Expression and characterization of a novel ß-1,4-endoglucanase from Bacillus subtilis strain isolated from a pulp and paper mill wastewater.

The production of fermentable sugars from lignocellulosic biomass is achieved by the synergistic action of a group of enzymes called cellulases. Cellulose is a long chain of chemically linked glucoses by ß-1,4 bonds. The enzyme ß-1,4-endoglucanase is the first cellulase involved in the degradation, breaking the bond of the amorphous regions. A ß-1,4-endoglucanase enzyme with high activity was obtained from a Bacillus subtilis strain isolated from wastewater of a pulp and paper mill. Sequencing and bioinformatic analysis showed that the gene amplified by PCR consisting of 1407 nucleotides and coding for a ß-1,4-endoglucanase enzyme of approximately 55 kDa. The open reading frame (ORF) encoding the mature endoglucanase (eglS) was successfully inserted in a modified cloning plasmid (pITD03) and into the pYD1 plasmid used for its expression in yeast. Carboxymethylcellulose (CMC) plate assay, SDS-PAGE, and zymogram confirmed the production and secretion by the transformed E. coli BL21-SI strain of a 39 kDa ß-1,4-endoglucanase consistent with the catalytic domain without the cellulose-binding module (CBM). The results showed that the truncated ß-1,4-endoglucanase had higher activity and stability.

Gradually supply of isoamyl alcohol increases the isoamyl acetate production in solid-state fermentation.

Nowadays, isoamyl acetate production is carried out by chemical synthesis with a recent interest in developing biological producing processes, mainly based on microorganisms in submerged fermentation. This work assayed producing isoamyl acetate through solid-state fermentation (SSF), feeding the precursor in the gas phase. Polyurethane foam functioned as the inert support to contain 20 ml of a solution of molasses (10% w/v, pH 5.0). The yeast Pichia fermentans was inoculated at 3 × 107 cells per gram of initial dry weight. The airstream to supply oxygen also served to supply the precursor. Slow supply was obtained using an isoamyl alcohol solution of 5 g l-1 in the bubbling columns and an air stream of 50 ml min-1. For fast supply, fermentations were aerated using 10 g l-1 and 100 ml min-1 for isoamyl alcohol solution and air stream, respectively. It demonstrated the feasibility of isoamyl acetate production in SSF. Moreover, the slow supply of the precursor increased isoamyl acetate production up to 390 mg l-1, which is 12.5 times higher than that obtained without precursor (32 mg l-1). On the other hand, fast supply caused an evident inhibition of the growth and production capacity of the yeast.

Search hours for food attractant by Anastrepha fraterculus and Ceratitis capitata (Diptera: Tephritidae) adults in guava orchards.

Anastrepha fraterculus (Wiedemann, 1830) and Ceratitis capitata (Wiedemann, 1824) are considered the main pests of Brazilian fruit production. Understanding the behavior of species is of great importance for the success of management strategies. This study was to determine the period and search time by attractive food for A. fraterculus and C. capitata adults by using three commercial food attractants: BioAnastrepha™ 5%; Isca Samaritá Tradicional™ 5% and Ceratrap™ 1.5%. The largest catches of A. fraterculus and C. capitata adults in McPhail traps occurred during the day between 6:30 am and 6:30 pm for both species. The BioAnastrepha™ food attractant provided the largest catches compared to Isca Samaritá Tradicional™ and Ceratrap™. In addition, there was a higher prevalence of capturing females than males, for both flies' species. The period of largest search activity for food attractant was observed from 12h:31 pm to 04:30 pm for A. fraterculus and C. capitata, time of greatest temperature on the day. The definition of the period of largest activity of A. fraterculus and C. capitata adults in the field helps in the elaboration of management strategies to be adopted.

Expression of a ß-glucosidase from Trichoderma reesei in Escherichia coli using a synthetic optimized gene and stability improvements by immobilization using magnetite nano-support.

