Purification and characterization of a thrombolytic enzyme produced by a new strain of Bacillus subtilis

Fibrinolytic enzymes with a direct mechanism of action and safer properties are currently requested for thrombolytic therapy. This paper reports on a new enzyme capable of degrading blood clots directly without impairing blood coagulation. This enzyme is also non-cytotoxic and constitutes an alternative to other thrombolytic enzymes known to cause undesired side effects. Twenty-four Bacillus isolates were screened for production of fibrinolytic enzymes using a fibrin agar plate. Based on produced activity, isolate S127e was selected and identified as B. subtilis using the 16S rDNA gene sequence. This strain is of biotechnological interest for producing high fibrinolytic yield and consequently has potential in the industrial field. The purified fibrinolytic enzyme has a molecular mass of 27.3 kDa, a predicted pI of 6.6, and a maximal affinity for Ala-Ala-Pro-Phe. This enzyme was almost completely inhibited by chymostatin with optimal activity at 48°C and pH 7. Specific subtilisin features were found in the gene sequence, indicating that this enzyme belongs to the BPN group of the S8 subtilisin family and was assigned as AprE127. This subtilisin increased thromboplastin time by 3.7% (37.6 to 39 s) and prothrombin time by 3.2% (12.6 to 13 s), both within normal ranges. In a whole blood euglobulin assay, this enzyme did not impair coagulation but reduced lysis time significantly. Moreover, in an in vitro assay, AprE127 completely dissolved a thrombus of about 1 cc within 50 min and, in vivo, reduced a thrombus prompted in a rat tail by 11.4% in 24 h compared to non-treated animals.This work was supported by Direcao Regional da Ciencia e Tecnologia - DRCT-Acores (Medida 2.2.2/I/025/2008) . Duarte Toubarro received a post-doctoral grant (SFRH/BPD/77483/2011; M3.1.a/F/050/2016) from FundacAo para a Ciencia e Tecnologia, Portugal; Jorge Frias received a doctoral grant (SFRH/BD/131698/2017) . The authors are grateful to Dr. Teresa Sampaio and Nelia Arruda, from Dr. Forjaz Sampaio Medical Center, for helping with the plasma anticoagulation assays. Duarte Toubarro and Nelson Simoes designed the study and helped with the paper's redaction. Jorge Frias performed the experiments, analyzed the data and wrote the paper. Alexandra Fraga and Jorge Pedrosa performed thrombolytic assays in a rat model. Claudia Botelho and Jose Teixeira participated in the discussion and redaction of the paper.info:eu-repo/semantics/publishedVersio

Gross motor coordination and weight status of Portuguese children aged 6-14 years

Objectives: To construct age- and gender-specific percentiles for gross motor coordination (MC) tests and to explore differences in gross MC in normal-weight, overweight and obese children. Methods: Data are from the "Healthy Growth of Madeira Study", a cross-sectional study carried out in children, aged 6–14 years. All 1,276 participants, 619 boys and 657 girls, were assessed for gross MC (Korperkoordinations Test fur Kinder, KTK), anthropometry (height and body mass), physical activity (Baecke questionnaire) and socioeconomic status (SES). Centile curves for gross MC were obtained for boys and girls separately using generalized additive models for location, scale and shape. Results: A significant main effect for age was found in walking backwards and moving sideways. Boys performed significantly better than girls on moving sideways. At the upper limit of the distributions, interindividual variability was higher in hopping on one leg (girls) and jumping and moving sideways (boys and girls). One-way ANCOVA, controlling for age, physical activity and SES, indicated that normal-weight children scored significantly better than their obese peers in all gross MC tests. Overweight boys and girls also scored significantly better than their obese colleagues in some MC tests. Conclusions: These centile curves can be used as reference data in Portuguese children and youth, aged 6–14 years. Being overweight or obese was a major limitation in MC tests and, therefore, of the children’s health- and performance related physical fitness

Predator Avoidance in the European Seabass After Recovery From Short-Term Hypoxia and Different CO2 Conditions

Short-term hypoxia that lasts just a few days or even hours is a major threat for the marine ecosystems. The single effect of the human-induced levels of hypoxia and other anthropogenic impacts such as elevated pCO2 can reduce the ability of preys to detect their predators across taxa. Moreover, both processes, hypoxia and elevated pCO2, are expected to co-occur in certain habitats, but the synergic consequences of both processes and the ability of fish to recover remain unknown. To provide empirical evidence to this synergy, we experimentally evaluated the risk-taking behavior in juveniles of the European seabass (Dicentrachus labrax), an important commercial fisheries species after recovering from short-term hypoxia and different pH scenarios. The behavior of seabass juveniles was monitored in an experimental arena before and after the exposure to a simulated predator and contrasted to control fish (BACI design) (current levels of hypoxia and elevated pCO2) using a mechanistic function-valued modeling trait approach. Results revealed that fish recovering from elevated pCO2, alone or combined with hypoxia, presented less avoidance behavior in failing to seek refuge when a simulated predator was present in the arena compared to those exposed to control pCO2 levels. Our results show that recovery from short-term exposure to acidification and hypoxia was not synergistic and suggest that recovery from acidification takes longer than from short-term hypoxia treatment through a potential effect on the sensorial and hence behavioral capacities of fish

