurred mainly in the roots. The results suggest that T. latifolia can be considered as an interesting alternative for treating aquatic effluents polluted with toxic trace elements.

Tuning the physicochemical features of titanium oxide nanomaterials by ultrasound: Elevating photocatalytic selective partial oxidation of lignin-inspired aromatic alcohols.

The research for "green" and economically feasible approaches such as (photo)catalysis especially for biomass valorization such as selective oxidation of biomass derived compounds like aromatic alcohols to corresponding aldehyde by avoiding the harsh reaction conditions and the addition of reagents concentrate the focus of attention the last years. Hence, design and development of novel photocatalyst for the partial selective oxidation is highly desirable. In this research work, ultrasonication of different frequencies (22, 40, 80 kHz) and different amplitudes was utilized as synthesis tool in order to obtain novel materials by precipitation method. The synthesized samples were characterized by using different techniques such as N2 sorption, TEM, XPS, XRD, thermal analysis, and diffuse reflectance spectroscopy. The synthesized sample by using low ultrasound frequency (22 kHz) and amplitude showed a mixed morphological and structural nature consisting of asymmetric 1-dimensional (nanorods-like), layered nano-structures and not well-defined areas, leading to elevate for metal oxide specific surface areas up to 155 m2/g. The observed 1-D nanostructures have diamentions in the range of 20-60 nm. This sample revealed the highest photo-oxidation efficiency for the selective conversion of two biomass-derived, and more specifically lignin-inspired model compounds, benzyl alcohol and cinnamyl alcohol to benzaldehyde and cinnamyl aldehyde, respectively, and hence the highest yield towards the desired aldehydes. The selective photo-oxidation activity was retained even after 5 photocatalytic cycles, while no leaching of Ti was recorded.

Diclofenac removal by the microalgae species Chlorella vulgaris, Nannochloropsis oculata, Scenedesmus acutus, and Scenedesmus obliquus.

In this work, we evaluated the removal efficiency of diclofenac by Chlorella vulgaris OW-01, Nannochloropsis oculata CCAP 849/7, Scenedesmus acutus UTEX 72, and Scenedesmus obliquus CCAP 276/2. Each microalga was grown in media with different concentrations (50 and 100% of the original formulation) of carbon, nitrogen, and phosphorus, to evaluate their effect on the removal of diclofenac. We also evaluated the photodegradation of diclofenac under the same conditions. The diclofenac removed from the media ranged from 59 to 92%, obtaining the highest removal with S. obliquus. The diclofenac adsorbed on the cell walls ranged from 12.2 to 26.5%, obtaining the highest adsorption with S. obliquus. The diclofenac degraded by light ranged from 15 to 28%. The nutrient deficit showed no influence on the removal of diclofenac in any of the microalgae under study. These results indicate that S. obliquus is the best alternative for the bioremediation of diclofenac. Supplementary Information: The online version contains supplementary material available at 10.1007/s13205-022-03268-2.

Carbon-Based Nanocatalysts (CnCs) for Biomass Valorization and Hazardous Organics Remediation.

The continuous increase of the demand in merchandise and fuels augments the need of modern approaches for the mass-production of renewable chemicals derived from abundant feedstocks, like biomass, as well as for the water and soil remediation pollution resulting from the anthropogenic discharge of organic compounds. Towards these directions and within the concept of circular (bio)economy, the development of efficient and sustainable catalytic processes is of paramount importance. Within this context, the design of novel catalysts play a key role, with carbon-based nanocatalysts (CnCs) representing one of the most promising class of materials. In this review, a wide range of CnCs utilized for biomass valorization towards valuable chemicals production, and for environmental remediation applications are summarized and discussed. Emphasis is given in particular on the catalytic production of 5-hydroxymethylfurfural (5-HMF) from cellulose or starch-rich food waste, the hydrogenolysis of lignin towards high bio-oil yields enriched predominately in alkyl and oxygenated phenolic monomers, the photocatalytic, sonocatalytic or sonophotocatalytic selective partial oxidation of 5-HMF to 2,5-diformylfuran (DFF) and the decomposition of organic pollutants in aqueous matrixes. The carbonaceous materials were utilized as stand-alone catalysts or as supports of (nano)metals are various types of activated micro/mesoporous carbons, graphene/graphite and the chemically modified counterparts like graphite oxide and reduced graphite oxide, carbon nanotubes, carbon quantum dots, graphitic carbon nitride, and fullerenes.