The enzymatic conversion of lignocellulosic biomass to fermentable sugars is determined by the enzymatic activity of cellulases; consequently, improving enzymatic activity has attracted great interest in the scientific community. Cocktails of commercial cellulase often have low ß-glucosidase content, leading to the accumulation of cellobiose. This accumulation inhibits the activity of the cellulolytic complex and can be used to determine the enzymatic efficiency of commercial cellulase cocktails. Here, a novel codon optimized ß-glucosidase gene (B-glusy) from Trichoderma reesei QM6a was cloned and expressed in three strains of Escherichia coli (E. coli). The synthetic sequence containing an open reading frame (ORF) of 1491 bp was used to encode a polypeptide of 497 amino acid residues. The ß-glucosidase recombinant protein that was expressed (57 kDa of molecular weight) was purified by Ni agarose affinity chromatography and visualized by SDS-PAGE. The recombinant protein was better expressed in E. coli BL21 (DE3), and its enzymatic activity was higher at neutral pH and 30 °C (22.4 U/mg). Subsequently, the ß-glucosidase was immobilized using magnetite nano-support, after which it maintained >65% of its enzymatic activity from pH 6 to 10, and was more stable than the free enzyme above 40 °C. The maximum immobilization yield had enzyme activity of 97.2%. In conclusion, ß-glucosidase is efficiently expressed in the microbial strain E. coli BL21 (DE3) grown in a simplified culture medium.

Spinocerebellar ataxia type 3: response to levodopa infusion in two cases.

INTRODUCTION: Spinocerebellar ataxia type 3 (SCA-ATXN3) is a genetic neurodegenerative disease characterized by progressive cerebellar ataxia and other variable findings, including Parkinsonian syndrome. There is no disease-modifying treatment for SCA-ATXN3, so symptom-based management predominates. We aim to illustrate the disease's phenotypic variability and describe the effectiveness of advanced therapies in Parkinsonian symptoms. CASES: We present two patients with a predominant levodopa-responsive Parkinsonian phenotype, combined with cerebellar features. We achieved an optimal control of Parkinsonian symptoms with a carbidopa-levodopa intestinal gel infusion pump. CONCLUSIONS: We should suspect an SCA-ATXN3 etiology in patients with syndromes resembling an early-onset Parkinson disease with an autosomal dominant pattern. These patients could benefit from anti-Parkinsonian treatments, including levodopa intestinal gel infusion pump.

Improvement in the Thermostability of a Recombinant ß-Glucosidase Immobilized in Zeolite under Different Conditions.

ß-Glucosidase is part of the cellulases and is responsible for degrading cellobiose into glucose, a compound that can be used to produce biofuels. However, the use of the free enzyme makes the process more expensive. Enzyme immobilization improves catalytic characteristics and supports, such as zeolites, which have physical-chemical characteristics and ion exchange capacity that have a promising application in the biotechnological industry. This research aimed to immobilize by adsorption a recombinant ß-glucosidase from Trichoderma reesei, obtained in Escherichia coli BL21 (DE3), in a commercial zeolite. A Box Behnken statistical design was applied to find the optimal immobilization parameters, the stability against pH and temperature was determined, and the immobilized enzyme was characterized by SEM. The highest enzymatic activity was determined with 100 mg of zeolite at 35 °C and 175 min. Compared to the free enzyme, the immobilized recombinant ß-glucosidase presented greater activity from pH 2 to 4 and greater thermostability. The kinetic parameters were calculated, and a lower KM value was obtained for the immobilized enzyme compared to the free enzyme. The obtained immobilization parameters by a simple adsorption method and the significant operational stability indicate promising applications in different fields.

Bronchiectasis in severe asthma: a distinct phenotype?

PURPOSE OF REVIEW: The presence of bronchiectasis has been described in about 30% of severe asthma patients. The coexistence of these two respiratory conditions poses new challenges from both clinical and research perspectives. We will review the available literature on this topic to discuss the existance of a specific clinical phenotype of asthma. RECENT FINDINGS: Despite the paucity of literature, the presence of bronchiectasis with severe asthma is associated with older age, chronic bronchial expectoration, rhinosinusitis, more frequent and severe exacerbations, neutrophilic airway inflammation and poor response to usual treatment. Conversely, asthma features are also described in bronchiectasis patients even in the absence of an appropriate diagnosis of asthma. In both cases, there is some evidence supporting the use of bronchodilators, macrolides and respiratory physiotherapy, while the use of inhaled corticosteroids and antibiotics is controversial. SUMMARY: Based on available evidence on the association between (severe) asthma and bronchiectasis, its pathophysiology, certain clinical aspects and prognosis are largely unclear. Although specific management appears to be required in most cases, in our opinion there is still insufficient evidence to consider it a distinct phenotype of severe asthma. Hopefully, future research will shed more light on this topic and define the best therapeutic approach.

