Iterative in Situ Click Chemistry Assembles a Branched Capture Agent and Allosteric Inhibitor for Akt1

We describe the use of iterative in situ click chemistry to design an Akt-specific branched peptide triligand that is a drop-in replacement for monoclonal antibodies in multiple biochemical assays. Each peptide module in the branched structure makes unique contributions to affinity and/or specificity resulting in a 200 nM affinity ligand that efficiently immunoprecipitates Akt from cancer cell lysates and labels Akt in fixed cells. Our use of a small molecule to preinhibit Akt prior to screening resulted in low micromolar inhibitory potency and an allosteric mode of inhibition, which is evidenced through a series of competitive enzyme kinetic assays. To demonstrate the efficiency and selectivity of the protein-templated in situ click reaction, we developed a novel QPCR-based methodology that enabled a quantitative assessment of its yield. These results point to the potential for iterative in situ click chemistry to generate potent, synthetically accessible antibody replacements with novel inhibitory properties

A Cocktail of Thermally Stable, Chemically Synthesized Capture Agents for the Efficient Detection of Anti-Gp41 Antibodies from Human Sera

We report on a method to improve in vitro diagnostic assays that detect immune response, with specific application to HIV-1. The inherent polyclonal diversity of the humoral immune response was addressed by using sequential in situ click chemistry to develop a cocktail of peptide-based capture agents, the components of which were raised against different, representative anti-HIV antibodies that bind to a conserved epitope of the HIV-1 envelope protein gp41. The cocktail was used to detect anti-HIV-1 antibodies from a panel of sera collected from HIV-positive patients, with improved signal-to-noise ratio relative to the gold standard commercial recombinant protein antigen. The capture agents were stable when stored as a powder for two months at temperatures close to 60°C

Shotgun metagenomic analysis of microbial communities in the surface waters of the Eastern South China Sea

Aims: The South China Sea (SCS) harbours a rich biodiversity. However, few studies have been published on its diverse communities, particularly its microbial counterparts. As key players behind many of the vital processes carried out in the ocean, microbes are the focus of this study, placing particular emphasis on community composition, structure, and function. Methodology and results: By employing next generation shotgun sequencing technologies (Illumina HiSeq2000), we assessed the taxonomic structure and functional diversity of the prokaryotic communities in surface waters collected from 3 representative sites in the Eastern SCS: Sarawak (Kuching), Sabah (Kota Kinabalu), and Philippines (Manila). Comparisons were undertaken to similar studies from coastal and open ocean environments. All 3 locations were dominated by members of the Proteobacteria (Alpha- and Gamma-) and Cyanobacteria (Synechococcus sp. and Prochlorococcus sp.). The highest proportion of Gammaproteobacteria was found in Sarawak, representing an approximate 20% of total sequences. Archaeal assemblages were made up largely of Euryarchaeota and unclassified sequences, while Crenarchaeota and Thaumarchaeota were present in much smaller proportions, except in the Philippines where Thaumarchaeota made up almost 40% of the entire taxa. Conclusion, significance and impact of study: The majority of the microbial communities adhered to a core set of functional genes across the different locations. However, differences existed particularly in Sarawak waters which are hypothesized to be due to local environmental parameters such as riverine influence. The results obtained from this study provide the first comparison of prokaryotic communities in the surface waters of the eastern SCS and will serve as a good platform for prospective studies in the field of environmental science

Reconciliation of essential process parameters for an enhanced predictability of Arctic stratospheric ozone loss and its climate interactions