Removal and accumulation of cadmium and lead by Typha latifolia exposed to single and mixed metal solutions.

We investigated the effect of Cd and Pb on the growth of the aquatic macrophyte Typha latifolia; the removal from the solution and the accumulation of these elements by the plant were also studied. Thus, small plants of T. latifolia, collected from a noncontaminated site, were exposed for 10 days to Cd and Pb, in a single solution or in mixture solutions, at two concentrations of the metals (5 and 7.5 mg/L). Our results showed that T. latifolia removed effectively Cd and Pb from solutions and was able to accumulate these metals in the roots and, to a lesser extent, in the leaves. Our findings suggested a synergistic effect of Cd and Pb with respect to the toxicity to T. latifolia. Additionally, Cd diminished the Pb absorption by T. latifolia. Our results confirmed, using scanning electron microscopy, the internalization of Cd and Pb in T. latifolia.

Inhibiting the stringent response blocks Mycobacterium tuberculosis entry into quiescence and reduces persistence.

The stringent response enables Mycobacterium tuberculosis (Mtb) to shut down its replication and metabolism under various stresses. Here we show that Mtb lacking the stringent response enzyme RelMtb was unable to slow its replication rate during nutrient starvation. Metabolomics analysis revealed that the nutrient-starved relMtb -deficient strain had increased metabolism similar to that of exponentially growing wild-type bacteria in nutrient-rich broth, consistent with an inability to enter quiescence. Deficiency of relMtb increased the susceptibility of mutant bacteria to killing by isoniazid during nutrient starvation and in the lungs of chronically infected mice. We screened a pharmaceutical library of over 2 million compounds for inhibitors of RelMtb and showed that the lead compound X9 was able to directly kill nutrient-starved M. tuberculosis and enhanced the killing activity of isoniazid. Inhibition of RelMtb is a promising approach to target M. tuberculosis persisters, with the potential to shorten the duration of TB treatment.

Perinatal risk factors for pneumothorax and morbidity and mortality in very low birth weight infants.

AIM: To determine the perinatal risk factors for pneumothorax in Very-Low-Birth-Weight (VLBW) infants and the associated morbidity and mortality in this population. METHODS: Retrospective analysis of data collected prospectively from a cohort of VLBW neonates assisted in our Unit (2006-2013). We included all consecutive in-born patients with ≤ 1500 g, without severe congenital anomalies. Perinatal history, demographics, interventions and clinical outcomes were collected. Associations were evaluated by logistic regression analysis. RESULTS: During the study period, 803 VLBW infants were assisted in our Unit, of whom 763 were inborn. Ten patients (1.2%) died in delivery room, and 18 (2.2%) with major congenital anomalies were excluded. Finally, 735 (91.5%) neonates were included in the study. Seventeen (2.3%) developed pneumothorax during the first week of life [median (IQR): 2 (1-2) days]. After correcting for GA and other confounders, prolonged rupture of membranes [aOR =1.002 (95% CI 1.000-1.003); p = 0.040] and surfactant administration [aOR = 6.281 (95% CI 1.688-23.373); p = 0.006] were the independent risk factors associated with pneumothorax. Patients with pneumothorax had lower probabilities of survival without major brain damage (MBD): aOR = 0.283 (95% CI = 0.095-0.879); p = 0.029. CONCLUSIONS: Pneumothorax in VLBW seems to be related to perinatal inflammation and surfactant administration, and it is significantly associated with a reduction in the probabilities of survival without MBD.

Utilization of substrate-induced quenching for screening targets promoting NADH and NADPH consumption.

