Insights into Campylobacter jejuni Desulforubrerythrin catalytic mechanism

Dissertation presented to obtain the Master Degree in Molecular Genetics and BiomedicineThe following work aims to contribute to a better understanding of systems involved in resistance to oxidative stress species, namely hydrogen peroxide. The work is focused in one protein from the pathogen Campylobacter jejuni: desulforubrerythrin. Desulforubrerythrin is a non-heme iron protein in which the catalytic centre harbours a diiron cluster. Besides, the protein has a desulforedoxin domain at the N-terminal and a rubredoxin domain at the C-terminal. With the objective of understanding the protein catalytic mechanism three site-directed mutants, as well the wild type protein, were over expressed in Escherichia coli, purified and studied through biochemical and spectroscopic techniques. The amino acid residues selected for mutations are two tyrosines near the catalytic centre (residues 59 and 127). These residues are strictly conserved in rubrerythrins; moreover in diiron centres containing proteins tyrosines play a role in dissipating oxidizing species of iron (IV) by forming a tyrosil radical. The selected residues were replaced by a phenalanine residue which gave rise to three mutants: Y59F, Y127F and Y59F Y127F. These were characterized having as reference the wild type protein. All proteins have a molecular mass of 24 kDa and are tetramers in solution. The EPR and UV-visible techniques confirmed the presence of the three metallic domains in the wild type and Y59F mutant. The Y127F mutant was successfully used to test a protocol for diiron centre reconstitution in desulforubrerythrin. Finally, crystals of the wild type and, for the first time, of the Y59F and double mutants were obtained. The X-ray data for the mutants were collected with a resolution of 1.9 Å and its structure will be determined

Co-immobilization of Liposomal amphotericin B and antimicrobial peptides to prevent multi-kingdom infections

The drawbacks arising from microbial colonisation of indwelling devices have been well established, as evidenced by a massive body research on antimicrobial coating strategies. Most of these strategies, however, are designed to target bacterial biofilms, being fungal biofilms or mixed biofilms much less taken into account. In real-life settings, fungi will be inevitably found in consortium with bacteria, so the development of antifungal coating strategies to be combined with antibacterial approaches will be pivotal for the fight of biomaterial-associated infections (BAI). This study aimed to engineer an effective strategy for the immobilization of liposomal amphotericin B (LAmB) on polydimethylsiloxane (PDMS) surfaces to be afterwards co-immobilized with antimicrobial peptides (AMP). Immobilization was performed using a two-step mussel-inspired coating strategy, in which PDMS were first immersed in dopamine solution and its self-polymerization leads to the deposition of a thin adherent film, called polydopamine (pDA), which allowed further incorporation of LAmB and/or AMP. Surface characterization confirmed the polymerization of dopamine and further functionalization with only LAmB yielded surfaces with less roughness and more hydrophilic features. It also rendered the surfaces of PDMS with the ability to prevent the attachment of Candida albicans and kill the adherent cells, without toxicity towards mammalian cells. The decreased number of adhered cells observed on these surfaces together with the no cytotoxicity found suggested the release of AmB but not in its free formulation. It was hypothesized that liposomes were released from the surfaces to target fungi cells, being able to cross the cell wall and reach the cell membrane where a higher affinity to ergosterol than the cholesterol of liposomes will allow AmB to be free and promote fungal death. AMP immobilization alone provided PDMS surfaces with contact-killing activity towards Proteus mirabilis in the first 4 h of colonization and no leaching was observed. AMP permanent immobilization comprised, however, a disadvantage for longer periods of exposure which was attributed to the fact that dead adhered cells can mask the antimicrobial features of contactkilling surfaces, providing a surface for new colonization. Co-immobilization of both LAmB and AMP showed that LAmB retained its antifungal activity towards C. albicans, even when challenged by a multi-kingdom consortium. Although AMP immobilization requires further optimization to promote its controlled release from the surfaces, overall results highlight the remarkable antifungal and biocompatible properties obtained with LAmB immobilization. This coating strategy holds, therefore, a great potential to be combined with antibacterial agents in the development of approaches to fight BAI.info:eu-repo/semantics/publishedVersio

Boas práticas ao Serviço do Utente - Centro Hospitalar do Tâmega e Sousa, EPE

O CHTS pretende e ambiciona na literacia em saúde, na vertente do cidadão, que haja mais igualdades em saúde e que, este cidadão perante a necessidade de tomar decisões de forma autónoma (muitas vezes de elevada complexidade), sobre a promoção de saúde, prevenção das doenças ou seu tratamento, esteja informado e com conhecimentos para o fazer. Pretende que o cidadão seja capaz de obter melhor acesso aos cuidados de saúde, usar e usufruir da forma mais adequada e, de forma intencional e consciente, possa obter os maiores benefíciospara a manutenção do seu estado de saúde. A OMS, define Literacia em Saúde como “o grau em que os indivíduos têm a capacidade de obter, processar e entender as informações básicas de saúde para utilizarem os serviços e tomarem decisões adequadas de saúde”, ou seja, a literacia em saúde contempla um conjunto de conhecimentos, atitudes, habilidades e até competências que capacitam a pessoa no acesso, compreensão das informações para que possa avaliar de forma critica a sua relevância no uso responsável desse conhecimento. Foi, neste contexto, que surgiu no CHTS uma nova ótica de leitura e de intervenção das suas equipas multidisciplinares, na consecução de projetos e ações que visam reforçar os níveis de literacia, de forma multidimensional e colaborativa, aproximando-se cada vez mais da centralidade no cidadão, bem como de uma maior eficiência e eficácia dos serviços, qualidade assistencial e satisfação do cidadão e profissional. Deve-se muito à capacidade dos profissionais de saúde, mesmo com diferenças de uns para outros, em identificar as necessidades das pessoas, em estarem disponíveis para promover mudança, a avaliar diariamente o nível de compreensão, capacidades para realizar tarefas prescritas, motivação e nível de mudança comportamental do cidadão, tendo em conta a sua idade e o seu estado de saúde. José Ribeiro Nunes, Enf. Diretor, Prefácioinfo:eu-repo/semantics/publishedVersio

