12,025 research outputs found

    A Robust and Universal Metaproteomics Workflow for Research Studies and Routine Diagnostics Within 24 h Using Phenol Extraction, FASP Digest, and the MetaProteomeAnalyzer

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    The investigation of microbial proteins by mass spectrometry (metaproteomics) is a key technology for simultaneously assessing the taxonomic composition and the functionality of microbial communities in medical, environmental, and biotechnological applications. We present an improved metaproteomics workflow using an updated sample preparation and a new version of the MetaProteomeAnalyzer software for data analysis. High resolution by multidimensional separation (GeLC, MudPIT) was sacrificed to aim at fast analysis of a broad range of different samples in less than 24 h. The improved workflow generated at least two times as many protein identifications than our previous workflow, and a drastic increase of taxonomic and functional annotations. Improvements of all aspects of the workflow, particularly the speed, are first steps toward potential routine clinical diagnostics (i.e., fecal samples) and analysis of technical and environmental samples. The MetaProteomeAnalyzer is provided to the scientific community as a central remote server solution at www.mpa.ovgu.de.Peer Reviewe

    iDNA from terrestrial haematophagous leeches as a wildlife surveying and monitoring tool - prospects, pitfalls and avenues to be developed

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    Invertebrate-derived DNA (iDNA) from terrestrial haematophagous leeches has recently been proposed as a powerful non-invasive tool with which to detect vertebrate species and thus to survey their populations. However, to date little attention has been given to whether and how this, or indeed any other iDNA-derived data, can be combined with state-of-the-art analytical tools to estimate wildlife abundances, population dynamics and distributions. In this review, we discuss the challenges that face the application of existing analytical methods such as site-occupancy and spatial capture-recapture (SCR) models to terrestrial leech iDNA, in particular, possible violations of key assumptions arising from factors intrinsic to invertebrate parasite biology. Specifically, we review the advantages and disadvantages of terrestrial leeches as a source of iDNA and summarize the utility of leeches for presence, occupancy, and spatial capture-recapture models. The main source of uncertainty that attends species detections derived from leech gut contents is attributable to uncertainty about the spatio-temporal sampling frame, since leeches retain host-blood for months and can move after feeding. Subsequently, we briefly address how the analytical challenges associated with leeches may apply to other sources of iDNA. Our review highlights that despite the considerable potential of leech (and indeed any) iDNA as a new survey tool, further pilot studies are needed to assess how analytical methods can overcome or not the potential biases and assumption violations of the new field of iDNA. Specifically we argue that studies to compare iDNA sampling with standard survey methods such as camera trapping, and those to improve our knowledge on leech (and other invertebrate parasite) physiology, taxonomy, and ecology will be of immense future value

    What is the evidence for giving chemoprophylaxis to children or students attending the same preschool, school or college as a case of meningococcal disease?

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    We performed a systematic literature review to assess the effectiveness of chemoprophylaxis for contacts of sporadic cases of invasive meningococcal disease (IMD) in educational settings. No studies directly compared IMD risk in contacts with/without chemoprophylaxis. However, compared to the background incidence, an elevated IMD risk was identified in settings without a general recommendation for chemoprophylaxis in pre-schools [pooled risk difference (RD) 58·2/10⁵, 95% confidence interval (CI) 27·3-89·0] and primary schools (pooled RD 4·9/10⁵, 95% CI 2·9-6·9) in the ~30 days after contact with a sporadic IMD case, but not in other educational settings. Thus, limited but consistent evidence suggests the risk of IMD in pre-school contacts of sporadic IMD cases is significantly increased above the background risk, but lower than in household contacts (pooled RD for household contacts with no chemoprophylaxis vs. background incidence: 480·1/10⁵, 95% CI 321·5-639·9). We recommend chemoprophylaxis for pre-school contacts depending on an assessment of duration and closeness of contact