Oxidation of reduced nicotinamide adenine dinucleotides is a common event for many biochemical reactions. However, its exploitation for ultrahigh-throughput screening purposes is not an easy task and is affected by various drawbacks. It is known that such nucleotides induce quenching on the fluorescence of several dyes and that this quenching disappears with oxidation of the nucleotide. We have made use of this property to develop an assay for high-throughput screening with NADH and NADPH-dependent reductases. Full screening campaigns have been run with excellent assay quality parameters, and interesting hits have been identified. The method is amenable to miniaturization and allows easy identification of false positives without needing extra secondary assays. Although it is based on monitoring substrate consumption, it is demonstrated that the effect of fractional conversion on assay sensitivity is negligible.

Discovery of Novel Inhibitors of the Tautomerase Activity of Macrophage Migration Inhibitory Factor (MIF).

Macrophage migration inhibitory factor (MIF) is a proinflammatory cytokine associated with multiple diseases, including neurodegenerative disorders. With the ultimate goal of providing novel chemotypes as starting points for development of disease-modifying therapeutics for neurodegeneration, we endeavored to screen the GSK compound collection for MIF inhibitors using a miniaturized, activity-based kinetic assay. The assay monitors the increase in absorbance at 320 nm resulting from keto-to-enol tautomerization of 4-hydroxyphenylpyruvate, a reaction catalyzed by MIF. We ran a full-diversity screen evaluating the inhibitory activity of 1.6 million compounds. Primary hits were confirmed and retested in an orthogonal assay measuring tautomerization of l-dopachrome methyl ester by the decrease in absorbance at 475 nm in kinetic mode. Selected compounds were progressed to medium-throughput mode-of-inhibition studies, which included time dependence, enzyme concentration dependence, and reversibility of their inhibitory effect. With these results and after inspection of the physicochemical properties of compounds, 17 chemotypes were prioritized and progressed to further stages of validation and characterization to better assess their therapeutic potential.

Discovery of Enhancers of the Secretion of Leukemia Inhibitory Factor for the Treatment of Multiple Sclerosis.

Multiple sclerosis (MS) is an autoimmune neurodegenerative disease that involves activation of T cells, microglia, and astrocytes. There is a clear unmet medical need for MS, as current therapies reduce the relapse rate, but are unable to prevent the neurological deterioration. Leukemia inhibitory factor (LIF) is a proinflammatory cytokine that can also positively modulate the immune response, by inducing the inhibition of myelin-reactive TH17 differentiation, and by promoting oligodendrocyte-mediated myelination. The aim of this project was to find central nervous system (CNS)-permeable and orally available small molecules that upregulate production of endogenous LIF. We describe here the development of a phenotypic assay and screening of 1.7 million compounds to identify LIF enhancers using U87 MG cells. Five chemically tractable series of compounds and a few singletons were selected for further progression. Some of them were also active in a different LIF-expressing cell line and in primary rat astrocytes. Although further studies would be required to deconvolute the targets involved in LIF induction and to confirm activity of hits in more disease-relevant assays, our results have demonstrated the potential of the phenotypic approach to identify specific and chemically tractable small molecules that trigger the production of LIF in relevant cell lines.

Novel inhibitors of Mycobacterium tuberculosis GuaB2 identified by a target based high-throughput phenotypic screen.

High-throughput phenotypic screens have re-emerged as screening tools in antibiotic discovery. The advent of such technologies has rapidly accelerated the identification of 'hit' compounds. A pre-requisite to medicinal chemistry optimisation programmes required to improve the drug-like properties of a 'hit' molecule is identification of its mode of action. Herein, we have combined phenotypic screening with a biased target-specific screen. The inosine monophosphate dehydrogenase (IMPDH) protein GuaB2 has been identified as a drugable target in Mycobacterium tuberculosis, however previously identified compounds lack the desired characteristics necessary for further development into lead-like molecules. This study has identified 7 new chemical series from a high-throughput resistance-based phenotypic screen using Mycobacterium bovis BCG over-expressing GuaB2. Hit compounds were identified in a single shot high-throughput screen, validated by dose response and subjected to further biochemical analysis. The compounds were also assessed using molecular docking experiments, providing a platform for their further optimisation using medicinal chemistry. This work demonstrates the versatility and potential of GuaB2 as an anti-tubercular drug target.

