PLEIOTROPIC PHENOTYPE OF PSEUDOMONAS AERUGINOSA MUTANTS DEFECTIVE IN GLUCOSE UPTAKE

Pseudomonas aeruginosa \ue8 un patogeno opportunista che provoca una vasta gamma di infezioni nell\u2019uomo. \uc8 noto che l\u2019incidenza delle infezioni da P. aeruginosa \ue8 maggiore in persone che presentano elevati livelli di glucosio nel sangue, a causa della capacit\ue0 del batterio di utilizzare il glucosio in eccesso come nutriente per la crescita. Pertanto, bloccare l\u2019attivit\ue0 di proteine coinvolte nel trasporto o nell\u2019utilizzazione del glucosio da parte del batterio potrebbe essere una buona strategia per lo sviluppo di farmaci anti-pseudomonas. Per valutare questa ipotesi, abbiamo costruito una serie di mutanti singoli e multipli difettivi nel trasporto del glucosio nel ceppo PAO1 di P. aeruginosa e li abbiamo analizzati per fenotipi correlati alla virulenza in saggi in vitro e in vivo, in due differenti modelli di infezione. In particolare, sono stati deleti i geni codificanti per i trasportatori della membrana interna Glt, GntP e Kgut, che mediano l\u2019ingresso del glucosio e dei suoi derivati ossidati gluconato e 2-ketogluconato. Un triplo mutante privo dei suddetti trasportatori si \ue8 rivelato incapace di crescere su glucosio come unica fonte di carbonio (mutante GUN, Glucose Uptake Null). Pi\uf9 di 500 geni, che controllano sia funzioni metaboliche che la virulenza, sono espressi in modo differenziale nel mutante GUN rispetto al ceppo parentale. Coerentemente con i dati dell\u2019analisi trascrittomica, i saggi di virulenza in vitro hanno mostrato che il mutante GUN si comporta diversamente dal ceppo parentale, avendo una ridotta capacit\ue0 di formare biofilm e una maggiore secrezione di proteasi, piocianina e pioverdina. Inoltre, questo mutante ha una produzione alterata delle molecole segnale che attivano il sistema del quorum sensing. \uc8 interessante notare che il profilo trascrizionale e la maggior parte dei tratti fenotipici analizzati differiscono tra il mutante GUN e il ceppo parentale indipendentemente dalla presenza del glucosio, suggerendo che uno (o pi\uf9) dei trasportatori deleti abbia una funzione addizionale non correlata al trasporto del glucosio. Infine, alcuni mutanti hanno mostrato un diverso grado di virulenza in saggi di infezione negli ospiti modello Caenorhabditis elegans e Galleria mellonella. In particolare, mentre in C. elegans la virulenza dei mutanti \ue8 risultata simile a quella del ceppo parentale PAO1, i mutanti deleti del gene kguT, che codifica per il trasportatore del 2-ketogluconato, sono meno virulenti di PAO1 in G. mellonella. L\u2019attenuazione \ue8 particolarmente significativa per il doppio mutante \u394gntP \u394kguT e per il mutante GUN. Questo risultato suggerisce che l\u2019efficacia antibatterica di composti che bloccano l\u2019utilizzo del glucosio da parte di P. aeruginosa potrebbe presumibilmente variare a seconda della capacit\ue0 del patogeno di adattarsi allo specifico contesto nutrizionale dell\u2019ospite infettato. Una maggiore conoscenza delle interazioni metaboliche che avvengono tra patogeno ed ospite nel sito d\u2019infezione diventa quindi sempre pi\uf9 necessaria per poter sviluppare terapie antibatteriche efficaci.Pseudomonas aeruginosa is an opportunistic pathogen causing a wide range of infections in humans. Pathologies leading to hyperglycaemia have been associated with augmented risk of developing serious P. aeruginosa infections due to the ability of the bacterium to utilize glucose as carbon source for the growth. Therefore, preventing the import of glucose might be a good strategy to develop anti-pseudomonas drugs. To address this hypothesis, a collection of single and multiple glucose uptake defective mutants was generated in P. aeruginosa PAO1 and tested for virulence-related phenotypes in in vitro assays and in vivo, in two different infection models. In particular, we engineered mutants in genes encoding inner membrane (IM) proteins involved in the internalization of glucose and its oxidized derivatives gluconate and 2-ketogluconate, i.e. the Glt, GntP and KguT transporters. A triple mutant lacking these transporters was demonstrated to be completely unable to grow on glucose as sole carbon source (Glucose Uptake Null mutant, GUN). The transcriptomic analysis revealed a strong divergence in the GUN transcriptional profile relative to the parental strain, with more than 500 differentially expressed genes, controlling both metabolic functions and virulence traits. Consistent with the transcriptomic data, the GUN mutant showed a pleiotropic phenotype in the in vitro assays, with a reduction in biofilm formation and an increased secretion of proteases, pyocyanin and pyoverdine. Furthermore, the production of quorum sensing signal molecules was altered in this mutant. Interestingly, the gene expression profile and most phenotypic traits differ between GUN and the parental strain irrespective of the presence of glucose, suggesting a possible additional role for the deleted transporter(s). Finally, the in vivo assays demonstrated that while the virulence of all mutants in the Caenorhabditis elegans infection model was essentially comparable to that of the wild type strain, all mutants lacking kguT (i.e. the 2-ketogluconate transporter gene) and especially the double \u394gntP \u394kguT and GUN mutants, were attenuated in the Galleria mellonella model. This suggests that targeting glucose metabolism with specific drugs may alter P. aeruginosa pathogenicity depending on the ability of this pathogen to adapt to the specific nutritional context encountered at the site of infection. Therefore, a deeper knowledge of host-pathogen metabolic transactions is pivotal to develop effective antibacterial strategies based on hampering bacterial metabolic functions