Characterisation of microbial attack on archaeological bone

As part of an EU funded project to investigate the factors influencing bone preservation in the archaeological record, more than 250 bones from 41 archaeological sites in five countries spanning four climatic regions were studied for diagenetic alteration. Sites were selected to cover a range of environmental conditions and archaeological contexts. Microscopic and physical (mercury intrusion porosimetry) analyses of these bones revealed that the majority (68%) had suffered microbial attack. Furthermore, significant differences were found between animal and human bone in both the state of preservation and the type of microbial attack present. These differences in preservation might result from differences in early taphonomy of the bones. © 2003 Elsevier Science Ltd. All rights reserved

Global variation in postoperative mortality and complications after cancer surgery: a multicentre, prospective cohort study in 82 countries

© 2021 The Author(s). Published by Elsevier Ltd. This is an Open Access article under the CC BY-NC-ND 4.0 licenseBackground: 80% of individuals with cancer will require a surgical procedure, yet little comparative data exist on early outcomes in low-income and middle-income countries (LMICs). We compared postoperative outcomes in breast, colorectal, and gastric cancer surgery in hospitals worldwide, focusing on the effect of disease stage and complications on postoperative mortality. Methods: This was a multicentre, international prospective cohort study of consecutive adult patients undergoing surgery for primary breast, colorectal, or gastric cancer requiring a skin incision done under general or neuraxial anaesthesia. The primary outcome was death or major complication within 30 days of surgery. Multilevel logistic regression determined relationships within three-level nested models of patients within hospitals and countries. Hospital-level infrastructure effects were explored with three-way mediation analyses. This study was registered with ClinicalTrials.gov, NCT03471494. Findings: Between April 1, 2018, and Jan 31, 2019, we enrolled 15 958 patients from 428 hospitals in 82 countries (high income 9106 patients, 31 countries; upper-middle income 2721 patients, 23 countries; or lower-middle income 4131 patients, 28 countries). Patients in LMICs presented with more advanced disease compared with patients in high-income countries. 30-day mortality was higher for gastric cancer in low-income or lower-middle-income countries (adjusted odds ratio 3·72, 95% CI 1·70–8·16) and for colorectal cancer in low-income or lower-middle-income countries (4·59, 2·39–8·80) and upper-middle-income countries (2·06, 1·11–3·83). No difference in 30-day mortality was seen in breast cancer. The proportion of patients who died after a major complication was greatest in low-income or lower-middle-income countries (6·15, 3·26–11·59) and upper-middle-income countries (3·89, 2·08–7·29). Postoperative death after complications was partly explained by patient factors (60%) and partly by hospital or country (40%). The absence of consistently available postoperative care facilities was associated with seven to 10 more deaths per 100 major complications in LMICs. Cancer stage alone explained little of the early variation in mortality or postoperative complications. Interpretation: Higher levels of mortality after cancer surgery in LMICs was not fully explained by later presentation of disease. The capacity to rescue patients from surgical complications is a tangible opportunity for meaningful intervention. Early death after cancer surgery might be reduced by policies focusing on strengthening perioperative care systems to detect and intervene in common complications. Funding: National Institute for Health Research Global Health Research Unit

Effects of hospital facilities on patient outcomes after cancer surgery: an international, prospective, observational study

© 2022 The Author(s). Published by Elsevier Ltd. This is an Open Access article under the CC BY 4.0 licenseBackground: Early death after cancer surgery is higher in low-income and middle-income countries (LMICs) compared with in high-income countries, yet the impact of facility characteristics on early postoperative outcomes is unknown. The aim of this study was to examine the association between hospital infrastructure, resource availability, and processes on early outcomes after cancer surgery worldwide. Methods: A multimethods analysis was performed as part of the GlobalSurg 3 study—a multicentre, international, prospective cohort study of patients who had surgery for breast, colorectal, or gastric cancer. The primary outcomes were 30-day mortality and 30-day major complication rates. Potentially beneficial hospital facilities were identified by variable selection to select those associated with 30-day mortality. Adjusted outcomes were determined using generalised estimating equations to account for patient characteristics and country-income group, with population stratification by hospital. Findings: Between April 1, 2018, and April 23, 2019, facility-level data were collected for 9685 patients across 238 hospitals in 66 countries (91 hospitals in 20 high-income countries; 57 hospitals in 19 upper-middle-income countries; and 90 hospitals in 27 low-income to lower-middle-income countries). The availability of five hospital facilities was inversely associated with mortality: ultrasound, CT scanner, critical care unit, opioid analgesia, and oncologist. After adjustment for case-mix and country income group, hospitals with three or fewer of these facilities (62 hospitals, 1294 patients) had higher mortality compared with those with four or five (adjusted odds ratio [OR] 3·85 [95% CI 2·58–5·75]; p<0·0001), with excess mortality predominantly explained by a limited capacity to rescue following the development of major complications (63·0% vs 82·7%; OR 0·35 [0·23–0·53]; p<0·0001). Across LMICs, improvements in hospital facilities would prevent one to three deaths for every 100 patients undergoing surgery for cancer. Interpretation: Hospitals with higher levels of infrastructure and resources have better outcomes after cancer surgery, independent of country income. Without urgent strengthening of hospital infrastructure and resources, the reductions in cancer-associated mortality associated with improved access will not be realised. Funding: National Institute for Health and Care Research