Spatial and temporal distribution of metals in PM2.5 during 2013: assessment of wind patterns to the impacts of geogenic and anthropogenic sources.

The Mexico City Metropolitan Area (MCMA) was the object of a chemical elemental characterization (Ti, V, Cr, Mn, Co, Ni, Cu, Mo, Ag, Cd, Sb, Pb, La, Sm, Ce, and Eu) of PM2.5 collected during 2013 and analyzed by inductively coupled plasma mass spectrometry (ICP-MS). Sampling campaigns were carried out at five locations simultaneously-northwest, northeast, center, southwest, and southeast-during dry-warm season (April), rainy season (August), and dry-cold season (November). By means of enrichment factor (EF) and principal component analysis (PCA), it was possible to attribute the analyzed elements to geogenic and anthropogenic sources, as well as to identify a group of elements with mixed provenance sources. The highest concentrations for most metals were found in northwest and northeast, and during dry-warm (DW), confirming the trend observed in PM2.5 samples collected in 2011. Despite similarities between 2011 and 2013, an increase of 17% in PM2.5 mass concentration was observed, mainly attributable to geogenic sources, whereby the importance of wind intensity to the impact of emission sources is highlighted. The effect of wind intensity was revealed, by means of polar plots, as the controlling mechanism for this increase. This allowed us to conclude that high-speed episodes (5 m s-1) were responsible for raising geogenic metal concentrations rather than wind direction.

Urban particulate matter induces the expression of receptors for early and late adhesion molecules on human monocytes.

Exposure to urban particulate matter (PM) is correlated with increases in the emergence of health services due to adverse events and deaths and is mainly related to cardiorespiratory complications. The translocation of particles from the lung into circulation has been proposed as a factor that may trigger systemic effects. Monocytes may be exposed to PM, and if the monocytes are activated, then they are likely to adhere to endothelial cells in a distant organ due to the expression of receptors for adhesion molecules. In the present study, we evaluated the expression of receptors for adhesion molecules (sLex, PSGL-1, LFA-1, VLA-4 and αVß3) in monocytes (U937 cells) exposed for 3 or 18 h to PM10 (0.001, 0.003, 0.010, 0.030, 0.300, 3 or 30 µg/mL). Exposed cells were co-cultured with human endothelial cells that were naive or previously exposed to the same particles. When U937 cells were exposed to PM10, similar levels of expression for early and late receptors for adhesion molecules were observed from 30 ng/mL as those induced by TNF-α. Cells exposed to particles at concentrations above 30 ng/mL were more adhesive to naive or exposed human endothelial cells. Taken together, our results suggest that it is plausible that activated monocytes may play a role in systemic effects induced by PM10 due to the size distribution of the particles and the concentrations required to trigger the expression of receptors for adhesion molecules in monocytes.

Recognition of the importance of geogenic sources in the content of metals in PM2.5 collected in the Mexico City Metropolitan Area.

The study of airborne metals in urban areas is relevant due to their toxic effects on human health and organisms. In this study, we analyzed metals including rare earth elements (REE) in particles smaller than 2.5 µm (PM2.5), collected at five sites around the Mexico City Metropolitan Area (MCMA), during three periods in 2011: April (dry-warm season, DW), August (rainy season, R), and November (dry-cold season, DC). Principal component analysis allowed identifying factors related to geogenic sources and factors related to anthropogenic sources. The recognition of the high impact of geogenic sources in PM2.5 is in agreement with the REE distribution patterns, which show similar behavior as those shown by igneous rocks, confirming the influence of the regional geogenic material. Metals associated to geogenic sources showed higher concentration (p < 0.05) at NE of the MCMA and a significant correlation with prevalent winds. Geogenic metals show similar seasonal distribution, with the highest concentration during DW (p < 0.05), suggesting a possible metal resuspension effect which affects more significantly at lower relative humidity (RH). The metals associated with anthropogenic sources are in agreement with the urban complexity of the area, showing homogenous distribution throughout MCMA (p > 0.05) and no similar seasonal pattern among them. These unexpected results exposed outstanding information regarding the identification of different geogenic sources as the main contributors of metals in the atmospheric environment in the MCMA and highlighted the importance of meteorology in the spatial and seasonal metal patterns.