Significant reductions in stratospheric ozone occur inside the polar vortices each spring when chlorine radicals produced by heterogeneous reactions on cold particle surfaces in winter destroy ozone mainly in two catalytic cycles, the ClO dimer cycle and the ClO/BrO cycle. Chlorofluorocarbons (CFCs), which are responsible for most of the chlorine currently present in the stratosphere, have been banned by the Montreal Protocol and its amendments, and the ozone layer is predicted to recover to 1980 levels within the next few decades. During the same period, however, climate change is expected to alter the temperature, circulation patterns and chemical composition in the stratosphere, and possible geo-engineering ventures to mitigate climate change may lead to additional changes. To realistically predict the response of the ozone layer to such influences requires the correct representation of all relevant processes. The European project RECONCILE has comprehensively addressed remaining questions in the context of polar ozone depletion, with the objective to quantify the rates of some of the most relevant, yet still uncertain physical and chemical processes. To this end RECONCILE used a broad approach of laboratory experiments, two field missions in the Arctic winter 2009/10 employing the high altitude research aircraft M55-Geophysica and an extensive match ozone sonde campaign, as well as microphysical and chemical transport modelling and data assimilation. Some of the main outcomes of RECONCILE are as follows: (1) vortex meteorology: the 2009/10 Arctic winter was unusually cold at stratospheric levels during the six-week period from mid-December 2009 until the end of January 2010, with reduced transport and mixing across the polar vortex edge; polar vortex stability and how it is influenced by dynamic processes in the troposphere has led to unprecedented, synoptic-scale stratospheric regions with temperatures below the frost point; in these regions stratospheric ice clouds have been observed, extending over >106km2 during more than 3 weeks. (2) Particle microphysics: heterogeneous nucleation of nitric acid trihydrate (NAT) particles in the absence of ice has been unambiguously demonstrated; conversely, the synoptic scale ice clouds also appear to nucleate heterogeneously; a variety of possible heterogeneous nuclei has been characterised by chemical analysis of the non-volatile fraction of the background aerosol; substantial formation of solid particles and denitrification via their sedimentation has been observed and model parameterizations have been improved. (3) Chemistry: strong evidence has been found for significant chlorine activation not only on polar stratospheric clouds (PSCs) but also on cold binary aerosol; laboratory experiments and field data on the ClOOCl photolysis rate and other kinetic parameters have been shown to be consistent with an adequate degree of certainty; no evidence has been found that would support the existence of yet unknown chemical mechanisms making a significant contribution to polar ozone loss. (4) Global modelling: results from process studies have been implemented in a prognostic chemistry climate model (CCM); simulations with improved parameterisations of processes relevant for polar ozone depletion are evaluated against satellite data and other long term records using data assimilation and detrended fluctuation analysis. Finally, measurements and process studies within RECONCILE were also applied to the winter 2010/11, when special meteorological conditions led to the highest chemical ozone loss ever observed in the Arctic. In addition to quantifying the 2010/11 ozone loss and to understand its causes including possible connections to climate change, its impacts were addressed, such as changes in surface ultraviolet (UV) radiation in the densely populated northern mid-latitudes

Reconciliation of essential process parameters for an enhanced predictability of Arctic stratospheric ozone loss and its climate interactions : (RECONCILE) ; activities and results

The international research project RECONCILE has addressed central questions regarding polar ozone depletion, with the objective to quantify some of the most relevant yet still uncertain physical and chemical processes and thereby improve prognostic modelling capabilities to realistically predict the response of the ozone layer to climate change. This overview paper outlines the scope and the general approach of RECONCILE, and it provides a summary of observations and modelling in 2010 and 2011 that have generated an in many respects unprecedented dataset to study processes in the Arctic winter stratosphere. Principally, it summarises important outcomes of RECONCILE including (i) better constraints and enhanced consistency on the set of parameters governing catalytic ozone destruction cycles, (ii) a better understanding of the role of cold binary aerosols in heterogeneous chlorine activation, (iii) an improved scheme of polar stratospheric cloud (PSC) processes that includes heterogeneous nucleation of nitric acid trihydrate (NAT) and ice on non-volatile background aerosol leading to better model parameterisations with respect to denitrification, and (iv) long transient simulations with a chemistry-climate model (CCM) updated based on the results of RECONCILE that better reproduce past ozone trends in Antarctica and are deemed to produce more reliable predictions of future ozone trends. The process studies and the global simulations conducted in RECONCILE show that in the Arctic, ozone depletion uncertainties in the chemical and microphysical processes are now clearly smaller than the sensitivity to dynamic variability

Uso de drogas e o aumento das infecções sexualmente transmissíveis: uma revisão sistemática: Drug use and the increase in sexually transmitted infections: a systematic review

Populações de usuários de drogas têm sido associadas a epidemias de infecções ou Infecções Sexualmente Transmissíveis, especialmente a infecção pelo HIV (que está associada a drogas injetáveis, uso de equipamentos contaminados para drogas injetáveis e sexo inseguro). A droga mais associada às DSTs é a cocaína fumável de base livre (crack), devido ao aumento dos comportamentos sexuais de risco. Diante disso, o presente estudo teve como objetivo compreender o impacto do uso de drogas no aumento das infecções sexualmente transmissíveis. Para isso, adotou-se como metodologia a revisão sistemática de literatura, realizando buscas nas bases de dados Scielo, Pubmed e BVS/Medline a partir do uso de descritores DeCS/MeSH e aplicação de critérios de inclusão e exclusão. A partir da análise e interpretação dos dados, concluiu-se que que pessoas que fazem uso abusivo de drogas lícitas ou ilícitas, sejam elas mulheres, homens, adolescentes, jovens, adultos, idosos, em situação de rua ou não, tendem a desenvolver comportamentos vulneráveis que pode resultar em IST. Somado a isso, enquanto comportamento de risco, tem-se a preferência por não usar preservativo, seja em relações sexuais com pessoas monogâmicas como com dois ou mais parceiros. Nesses casos, tanto o uso exacerbado de drogas como a falta de informação sobre comportamento sexual demonstram-se insuficientes