Gap-analysis and annotated reference library for supporting macroinvertebrate metabarcoding in Atlantic Iberia

DNA metabarcoding provides a rapid and effective identification tool of macroinvertebrate species. The accuracy of species-level assignment, and consequent taxonomic coverage, relies on comprehensive DNA barcode reference libraries, which, due to incompleteness, are currently a recognized limitation for metabarcoding applications. In this study, we assembled a comprehensive reference library of DNA barcodes for Atlantic Iberia marine macroinvertebrate species, assessed gaps in species coverage and examined data ambiguities. Initially, an Iberian species checklist for the three dominant groups of marine macroinvertebrates was compiled, comprising 2827 species (926 Annelida, 638 Crustacea and 1263 Mollusca). A total of 18162 DNA sequences of the cytochrome c oxidase subunit I barcode region (COI-5P) matching the species checklist were compiled in a BOLD dataset, where taxonomic discordances were evaluated and cases of deep intraspecific divergence flagged. Gap-analysis showed that 63% of the Iberian macroinvertebrate species still lack a DNA barcode. Coverage gaps varied considerably across taxonomic groups with Mollusca displaying the highest sequence representation in the dataset (427 species, 49% of the total number of sequences), and Crustacea the highest species coverage with 338 species barcoded (53% of the checklist). In contrast, Polychaeta displayed the lower levels of completion (288 species, 16% of the total number of sequences). In total, 1545 Barcode Index Numbers (BINs) were assigned to 1053 barcoded species, of which 66% were taxonomically concordant, 26% displayed multiple BINs and 8% were discordant. Overall, results show that there is still a large portion of marine invertebrate taxa in this region of Europe pending barcode coverage, even considering only the dominant groups. However, the most notable finding was the relevant proportion of species flagged for significant intraspecific divergence and possible hidden diversity. The annotated reference library and gap-analysis here provided can thereThis study was supported by the project The NextSea [NORTE-01-0145-FEDER-000032], under the PORTUGAL 2020 Partnership Agreement, through the European Regional Development Fund (ERDF). This work was supported by the "Contrato-Programa'' UIDB/04050/2020 funded by national funds through the FCT I.P. SD and PEV work was supported by national funds through the Portuguese Foundation for Science and Technology (FCT, I.P.) in the scope of the project NIS-DNA [PTDC/BIA-BMA/29754/2017]. BRL benefitted from an FCT fellowship PD/BD/127994/2016. MALT benefitted from an FCT fellowship co-financed by European Social Fund (ESF) SFRH/BD/131527/2017

Revision and annotation of DNA barcode records for marine invertebrates: Report of the 8th iBOL conference hackathon

The accuracy of specimen identification through DNA barcoding and metabarcoding relies on reference libraries containing records with reliable taxonomy and sequence quality. The considerable growth in barcode data requires stringent data curation, especially in taxonomically difficult groups such as marine invertebrates. A major effort in curating marine barcode data in the Barcode of Life Data Systems (BOLD) was undertaken during the 8th International Barcode of Life Conference (Trondheim, Norway, 2019). Major taxonomic groups (crustaceans, echinoderms, molluscs, and polychaetes) were reviewed to identify those which had disagreement between Linnaean names and Barcode Index Numbers (BINs). The records with disagreement were annotated with four tags: A) MIS-ID (misidentified, mislabeled, or contaminated records), b) AMBIG (ambiguous records unresolved with the existing data), c) COMPLEX (species names occurring in multiple BINs), and d) SHARE (barcodes shared between species). A total of 83,712 specimen records corresponding to 7,576 species were reviewed and 39% of the species were tagged (7% MIS-ID, 17% AMBIG, 14% COMPLEX, and 1% SHARE). High percentages (>50%) of AMBIG tags were recorded in gastropods, whereas COMPLEX tags dominated in crustaceans and polychaetes. The high proportion of tagged species reflects either flaws in the barcoding workflow (e.g., misidentification, cross-contamination) or taxonomic difficulties (e.g., synonyms, undescribed species). Although data curation is essential for barcode applications, such manual attempts to examine large datasets are unsustainable and automated solutions are extremely desirable.The hackathon was organized with financial support from the European Union COST Action DNAqua-Net (CA 15219 https://dnaqua.net/) in the scope of the 8th International Barcode of Life Conference in Trondheim, Norway on 16 June 2019. DNAqua-Net is acknowledged for the funding provided and the local conference organizers for all the logistical support that ensured a successful event. Tyler Elliot and the rest of the BOLD team are acknowledged for their help with data queries and analytics. The authors also thank the hackathon participants for vibrant discussions during and after the event: Berry van der Hoorn, Katrine Konsghavn, Guy Paz, Mouna Rifi, Malin Strand, Anne Helene Tandberg, Adam Wall, and Endre Willassen. Marcos A. L. Teixeira was supported by a PhD grant from the Portuguese Foundation for Science and Technology (FCT I.P.) co-financed by ESF (SFRH/BD/131527/2017). Financial support granted by FCT to Sofia Duarte (CEECIND/00667/2017) and to Pedro E. Vieira (project NIS-DNA, PTDC/BIA-BMA/29754/2017) is also acknowledged. Sanna Majaneva was financially supported by the Norwegian Taxonomy Initiative (project no. 70184235). The authors thank the five reviewers who provided valuable input into the earlier version of the manuscript