Atmospheric PM2.5 Mercury in the Metropolitan Area of Mexico City.

In this study, atmospheric mercury concentration in airborne particulate matter with an aerodynamic diameter ≤ 2.5 µm (PM2.5) was analyzed by ICP-MS. Samples were collected in the Mexico City Metropolitan Area (MCMA), during 2013, in five locations, Northwest, Northeast (NE), Central, Southwest and Southeast, along three seasons: dry warm, rainy, and dry cold (DC). It can be observed that NE shows the highest mercury concentration (p < 0.05), where pollution events were identified. The seasonal distribution shows that samples collected during DC present the highest concentration (p < 0.05). These results are in agreement with the distribution of important mercury industrial sources located in the northern urban area as well with the temperature and wind conditions during 2013. The comparison of data obtained in this work with those of similar previous studies clearly indicates a decrease, between 2006 and 2013, of mercury content in PM2.5 collected in MCMA.

Prophylaxis with enoxaparin for prevention of venous thromboembolism after lung transplantation: a retrospective study.

Venous thromboembolism (VTE) is a frequent complication after solid organ transplantation (SOT) and, specifically, after lung transplantation (LT). The objectives of this study were to evaluate prophylaxis with enoxaparin and to describe risk factors for VTE after LT. We retrospectively reviewed the clinical records of 333 patients who underwent LT in our institution between 2009 and 2014. We compared two consecutive cohorts: one that received enoxaparin only during post-transplant hospital admissions and a second cohort that received 90-day extended prophylaxis with enoxaparin. Cumulative incidence function for competing risk analysis was used to determine incidence of VTE during the first year after transplantation. Risk factors were analyzed using a Cox proportional hazards regression model. The cumulative incidence of VTE was 15.3% (95% CI: 11.6-19.4). Median time from transplant to the event was 40 (p25-p75, 14-112) days. Ninety-day extended prophylaxis did not reduce the incidence of VTE. In this study, the risk factors associated with VTE were male gender and interstitial lung disease. VTE is a major complication after LT, and 90-day extended prophylaxis was not able to prevent it. Large, multicenter, randomized clinical trials should be performed to define the best strategy for preventing VTE.

Impacts of commodity prices and governance on the expansion of tropical agricultural frontiers.

Deforestation in the tropics remains a significant global challenge linked to carbon emissions and biodiversity loss. Agriculture, forestry, wildfires, and urbanization have been repeatedly identified as main drivers of tropical deforestation. Understanding the underlying mechanisms behind these direct causes is crucial to navigate the multiple tradeoffs between competing forest uses, such as food and biomass production (SDG 2), climate action (SDG 13), and life on land (SDG 15). This paper develops and implements a global-scale empirical approach to quantify two key factors affecting land use decisions at tropical forest frontiers: agricultural commodity prices and national governance. It relies on data covering the period 2004-2015 from multiple public sources, aggregated to countries and agro-ecological zones. Our analysis confirms the persistent influence of commodity prices on agricultural land expansion, especially in forest-abundant regions. Economic and environmental governance quality co-determines processes of expansion and contraction of agricultural land in the tropics, yet at much smaller magnitudes than other drivers. We derive land supply elasticities for direct use in standard economic impact assessment models and demonstrate that our results make a difference in a Computable General Equilibrium framework.

Sampling, composition, and biological effects of Mexico City airborne particulate matter from multiple periods.