Alterações cognitivas na infecção pelo HIV: uma revisão sistemática: Cognitive changes in HIV infection: a systematic review

Provocada pelo vírus da imunodeficiência humana, com a síndrome da imunodeficiência adquirida, numa pessoa tem o seu sistema imunológico prejudicado, tornando-se suscetível a outras doenças e infecção. Tem-se a estimativa de que 50% dos infectados com o referido vírus podem sofrer alterações cognitivas. Diante disso, este estudo tem como objetivo refletir sobre mudanças estruturais cerebrais e comprometimento cognitivo em pacientes com HIV. Portanto, trata-se de uma revisão sistemática de literatura, desenvolvida a partir da seleção de estudos nas bases de dados Scielo, Pubmed e BVS/Medline a partir do uso de descritores DeCS/MeSH e aplicação de critérios de inclusão e exclusão. Após a análise e interpretação dos dados, concluiu-se que há uma significativa prevalência de HAND em adultos infectados por HIV, no que se refere a alterações cognitivas, especialmente entre pacientes do sexo feminino, de baixa escolaridade e renda, com diagnóstico tardio e baixa quantidade de linfócitos CD4 no início do tratamento. Entre essas pessoas, revelam-se comprometimentos quanto à memória, atenção, controle de impulsos, velocidade de processamento e motora, dentre outros

A Cocktail of Thermally Stable, Chemically Synthesized Capture Agents for the Efficient Detection of Anti-gp41 Antibodies from Human Sera and Techniques

This thesis reports on a method to improve in vitro diagnostic assays that detect immune response, with specific application to HIV-1. The inherent polyclonal diversity of the humoral immune response was addressed by using sequential in situ click chemistry to develop a cocktail of peptide-based capture agents, the components of which were raised against different, representative anti-HIV antibodies that bind to a conserved epitope of the HIV-1 envelope protein gp41. The cocktail was used to detect anti-HIV-1 antibodies from a panel of sera collected from HIV-positive patients, with improved signal-to-noise ratio relative to the gold standard commercial recombinant protein antigen. The capture agents were stable when stored as a powder for two months at temperatures close to 60°C

Data from: An immunosignature test distinguishes Trypanosoma cruzi, hepatitis B, hepatitis C and West Nile Virus seropositivity among asymptomatic blood donors

Background: The complexity of the eukaryotic parasite Trypanosoma (T.) cruzi manifests in its highly dynamic genome, multi-host life cycle, progressive morphologies and immune-evasion mechanisms. Accurate determination of infection or Chagas’ disease activity and prognosis continues to challenge researchers. We hypothesized that a diagnostic platform with higher ligand complexity than previously employed may hold value. Methodology: We applied the ImmunoSignature Technology (IST) for the detection of T. cruzi-specific antibodies among healthy blood donors. IST is based on capturing the information in an individual’s antibody repertoire by exposing their peripheral blood to a library of >100,000 position-addressable, chemically-diverse peptides. Principal findings: Initially, samples from two Chagas cohorts declared positive or negative by bank testing were studied. With the first cohort, library-peptides displaying differential binding signals between T. cruzi sero-states were used to train an algorithm. A classifier was fixed and tested against the training-independent second cohort to determine assay performance. Next, samples from a mixed cohort of donors declared positive for Chagas, hepatitis B, hepatitis C or West Nile virus were assayed on the same library. Signals were used to train a single algorithm that distinguished all four disease states. As a binary test, the accuracy of predicting T. cruzi seropositivity by IST was similar, perhaps modestly reduced, relative to conventional ELISAs. However, the results indicate that information beyond determination of seropositivity may have been captured. These include the identification of cohort subclasses, the simultaneous detection and discerning of other diseases, and the discovery of putative new antigens. Conclusions & significance: The central outcome of this study established IST as a reliable approach for specific determination of T. cruzi seropositivity versus disease-free individuals or those with other diseases. Its potential contribution for monitoring and controlling Chagas lies in IST’s delivery of higher resolution immune-state readouts than obtained with currently-used technologies. Despite the complexity of the ligand presentation and large quantitative readouts, performing an IST test is simple, scalable and reproducible