Utilización de un dispositivo móvil de tecnología electrónica para tamizaje de fibrilación auricular: estudio piloto / Use of a mobile electronic technology device for atrial fibrillation screening: pilot study / Uso de dispositivo de tecnologia eletrônica móvel para rastreamento de fibrilação atrial: estudo piloto

La fibrilación auricular (FA) es un problema de salud pública que genera mortalidad y morbilidad, con su mayor impacto en mayores de 65 años. Su detección es especialmente relevante para la población de riesgo. Este estudio piloto propone valorar la utilidad de un dispositivo móvil de tecnología electrónica (DMTE) para el tamizaje de FA. Objetivo primario: evaluar la validez y confiabilidad de un DMTE para identificar FA. Objetivo secundario: validar los procesos de recolección, transmisión, almacenamiento, procesamiento e interpretación de los datos obtenidos. Método: los asistentes a 5 talleres del Plan Ibirapitá (entre el 15 de octubre y el 30 de noviembre de 2018) fueron invitados a participar del estudio. Se utilizó un DMTE (sensor KardiaMobile de AliveCor®) cuyo registro se contrastó con el trazado del ECG de 12 derivaciones realizado en forma casi simultánea. Se compararon los diagnósticos automáticos con los realizados por dos cardiólogos expertos de forma independiente, a partir de la lectura del registro realizado con el DMTE y de un ECG de 12 derivaciones. Resultados: participaron del estudio 114 beneficiarios del Plan Ibirapitá, 78 del sexo femenino. La edad fue de 72,5 ± 5,36 años (rango: 53-87 años). La sensibilidad para el diagnóstico de FA del DMTE fue de 100%, con una especificidad de 96,6%. (VPP = 57,1% VPN = 100%) y una proporción de diagnóstico correcto de 96,8%. El DMTE catalogó como "sin clasificar" al 18,4% de los trazados. Considerando esto, la proporción de diagnóstico correcto disminuyó a 78,9%, sin presentar falsos negativos. Conclusiones: el cribado de FA con la utilización de un DMTE en una población de adultos mayores es factible y confiable. El hallazgo de un 18,4% de trazados con diagnóstico automático "sin clasificar" hace necesario contar con recursos humanos calificados para realizar la confirmación diagnóstica en esos registros.

Atrial fibrillation (AF) is a public health problem generating important morbidity and mortality mainly in > 65 years old population. Detection is key in the population at risk. This pilot study was designed to assess a mobile electronic technology device (METD) usefulness in AF screening. Objective: evaluate validity and reliability of a METD in AF identification. Secondary objective: to validate the process of collection, transmission, storage, method and interpretation of obtained data. Method: participants in 5 Ibirapitá Plan workshops (October 15-November 30, 2018) were invited to participate in the study. A KardiaMobile Alive Cor® METD was used, whose ECG recording was compared with a 12L ECG taken almost at the same time. Automatic METD report was compared with 2 independent cardiologists report considering the METD recording and the 12L ECG. Results: 114 participants (78 female), mean age 72,5 y.o. (SD 5,36; range 53-87) were included. METD sensitivity for AF diagnosis was 100% with a 96.6% specificity (PPV=57,1% NPV=100%), and a 96.8% number of correct diagnosis. A 18.4% of recordings were catalogued as "unclassified" by the METD. Considering this, the proportion of correct diagnosis decreased to 78.9%; there were not false negatives. Conclusions: AF screening with a METD in an older population is feasible and reliable. The finding of 18.4% METD recordings as "unclassified" raises the needs for experts review during diagnosis confirmation.