Exploring the effectiveness of a rapid participatory method in mapping the role of agricultural biodiversity in local food systems to identify potential entry points to improve peoples' capabilities to be nutrition secure

Vietnam has improved its food security situation at the price of losing traditional varieties adapted to local tastes and conditions due to increased production and use of high-yield crop varieties (mainly large scale mono-cropping). Decreased diversity of foods is a growing issue, and the existence of malnutrition in regions where staple crop production and food availability are sufficient highlights the necessity for a multidisciplinary approach to design interventions that target the four dimensions of food security - availability, access, stability and utilisation, of a diverse group of locally available nutritious foods. This paper demonstrates the use of the Four Cell Approach to participatory research to rapidly identify trends in a) species and food usage in a landscape and b) the dynamics of the food system which may contribute to, or be leveraged to improve, peoples' capabilities to be nutrition secure. In the case of Son La Province, Vietnam, explored in this paper, it is possible to conclude a positive relationship between increased production diversity and dietary diversity, and highlight the importance of markets for diffusion of diverse foods. Diversification of species production and consumption has a positive effect on resilience during the off-season, and diversification of coping mechanisms utilised by communities can build resilience for future shocks. Food-based approaches that promote diversified production and consumption of locally available nutritious foods that local people value, while ensuring access to markets, have the potential to improve capabilities of local people to provide nutritious foods for themselves and their communities while improving resilience to be food and nutrition secure at all time

Neutron radiography study of laboratory ageing and treatment applications with stone consolidants

A nano-silica consolidant and nano-titania modified tetraethyl-orthosilicate were applied on two building stones, a carbonate and a silicate, by brush, poultice or capillary absorption. Neutron radiography was used to monitor capillary water absorption, and to analyse changes in physical properties caused by heat treatment of specimens for the purposes of artificially ageing and different treatment applications with stone consolidants. Moreover, ultrasonic pulse velocity and gravimetrically determined water absorption were analysed to cross-validate neutron radiography. The results reveal that reactive systems like tetraethyl-orthosilicates need an unknown period for polymerisation, which makes nano-silica consolidants more favourable for construction follow-up work. While polymerisation is incomplete, hydrophobic behaviour, water trapping and pore clogging are evident. Within the tetraethyl-orthosilicate treatment, poultice and brushing are strongly influenced by the applicant, which results in wide ranging amounts of water absorbed and anomalous water distributions and kinetics. The carbonate lithotype displays polymerisation initiated in the core of the specimen, while the lateral surfaces are still mostly hydrophobic. Reaction time differences can be attributed to the different amounts of consolidants applied, which is a result of the chosen application settings. Artificial ageing of stone specimens is a prerequisite when mechanical strength gain is studied, as demonstrated by sound speed propagation

Integrated systems research for sustainable smallholder agriculture in the Central Mekong: Achievements and challenges of implementing integrated systems research

Internal Revie

Acute subdural hematoma in the elderly. outcome analysis in a retrospective multicentric series of 213 patients