Tracking prostate carcinogenesis over time through urine proteome profiling in an animal model: an exploratory approach

Prostate cancer (PCa) is one of the most lethal diseases in men, which justifies the search for new diagnostic tools. The aim of the present study was to gain new insights into the progression of prostate carcinogenesis by analyzing the urine proteome. To this end, urine from healthy animals and animals with prostate adenocarcinoma was analyzed at two time points: 27 and 54 weeks. After 54 weeks, the incidence of pre-neoplastic and neoplastic lesions in the PCa animals was 100%. GeLC-MS/MS and subsequent bioinformatics analyses revealed several proteins involved in prostate carcinogenesis. Increased levels of retinol-binding protein 4 and decreased levels of cadherin-2 appear to be characteristic of early stages of the disease, whereas increased levels of enolase-1 and T-kininogen 2 and decreased levels of isocitrate dehydrogenase 2 describe more advanced stages. With increasing age, urinary levels of clusterin and corticosteroid-binding globulin increased and neprilysin levels decreased, all of which appear to play a role in prostate hyperplasia or carcinogenesis. The present exploratory analysis can be considered as a starting point for studies targeting specific human urine proteins for early detection of age-related maladaptive changes in the prostate that may lead to cancer.info:eu-repo/semantics/publishedVersio

Insect chemical ecology: chemically mediated interactions and novel applications in agriculture

Forum PaperInsect chemical ecology (ICE) evolved as a discipline concerned with plant–insect interactions, and also with a strong focus on intraspecific pheromone-mediated communication. Progress in this field has rendered a more complete picture of how insects exploit chemical information in their surroundings in order to survive and navigate their world successfully. Simultaneously, this progress has prompted new research questions about the evolution of insect chemosensation and related ecological adaptations, molecular mechanisms that mediate commonly observed behaviors, and the consequences of chemically mediated interactions in different ecosystems. Themed meetings, workshops, and summer schools are ideal platforms for discussing scientific advancements as well as identifying gaps and challenges within the discipline. From the 11th to the 22nd of June 2018, the 11th annual PhD course in ICE was held at the Swedish University of Agricultural Sciences (SLU) Alnarp, Sweden. The course was made up of 35 student participants from 22 nationalities (Fig. 1a) as well as 32 lecturers. Lectures and laboratory demonstrations were supported by literature seminars, and four broad research areas were covered: (1) multitrophic interactions and plant defenses, (2) chemical communication focusing on odor sensing, processing, and behavior, (3) disease vectors, and (4) applied aspects of basic ICE research in agriculture. This particular article contains a summary and brief synthesis of these main emergent themes and discussions from the ICE 2018 course. In addition, we also provide suggestions on teaching the next generation of ICE scientists, especially during unprecedented global situationsinfo:eu-repo/semantics/publishedVersio

Um guia digital para a Natureza : promover a floresta dos Açores com aprendizagem baseada no lugar

A secção UAciência é coordenada pelo Professor Universitário Armindo Rodrigues.O projeto Guia de Campo (GC) no âmbito da transformação digital operada nos modos de aprendizagem característicos das sociedades contemporâneas. Hoje, as ferramentas móveis interativas e a aprendizagem baseada no lugar são recursos pedagógicos cada vez mais presentes na educação ambiental. O projeto GC pretende investigar o papel das tecnologias digitais interativas no desenvolvimento da literacia científica, ambiental e da conservação, entre jovens residentes nos Açores, ajudando a tornar mais significativas e apelativas as experiências educativas desenvolvidas em contextos naturais. A instrumentalidade da videofilia na estimulação da biofilia e testada neste projeto, com vista à promoção de conhecimentos, atitudes e práticas conservacionistas, uma vez que vários estudos evidenciam que os jovens só tendem a proteger o que conhecem e entendem, e que a promoção de experiências ativas na natureza (ou sobre esta) estimula a biofilia e a curiosidade das crianças. […].info:eu-repo/semantics/publishedVersio

Fractional order dynamical systems and its applications

This article illustrates several applications of fractional calculus (FC) in science and engineering. It has been recognized the advantageous use of this mathematical tool in the modeling and control of many dynamical systems. In this perspective, this paper investigates the use of FC in the following fields: Controller tuning; Electrical systems; Traffic systems; Digital circuit synthesis; Evolutionary computing; Redundant robots; Legged robots; Robotic manipulators; Nonlinear friction; Financial modeling.N/