A fibrilação atrial (FA) é um problema de saúde pública que gera mortalidade e morbidade, com maior impacto em pessoas com mais de 65 anos. Sua detecção é especialmente relevante para a população de risco. Este estudo piloto teve como objetivo avaliar a utilidade de um dispositivo móvel de tecnologia eletrônica (DMTE) para o monitoramento da FA. Objetivo principal: avaliar a validade e confiabilidade de um DMTE para identificar FA. Objetivo secundário: validar os processos de coleta, transmissão, armazenamento, processamento e interpretação dos dados obtidos. Método: participantes de 5 oficinas do Plano Ibirapitá (entre 15 de outubro e 30 de novembro de 2018) foram convidados a participar do estudo. Foi utilizado um DMTE (sensor AliveCor® KardiaMobile), cujo registro foi contrastado com o traçado do ECG de 12 derivações realizado quase simultaneamente. Os diagnósticos automatizados foram comparados com aqueles realizados por dois cardiologistas especialistas independentes, com base na leitura gráfica do DMTE e no ECG de 12 derivações. Resultados: participaram do estudo 114 beneficiários do Plano Ibirapitá, sendo 78 mulheres. A média de idade foi de 72,5 anos (DP 5,36; variação de 53-87 anos). A sensibilidade para o diagnóstico de FA no DMTE foi de 100% com especificidade de 96,6%. (VPP = 57,1% VPN = 100%) e proporção de diagnóstico correto de 96,8%. O DMTE definiu 18,4% dos registros como "não classificados". Portanto, a proporção de diagnósticos corretos diminuiu para 78,9% e não houve falsos negativos. Conclusões: a triagem para FA por meio de DMTE em uma população idosa é viável e confiável. A constatação de 18,4% dos registros com diagnóstico automático "não classificado" torna necessária a existência de recursos humanos qualificados para a realização da confirmação diagnóstica.

Inhibition of polyamine oxidase activity affects tumor development during the maize-Ustilago maydis interaction.

Ustilago maydis is a biotrophic plant pathogenic fungus that leads to tumor development in the aerial tissues of its host, Zea mays. These tumors are the result of cell hypertrophy and hyperplasia, and are accompanied by the reprograming of primary and secondary metabolism of infected plants. Up to now, little is known regarding key plant actors and their role in tumor development during the interaction with U. maydis. Polyamines are small aliphatic amines that regulate plant growth, development and stress responses. In a previous study, we found substantial increases of polyamine levels in tumors. In the present work, we describe the maize polyamine oxidase (PAO) gene family, its contribution to hydrogen peroxide (H2O2) production and its possible role in tumor development induced by U. maydis. Histochemical analysis revealed that chlorotic lesions and maize tumors induced by U. maydis accumulate H2O2 to significant levels. Maize plants inoculated with U. maydis and treated with the PAO inhibitor 1,8-diaminooctane exhibit a notable reduction of H2O2 accumulation in infected tissues and a significant drop in PAO activity. This treatment also reduced disease symptoms in infected plants. Finally, among six maize PAO genes only the ZmPAO1, which encodes an extracellular enzyme, is up-regulated in tumors. Our data suggest that H2O2 produced through PA catabolism by ZmPAO1 plays an important role in tumor development during the maize-U. maydis interaction.

L-cystine inhibits aspartate-beta-semialdehyde dehydrogenase by covalently binding to the essential 135Cys of the enzyme.

Aspartate-beta-semialdehyde dehydrogenase (ASADH) from Escherichia coli is inhibited by L- and D-cystine, and by other cystine derivatives. Enzyme inhibition is quantitatively reversed by addition of dithiothreitol (DTT), dithioerythrytol, beta-mercaptoethanol, di-mercaptopropanol or glutathione to the cystine-inactivated enzyme. Cystine labeling of the enzyme is a pH dependent process and is optimal at pH values ranging from 7.0 to 7.5. Both the cysteine incorporation profile and the inactivation curve of the enzyme as a function of pH suggest that a group(s) with pK(a) of 8.5 could be involved in cystine binding. Stoichiometry of the inactivation reaction indicates that one cysteine residue from the enzyme subunit is reactive against cystine, as found by direct incorporation of radioactive cystine into the enzyme and by free-thiol titration of the enzyme with 5,5'-dithiobis-2-nitrobenzoic acid (DTNB) before and after the cystine treatment. One mole of cysteine is released from each mol of cystine after reaction with the enzyme. ASA, NADP and NADPH did not prevent cystine inhibition. The [35S]cysteine-labelled enzyme can be visualized after electrophoresis in polyacrylamide gels and further detection by autoradiography. After pepsin treatment of the [35S]cysteine-inactivated enzyme, a main radioactive peptide was isolated by HPLC. The amino acid sequence of this peptide was determined as FVGGN(Cys)(2)TVSL, thus demonstrating that the essential 135Cys is the amino acid residue modified by the treatment with cystine.