OBJECTIVE: The objective of this study was to analyze the risk factors associated with the outcome of acute subdural hematoma (ASDH) in elderly patients treated either surgically or nonsurgically. METHODS: The authors performed a retrospective multicentric analysis of clinical and radiological data on patients aged ≥ 70 years who had been consecutively admitted to the neurosurgical department of 5 Italian hospitals for the management of posttraumatic ASDH in a 3-year period. Outcome was measured according to the Glasgow Outcome Scale (GOS) at discharge and at 6 months' follow-up. A GOS score of 1-3 was defined as a poor outcome and a GOS score of 4-5 as a good outcome. Univariate and multivariate statistics were used to determine outcome predictors in the entire study population and in the surgical group. RESULTS: Overall, 213 patients were admitted during the 3-year study period. Outcome was poor in 135 (63%) patients, as 65 (31%) died during their admission, 33 (15%) were in a vegetative state, and 37 (17%) had severe disability at discharge. Surgical patients had worse clinical and radiological findings on arrival or during their admission than the patients undergoing conservative treatment. Surgery was performed in 147 (69%) patients, and 114 (78%) of them had a poor outcome. In stratifying patients by their Glasgow Coma Scale (GCS) score, the authors found that surgery reduced mortality but not the frequency of a poor outcome in the patients with a moderate to severe GCS score. The GCS score and midline shift were the most significant predictors of outcome. Antiplatelet drugs were associated with better outcomes; however, patients taking such medications had a better GCS score and better radiological findings, which could have influenced the former finding. Patients with fixed pupils never had a good outcome. Age and Charlson Comorbidity Index were not associated with outcome. CONCLUSIONS: Traumatic ASDH in the elderly is a severe condition, with the GCS score and midline shift the stronger outcome predictors, while age per se and comorbidities were not associated with outcome. Antithrombotic drugs do not seem to negatively influence pretreatment status or posttreatment outcome. Surgery was performed in patients with a worse clinical and radiological status, reducing the rate of death but not the frequency of a poor outcome

Food for thought: The underutilized potential of tropical tree-sources foods for 21st century sustainable food systems

1. The global food system is causing large-scale environmental degradation and is a major contributor to climate change. Its low diversity and failure to produce enough fruits and vegetables is contributing to a global health crisis. 2. The extraordinary diversity of tropical tree species is increasingly recognized to be vital to planetary health and especially important for supporting climate change mitigation. However, they are poorly integrated into food systems. Tropical tree diversity offers the potential for sustainable production of many foods, providing livelihood benefits and multiple ecosystem services including improved human nutrition. 3. First, we present an overview of these environmental, nutritional and livelihood benefits and show that tree-sourced foods provide important contributions to critical fruit and micronutrient (vitamin A and C) intake in rural populations based on data from sites in seven countries. 4. Then, we discuss several risks and limitations that must be taken into account when scaling-up tropical tree-based food production, including the importance of production system diversity and risks associated with supply to the global markets. 5. We conclude by discussing several interventions addressing technical, financial, political and consumer behaviour barriers, with potential to increase the consumption and production of tropical tree-sourced foods, to catalyse a transition towards more sustainable global food systems

Reducing repeat pregnancies in adolescence: applying realist principles as part of a mixed-methods systematic review to explore what works, for whom, how and under what circumstances

BACKGROUND: Previous research has demonstrated emotional, psychological and educational harm to young mothers following unintended conceptions. The UK has one of the highest rates of pregnancies in adolescence in Western Europe with a high proportion of these being repeat pregnancies, making it a topic of interest for public health policy makers, and health and social care practitioners. As part of a wider mixed-methods systematic review, realist principles were applied to synthesise evidence about interventions aiming to reduce repeat pregnancies in adolescence.METHODS: A multi-streamed, mixed-methods systematic review was conducted searching 11 major electronic databases and 9 additional databases from 1995 onwards, using key terms such as pregnancy, teen or adolescent. The principles of realist synthesis were applied to all included literature to uncover theories about what works, for whom, how and in what context. Initial theory areas were developed through evidence scoping, group discussion by the authors and stakeholder engagement to uncover context + mechanism = outcome (CMO) configurations and related narratives.RESULTS: The searches identified 8,664 documents initially, and 403 in repeat searches, filtering to 81 included studies, including qualitative studies, randomised controlled trials, quantitative studies and grey literature. Three CMO configurations were developed. The individual experiences of young mothers' triggered self-efficacy, notions of perceived risks, susceptibility and benefits of pregnancy, resulting in the adolescent taking control of their fertility and sexual encounters. The choice between motherhood and other goals triggered notions of motivations, resulting in the adolescent managing their expectations of motherhood and controlling their fertility and sexual encounters. Barriers and facilitators to accessing services triggered notions of connectedness and self-determination; resulting in interventions that are tailored so they are relevant to young persons, and improve access to services and engagement with the issue of pregnancy in adolescence.CONCLUSIONS: Pregnancy in adolescence is a complex issue with many factors to consider. The conceptual platform described here could help guide policy makers and professionals towards a number of areas that need to be attended to in order to increase the likelihood of an intervention working to prevent rapid repeat pregnancy in adolescence.TRIAL REGISTRATION: PROSPERO CRD42012003168