    Malaria parasite detection increases during pregnancy in wild chimpanzees

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    Background: The diversity of malaria parasites (Plasmodium sp.) infecting chimpanzees (Pan troglodytes) and their close relatedness with those infecting humans is well documented. However, their biology is still largely unexplored and there is a need for baseline epidemiological data. Here, the effect of pregnancy, a well-known risk factor for malaria in humans, on the susceptibility of female chimpanzees to malaria infection was investigated. Methods: A series of 384 faecal samples collected during 40 pregnancies and 36 post-pregnancies from three habituated groups of wild chimpanzees in the Tai National Park, Cote d'Ivoire, were tested. Samples were tested for malaria parasites by polymerase chain reaction (PCR) and sequencing. Data were analysed using a generalized linear mixed model. Results: Probability of malaria parasite detection significantly increased towards the end of pregnancy and decreased with the age of the mother. Conclusions: This study provides evidence that susceptibility to malaria parasite infection increases during pregnancy, and, as shown before, in younger individuals, which points towards similar dynamics of malaria parasite infection in human and chimpanzee populations and raises questions about the effects of such infections on pregnancy outcome and offspring morbidity/mortality

    Changes to cholesterol trafficking in macrophages by Leishmania parasites infection

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    Leishmania spp. are protozoan parasites that are transmitted by sandfly vectors during blood sucking to vertebrate hosts and cause a spectrum of diseases called leishmaniases. It has been demonstrated that host cholesterol plays an important role during Leishmania infection. Nevertheless, little is known about the intracellular distribution of this lipid early after internalization of the parasite. Here, pulse‐chase experiments with radiolabeled cholesteryl esterified to fatty acids bound to low‐density lipoproteins indicated that retention of this source of cholesterol is increased in parasite‐containing subcellular fractions, while uptake is unaffected. This is correlated with a reduction or absence of detectable NPC1 (Niemann–Pick disease, type C1), a protein responsible for cholesterol efflux from endocytic compartments, in the Leishmania mexicana habitat and infected cells. Filipin staining revealed a halo around parasites within parasitophorous vacuoles (PV) likely representing free cholesterol accumulation. Labeling of host cell membranous cholesterol by fluorescent cholesterol species before infection revealed that this pool is also trafficked to the PV but becomes incorporated into the parasites’ membranes and seems not to contribute to the halo detected by filipin. This cholesterol sequestration happened early after infection and was functionally significant as it correlated with the upregulation of mRNA‐encoding proteins required for cholesterol biosynthesis. Thus, sequestration of cholesterol by Leishmania amastigotes early after infection provides a basis to understand perturbation of cholesterol‐dependent processes in macrophages that were shown previously by others to be necessary for their proper function in innate and adaptive immune responses

    A genomic portrait of the emergence, evolution, and global spread of a methicillin-resistant staphylococcus aureus pandemic

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    The widespread use of antibiotics in association with high-density clinical care has driven the emergence of drug-resistant bacteria that are adapted to thrive in hospitalized patients. Of particular concern are globally disseminated methicillin-resistant Staphylococcus aureus (MRSA) clones that cause outbreaks and epidemics associated with health care. The most rapidly spreading and tenacious health-care-associated clone in Europe currently is EMRSA-15, which was first detected in the UK in the early 1990s and subsequently spread throughout Europe and beyond. Using phylogenomic methods to analyze the genome sequences for 193 S. aureus isolates, we were able to show that the current pandemic population of EMRSA-15 descends from a health-care-associated MRSA epidemic that spread throughout England in the 1980s, which had itself previously emerged from a primarily community-associated methicillin-sensitive population. The emergence of fluoroquinolone resistance in this EMRSA-15 subclone in the English Midlands during the mid-1980s appears to have played a key role in triggering pandemic spread, and occurred shortly after the first clinical trials of this drug. Genome-based coalescence analysis estimated that the population of this subclone over the last 20 yr has grown four times faster than its progenitor. Using comparative genomic analysis we identified the molecular genetic basis of 99.8% of the antimicrobial resistance phenotypes of the isolates, highlighting the potential of pathogen genome sequencing as a diagnostic tool. We document the genetic changes associated with adaptation to the hospital environment and with increasing drug resistance over time, and how MRSA evolution likely has been influenced by country-specific drug use regimens

    Intersection problem for Droms RAAGs

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    We solve the subgroup intersection problem (SIP) for any RAAG G of Droms type (i.e., with defining graph not containing induced squares or paths of length 3): there is an algorithm which, given finite sets of generators for two subgroups H,K of G, decides whether HKH \cap K is finitely generated or not, and, in the affirmative case, it computes a set of generators for HKH \cap K. Taking advantage of the recursive characterization of Droms groups, the proof consists in separately showing that the solvability of SIP passes through free products, and through direct products with free-abelian groups. We note that most of RAAGs are not Howson, and many (e.g. F_2 x F_2) even have unsolvable SIP.Comment: 33 pages, 12 figures (revised following the referee's suggestions