Air pollution is a worldwide environmental problem with an impact on human health. Particulate matter of ten micrometers or less aerodynamic diameter (PM10) as well as its fine fraction (PM2.5) is related to multiple pulmonary diseases. The impact of air pollution in Mexico City, and importantly, particulate matter has been studied and considered as a risk factor for two decades ago. Previous studies have reported the composition of Mexico City particulate matter, as well as the biological effects induced by this material. However, material collected and used in previous studies is a limited resource, and sampling and particle recovery techniques have been improved. In this study, we describe the methods used in our laboratory for Mexico City airborne particulate matter PM10 and PM2.5 sampling, considering the years 2017, 2018 and 2019. We also analyzed the PM10 and PM2.5 samples obtained to determine their composition. Finally, we exposed lung cell line cultures to PM10 and PM2.5 to evaluate the biological effect of the material in terms of cell viability, cell death, inflammatory response, and cytogenetic alterations. Our results showed that PM10 composition includes inorganic, organic and biological compounds, while PM2.5 is a mixture of more enriched organic compounds. PM10 and PM2.5 treatment in lung cells does not significantly impact cell viability/cell death. However, PM10 and PM2.5 increase the secretion levels of IL-6. Moreover, PM10 as well as PM2.5 induce cytogenetic alterations, such as micronuclei, anaphase bridges, trinucleated cells and apoptotic cells in lung cells. Our results update the evidence of the composition and biological effects of Mexico City particulate matter and provide us a reliable basis for future approaches.

Maternal and fetal outcomes in multiparous women with Cystic Fibrosis.

BACKGROUND: Quality of life and survival in Cystic Fibrosis (CF) have improved dramatically, making family planning a feasible option. Maternal and perinatal outcomes in women with CF (wwCF) are similar to those seen in the general population. However, the effect of undergoing multiple pregnancies is unknown. METHODS: A multinational-multicenter retrospective cohort study. Data was obtained from 18 centers worldwide, anonymously, on wwCF 18-45 years old, including disease severity and outcome, as well as obstetric and newborn complications. Data were analyzed, within each individual patient to compare the outcomes of an initial pregnancy (1st or 2nd) with a multigravid pregnancy (≥3) as well as secondary analysis of grouped data to identify risk factors for disease progression or adverse neonatal outcomes. Three time periods were assessed - before, during, and after pregnancy. RESULTS: The study population included 141 wwCF of whom 41 (29%) had ≥3 pregnancies, "multiparous". Data were collected on 246 pregnancies, between 1973 and 2020, 69 (28%) were multiparous. A greater decline in ppFEV1 was seen in multiparous women, primarily in pancreatic insufficient (PI) wwCF and those with two severe (class I-III) mutations. Multigravid pregnancies were shorter, especially in wwCF over 30 years old, who had high rates of prematurity and newborn complications. There was no effect on pulmonary exacerbations or disease-related complications. CONCLUSIONS: Multiple pregnancies in wwCF are associated with accelerated respiratory deterioration and higher rates of preterm births. Therefore, strict follow-up by a multidisciplinary CF and obstetric team is needed in women who desire to carry multiple pregnancies.

Human health risk assessment of PM10-bound heavy metals and PAHs around the Latin America's Largest opencast coal mine.

Air pollution in opencast coal mine areas is a critical issue, resulting in harmful severe effects on human health. Therefore, it is essential to understand the air pollution factors and to assess the risks to humans. This study evaluated the potential risks (carcinogen and non-carcinogen) of inhalation exposure to PM10-bound heavy metals and Polycyclic Aromatic Hydrocarbons (PAHs) in an open pit mine in northern Colombia. During February-May 2022, PM10 samples were collected at eight sites. Heavy metals (i.e., Al, Cr(VI), Mn, Cu, Zn, As, Pb) and PAHs (thirteen priority PAHs, and one non-priority PAH) levels linked to PM10 were analyzed by X-ray fluorescence and gas chromatography-mass spectrometry, respectively. PM10 concentrations were found to range between 4.70 and 59.90 µg m-3. Out of the three different zones of the study area (i.e., North Zone, South Zone, and Populated Zone), the North Zone recorded the highest daily average concentrations of Cr(VI) (104.16 ng m-3), Mn (28.39 ng m-3), Cu (33.75 ng m-3), Zn (57.99 ng m-3), As (44.92 ng m-3), and Pb (27.13 ng m-3). The fraction of the analyzed heavy metals at all monitoring sites was 82%-89% for Al, followed by Cr(VI) with 3%-6%. Cr(VI) was the major contributor to the carcinogenic risk values, while Cu, Cr(VI), and As were the main drivers for the non-carcinogenic risk. The average cancer risk range for heavy metals was 3.30 × 10-04 -5.47 × 10-04. On the other hand, the cancer risk for PAHs exposure was acceptable. The average incremental lifetime cancer risk (ILCR) values varied between 2.87 × 10-07 and 4.21 × 10-07. Benzo[a]pyrene contributed to 54%-56% of the total risk from inhalation of PM10-bound PAHs, while Indeno[1,2,3-cd]pyrene contributed to 16%-19%. Based on the Monte Carlo sensitivity analysis, exposure to Cr(VI) was the main factor affecting cancer risk in the North, South, and Populated Zones. A suitable risk assessment and management plan requires understanding PM10-bound heavy metals and PAHs concentration levels as well as their potential health risks, mainly in open-cast coal mine zones. Our study found that people living near open-pit mines face potential health risks, so it is crucial to establish policies and regulations to control emission sources.

