Murine infection with bioluminescent Leishmania infantum axenic amastigotes applied to drug discovery

Leishmaniasis is an important vector-borne neglected tropical disease caused by Leishmania parasites. Current anti-Leishmania chemotherapy is unsatisfactory, justifying the continued search for alternative treatment options. Herein, we demonstrate that luciferase-expressing Leishmania infantum axenic amastigotes, unlike promastigotes, are highly infectious to BALB/c mice and thus generate a robust bioluminescent signal in target organs, such as the liver and the spleen, as early as two weeks after infection. Treatment with the reference drugs amphotericin B and miltefosine was effective at reducing parasite burdens. This model allows the assessment of treatment efficacy using whole-mouse bioluminescence imaging without the need to wait several weeks for spleen infections to be detectable by this non-invasive method. In conclusion, we propose the use of this model in an initial approach to evaluate the treatment efficacy of promising chemical entities without having to sacrifice large numbers of animals or to wait several days for a readout.We thank Carla Oliveira from i3S for the support with the statistical analysis. Part of the content of this manuscript has been released as a pre-print in BioRxiv (https://doi.org/10.1101/326355). This work was financed by FEDER - Fundo Europeu de Desenvolvimento Regional funds through the COMPETE 2020 - Operacional Programme for Competitiveness and Internationalisation (POCI), Portugal 2020, and by Portuguese funds through FCT - Fundação para a Ciência e a Tecnologia/Ministério da Ciência, Tecnologia e Ensino Superior in the framework of the project POCI-01-0145-FEDER-031013 (PTDC/SAU-PAR/31013/2017). This work also received funds from: Norte-01-0145-FEDER-000012 “Structured program on bioengineered therapies for infectious diseases and tissue regeneration” of Norte Portugal Regional Operational Programme (NORTE 2020) under the PORTUGAL 2020 Partnership Agreement, through the European Regional Development Fund (FEDER) and through the Research Unit No. 4293; Individual funding from FCT through SFRH/BD/123734/2016 (to DC), SFRH/BD/121252/2016 (to PC) and CEECIND/02362/2017 (to JT)

Prevalence of antimicrobial resistance in enteric Escherichia coli from domestic pets and assessment of associated risk markers using a generalized linear mixed model

Antimicrobial resistance (AMR) is a growing global public health problem, which is caused by the use of antimicrobials in both human and animal medical practice. The objectives of the present cross-sectional study were as follows: (1) to determine the prevalence of resistance in Escherichia coli isolated from the feces of pets from the Porto region of Portugal against 19 antimicrobial agents and (2) to assess the individual, clinical and environmental characteristics associated with each pet as risk markers for the AMR of the E. coli isolates. From September 2009 to May 2012, rectal swabs were collected from pets selected using a systematic random procedure from the ordinary population of animals attending the Veterinary Hospital of Porto University. A total of 78 dogs and 22 cats were sampled with the objective of isolating E. coli. The animals’ owners, who allowed the collection of fecal samples from their pets, answered a questionnaire to collect information about the markers that could influence the AMR of the enteric E. coli. Chromocult tryptone bile X-glucuronide agar was used for E. coli isolation, and the disk diffusion method was used to determine the antimicrobial susceptibility. The data were analyzed using a multilevel, univariable and multivariable generalized linear mixed model (GLMM). Several (49.7%) of the 396 isolates obtained in this study were multidrug-resistant. The E. coli isolates exhibited resistance to the antimicrobial agent's ampicillin (51.3%), cephalothin (46.7%), tetracycline (45.2%) and streptomycin (43.4%). Previous quinolone treatment was the main risk marker for the presence of AMR for 12 (ampicillin, cephalothin, ceftazidime, cefotaxime, nalidixic acid, ciprofloxacin, gentamicin, tetracycline, streptomycin, chloramphenicol, trimethoprim–sulfamethoxazole and aztreonam) of the 15 antimicrobials assessed. Coprophagic habits were also positively associated with an increased risk of AMR for six drugs, ampicillin, amoxicillin–clavulanic acid, cephamycin, ciprofloxacin, streptomycin, and trimethoprim–sulfamethoxazole. In summary, pets with a record of one or more previous quinolone treatments and exhibiting coprophagic habits were at an increased risk of harboring multidrug-resistant E. coli strains in their feces compared to pets without these characteristics. AMR is a serious global problem, and assessing the risk markers for the presence of drug-resistant bacteria in pets, a very close source of resistance determinants to humans, is essential for the implementation of safe handling procedures for companion animals and for the prudent selection of antimicrobial compounds in veterinary practice

Decision Making for Inconsistent Expert Judgments Using Negative Probabilities

In this paper we provide a simple random-variable example of inconsistent information, and analyze it using three different approaches: Bayesian, quantum-like, and negative probabilities. We then show that, at least for this particular example, both the Bayesian and the quantum-like approaches have less normative power than the negative probabilities one.Comment: 14 pages, revised version to appear in the Proceedings of the QI2013 (Quantum Interactions) conferenc