    PRECEPT: an evidence assessment framework for infectious disease epidemiology, prevention and control

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    Decisions in public health should be based on the best available evidence, reviewed and appraised using a rigorous and transparent methodology. The Project on a Framework for Rating Evidence in Public Health (PRECEPT) defined a methodology for evaluating and grading evidence in infectious disease epidemiology, prevention and control that takes different domains and question types into consideration. The methodology rates evidence in four domains: disease burden, risk factors, diagnostics and intervention. The framework guiding it has four steps going from overarching questions to an evidence statement. In step 1, approaches for identifying relevant key areas and developing specific questions to guide systematic evidence searches are described. In step 2, methodological guidance for conducting systematic reviews is provided; 15 study quality appraisal tools are proposed and an algorithm is given for matching a given study design with a tool. In step 3, a standardised evidence-grading scheme using the Grading of Recommendations Assessment, Development and Evaluation Working Group (GRADE) methodology is provided, whereby findings are documented in evidence profiles. Step 4 consists of preparing a narrative evidence summary. Users of this framework should be able to evaluate and grade scientific evidence from the four domains in a transparent and reproducible way.Funding Agencies|European Centre for Disease Prevention and Control (ECDC) [2012/040, 2014/008]</p

    Tolerance and safety of the potentially probiotic strain Lactobacillus rhamnosus PRSF-L477 : a randomised, double-blind placebo-controlled trial in healthy volunteers

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    In Europe, the species Lactobacillus rhamnosus is currently on the Qualified Presumption of Safety list used by the European Food Safety Authority (EFSA) for internal safety assessment, but according to the EFSA the species should remain a topic of surveillance. In the present study, the safety and tolerance of the potentially probiotic strain L. rhamnosus PRSF-L477 was investigated in a placebo-controlled double-blind volunteer trial following FAO/WHO guidelines. A total of thirty-four subjects received daily doses of 1 x 10(11) colony-forming units (cfu) of L. rhamnosus PRSF-L477 (n 17) or placebo (n 17) for a period of 3 weeks, followed by a wash-out period of another 3 weeks. A questionnaire on gastrointestinal tolerance and a diary was kept daily to record compliance throughout these 6 weeks. Faecal and blood samples were collected for microbiological and haematological analysis. The recorded gastrointestinal symptoms, defecation frequency and stool consistency were not influenced indicating that L. rhamnosus PRSF-L477 was well tolerated. The species L. rhamnosus was detected in the faeces of sixteen out of seventeen subjects of the probiotic group during the intervention period. Using pulsed-field gel electrophoresis, re-isolates of L. rhamnosus PRSF-L477 were confirmed in nine of these subjects. Antibiotic susceptibility profiles of these re-isolates were unchanged compared with PRSF-L477. No clinically relevant changes in blood parameters such as liver and kidney function and no serious adverse events appeared during and after administration. Therefore, we conclude that L. rhamnosus PRSF-L477 can safely be administrated to healthy subjects at a daily dose of 1 x 10(11) cfu

    Recombinant Envelope-Proteins with Mutations in the Conserved Fusion Loop Allow Specific Serological Diagnosis of Dengue-Infections

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    Dengue virus (DENV) is a mosquito-borne flavivirus and a major international public health concern in many tropical and sub-tropical areas worldwide. DENV is divided into four major serotypes, and infection with one serotype leads to immunity against the same, but not the other serotypes. The specific diagnosis of DENV-infections via antibody-detection is problematic due to the high degree of cross-reactivity displayed by antibodies against related flaviviruses, such as West Nile virus (WNV), Yellow Fever virus (YFV) or Tick-borne encephalitis virus (TBEV). Especially in areas where several flaviviruses co-circulate or in the context of vaccination e.g. against YFV or TBEV, this severely complicates diagnosis and surveillance. Most flavivirus cross-reactive antibodies are produced against the highly conserved fusion loop (FL) domain in the viral envelope (E) protein. We generated insect-cell derived recombinant E-proteins of the four DENV-serotypes which contain point mutations in the FL domain. By using specific mixtures of these mutant antigens, cross-reactivity against heterologous flaviviruses was strongly reduced, enabling sensitive and specific diagnosis of the DENV-infected serum samples in IgG and IgM-measurements. These results have indications for the development of serological DENV-tests with improved specificity
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