Lethal and sublethal effects of toxic bait formulations on Doryctobracon areolatus (Hymenoptera: Braconidae) and implications for integrated fruit fly management.

The use of toxic baits has become one of the main methods of management of fruit flies in Brazil. The application of toxic baits may cause side effects on the native parasitoid Doryctobracon areolatus (Hymenoptera: Braconidae). Based on the results, formulations made from the food attractants 3% Biofruit, 1.5% Ceratrap, 1.25% Flyral, 3% Isca Samaritá, 3% Isca Samaritá Tradicional, and 7% sugarcane molasses associated with the Malathion 1000 EC and the ready-to-use toxic bait Gelsura (containing the active ingredient alpha-cypermethrin) were classified as harmful (class 4) to D. areolatus (mortality > 85% at 96 HAE). In contrast, for toxic baits formulated with insecticide phosmet, the mortality ranged from 38% to 72%, classified as slightly harmful or moderately harmful. However, when phosmet was added to the 3% Samaritá Tradicional bait, the mortality was only 3.9% (class 1-harmless), similar to the toxicity observed for the Success 0.02 CB ready-to-use bait (0.24 g a.i. spinosad/l) (<5% mortality). Although toxic baits were formulated with spinosyn-based insecticides, all toxic bait formulations were classified as harmless or slightly harmful (<50% mortality) to D. areolatus, with the exception of 1.5% Ceratrap + spinetoram and 7% Sugarcane molasses + spinosad (≈ 60% mortality-moderately harmful). In addition, these formulations did not show sublethal effects in reducing the parasitism and emergence rate of the F1 generation of D. areolatus in A. fraterculus larvae. The results serve as a basis for the correct use of toxic food baits without affecting the biological control.

How can I assess my patients with Parkinson's disease during a busy clinic day?

BACKGROUND: The evaluation of motor impairment in Parkinson's disease (PD) is mainly assessed with the motor subdomain of the Unified Parkinson's Disease Rating scale (UPDRS part III) and, lately, with the MDS-UPDRS part III. To optimize efforts and special needs during specific circumstances in clinical practice, we sought to identify the most sensitive items to assess motor impairment in PD. METHODS: We included the COPPADIS-PD cohort and collected the UPDRS part III at baseline (V0), 12 months (V1), and 24 months (V2). Factor analysis and effect size using Cohen's d formula were performed in the Off and On states at V0, V1, and V2. RESULTS: We included 667 patients with PD, mean age of 62.59 ± 8.91 years, 410 (60.2%) males, with a median HY stage of 2.00 (1.00; 4.00) at baseline. Over time, the most discriminating items were postural stability and body bradykinesia ("arise from chair" and "gait") in the Off state, right and left upper extremity bradykinesia ("finger tap", "hand movements" and "prono/supination") in the On state. Body bradykinesia and right-left finger tapping were the items with the largest effect size (0.93, 0.84, 0.83, respectively) to assess motor improvement after receiving antiparkinsonian medications over time. CONCLUSION: Under specific circumstances, selecting a few items of the UPDRS part III, including postural stability, body bradykinesia, and upper extremity bradykinesia, could be used to create a quick clinical judgment of motor status and improvement in PD.