The EMBLA survey - metal-poor stars in the Galactic bulge

Cosmological models predict the oldest stars in the Galaxy should be found closest to the centre of the potential well, in the bulge. The Extremely Metal-poor BuLge stars with AAOmega survey (EMBLA) successfully searched for these old, metal-poor stars by making use of the distinctive SkyMapper photometric filters to discover candidate metal-poor stars in the bulge. Their metal-poor nature was then confirmed using the AAOmega spectrograph on the Anglo-Australian Telescope. Here we present an abundance analysis of 10 bulge stars with −2.8 < [Fe/H] < −1.7 from MIKE/Magellan observations, in total determining the abundances of 22 elements. Combining these results with our previous high-resolution data taken as part of the Gaia-ESO Survey, we have started to put together a picture of the chemical and kinematic nature of the most metal-poor stars in the bulge. The currently available kinematic data are consistent with the stars belonging to the bulge, although more accurate measurements are needed to constrain the stars’ orbits. The chemistry of these bulge stars deviates from that found in halo stars of the same metallicity. Two notable differences are the absence of carbon-enhanced metal-poor bulge stars, and the α element abundances exhibit a large intrinsic scatter and include stars which are underabundant in these typically enhanced elements.LMH and MA have been supported by the Australian Research Council (grant FL110100012). ARC acknowledges support from the European Union FP7 programme through ERC grant number 320360. DY is supported through an Australian Research Council Future Fellowship (FT140100554). Research on metal-poor stars with SkyMapper is supported through Australian Research Council Discovery Projects grants DP120101237 and DP150103294 (PI: Da Costa). This publication makes use of data products from the Two Micron All Sky Survey, which is a joint project of the University of Massachusetts and the Infrared Processing and Analysis Center/California Institute of Technology, funded by the National Aeronautics and Space Administration and the National Science Foundation. This paper includes data gathered with the 6.5 metre Magellan Telescopes located at Las Campanas Observatory, Chile.This is the final version of the article. It first appeared from Oxford University Press via http://dx.doi.org/10.1093/mnras/stw100

Quantifying functional consequences of habitat degradation on a Caribbean coral reef

This is the final version. Available on open access from the European Geosciences Union via the DOI in this recordCode and data availability: Data and R code will be made available on requestCoral reefs are declining worldwide. The abundance of corals has decreased alongside a rise of filter feeders, turf, and algae in response to intensifying human pressures. This shift in prevalence of functional groups alters the biogeochemical processes in tropical water ecosystems, thereby influencing reef functioning. An urgent challenge is to understand the functional consequences of these shifts to develop suitable management strategies that aim at preserving the biological functions of reefs. Here, we quantify biogeochemical processes supporting key reef functions (i.e. net community calcification (NCC) and production (NCP) and nutrient recycling) in situ for five different benthic assemblages currently dominating shallow degraded Caribbean reef habitats. To this end, a transparent custom-made enclosure was placed over communities dominated by either one of five functional groups - coral, turf and macroalgae, bioeroding sponges, cyanobacterial mats, or sand - to determine chemical fluxes between these communities and the overlying water, during both day and night. To account for the simultaneous influence that distinct biogeochemical processes have on measured variables, the rates were then derived by solving a model consisting of differential equations describing the contribution of each process to the measured chemical fluxes. Inferred rates were low compared to those known for reef flats worldwide. Reduced accretion potential was recorded, with negative or very modest net community calcification rates for all communities. Net production during the day was also low, suggesting limited accumulation of biomass through photosynthesis and remineralisation of organic matter at night was relatively high in comparison, resulting in net heterotrophy over the survey period for most communities. Estimated recycling processes (i.e. nitrification and denitrification) were high but did not fully counterbalance nutrient release from aerobic mineralisation, rendering all substrates sources of nitrogen. Results suggest similar directions and magnitudes of key biogeochemical processes of distinct communities on this shallow Curaçaoan reef. We infer that the amount and type of organic matter released by abundant algal turfs and cyanobacterial mats on this reef likely enhances heterotroph activity and stimulates the proliferation of less diverse copiotrophic microbial populations, rendering the studied reef net heterotrophic and drawing the biogeochemical "behaviour"of distinct communities closer to each other

JAK2 V617F Mutation Prevalence in Myeloproliferative Neoplasms in Pernambuco, Brazil

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Background: The JAK2 V617F mutation is associated with three myeloproliferative neoplasms (MPNs): polycythemia vera (PV), essential thrombocythemia (ET), and primary myelofibrosis (PMF). It generates an unregulated clonal hematopoietic progenitor and leads to abnormal increased proliferation of one or more myeloid lineages. Subjects bearing this mutation may present more frequently with complications such as thrombosis and bleeding, and no specific treatment has yet been developed for BCR-ABL-negative JAK2 V617F-negative MPNs. Aims: To determine the prevalence of JAK2 V617F in MPNs in Pernambuco, Brazil, and to compare it with previous studies. Material and Methods: 144 blood samples were collected at the Hospital of Hematology of the HEMOPE Foundation and were genotyped by polymerase chain reaction-restriction fragment length polymorphism with BsaXI enzymatic digestion. Results and Discussion: 88% (46/52) of the patients with PV, 47% (39/81) with ET, and 77% (8/11) with PMF were positive for JAK2 V617F, while more than 35% of the individuals were JAK2 V617F-negative, confirming a high prevalence of this abnormality in MPNs, more frequently with a low mutated allele burden, similar to what has been reported in other Western countries, despite differences among methods used to detect this mutation. Screening for JAK2 V617F may allow specific management of these diseases with JAK2 inhibitors in the future and highlights the need for further studies on the pathogenesis of BCR-ABL-negative JAK2 V617F-negative MPNs.167802805Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Fundacao de Amparo a Ciencia e Tecnologia do Estado de Pernambuco (FACEPE)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq

Protein profiling in hepatocellular carcinoma by label-free quantitative proteomics in two west african populations.

Background Hepatocellular Carcinoma is the third most common cause of cancer related death worldwide, often diagnosed by measuring serum AFP; a poor performance stand-alone biomarker. With the aim of improving on this, our study focuses on plasma proteins identified by Mass Spectrometry in order to investigate and validate differences seen in the respective proteomes of controls and subjects with LC and HCC. Methods Mass Spectrometry analysis using liquid chromatography electro spray ionization quadrupole time-of-flight was conducted on 339 subjects using a pooled expression profiling approach. ELISA assays were performed on four significantly differentially expressed proteins to validate their expression profiles in subjects from the Gambia and a pilot group from Nigeria. Results from this were collated for statistical multiplexing using logistic regression analysis. Results Twenty-six proteins were identified as differentially expressed between the three subject groups. Direct measurements of four; hemopexin, alpha-1-antitrypsin, apolipoprotein A1 and complement component 3 confirmed their change in abundance in LC and HCC versus control patients. These trends were independently replicated in the pilot validation subjects from Nigeria. The statistical multiplexing of these proteins demonstrated performance comparable to or greater than ALT in identifying liver cirrhosis or carcinogenesis. This exercise also proposed preliminary cut offs with achievable sensitivity, specificity and AUC statistics greater than reported AFP averages. Conclusions The validated changes of expression in these proteins have the potential for development into high-performance tests usable in the diagnosis and or monitoring of HCC and LC patients. The identification of sustained expression trends strengthens the suggestion of these four proteins as worthy candidates for further investigation in the context of liver disease. The statistical combinations also provide a novel inroad of analyses able to propose definitive cut-offs and combinations for evaluation of performance

Multicenter study of the natural history and therapeutic responses of patients with chikungunya, focusing on acute and chronic musculoskeletal manifestations - a study protocol from the clinical and applied research in Chikungunya (REPLICK network)

BACKGROUND: Chikungunya is associated with high morbidity and the natural history of symptomatic infection has been divided into three phases (acute, post-acute, and chronic) according to the duration of musculoskeletal symptoms. Although this classification has been designed to help guide therapeutic decisions, it does not encompass the complexity of the clinical expression of the disease and does not assist in the evaluation of the prognosis of severity nor chronic disease. Thus, the current challenge is to identify and diagnose musculoskeletal disorders and to provide the optimal treatment in order to prevent perpetuation or progression to a potentially destructive disease course. METHODS: The study is the first product of the Clinical and Applied Research Network in Chikungunya (REPLICK). This is a prospective, outpatient department-based, multicenter cohort study in Brazil. Four work packages were defined: i. Clinical research; ii) Translational Science - comprising immunology and virology streams; iii) Epidemiology and Economics; iv) Therapeutic Response and clinical trials design. Scheduled appointments on days 21 (D21) ± 7 after enrollment, D90 ± 15, D120 ± 30, D180 ± 30; D360 ± 30; D720 ± 60, and D1080 ± 60 days. On these visits a panel of blood tests are collected in addition to the clinical report forms to obtain data on socio-demographic, medical history, physical examination and questionnaires devoted to the evaluation of musculoskeletal manifestations and overall health are performed. Participants are asked to consent for their specimens to be maintained in a biobank. Aliquots of blood, serum, saliva, PAXgene, and when clinically indicated to be examined, synovial fluid, are stored at -80° C. The study protocol was submitted and approved to the National IRB and local IRB at each study site. DISCUSSION: Standardized and harmonized patient cohorts are needed to provide better estimates of chronic arthralgia development, the clinical spectra of acute and chronic disease and investigation of associated risk factors. This study is the largest evaluation of the long-term sequelae of individuals infected with CHIKV in the Brazilian population focusing on musculoskeletal manifestations, mental health, quality of life, and chronic pain. This information will both define disease burden and costs associated with CHIKV infection, and better inform therapeutic guidelines