Abstract
The study of bacterial isolates or communities requires the analysis of the therein included plasmids in order to provide an extensive characterization of the organisms. Plasmids harboring ...resistance and virulence factors are of especial interest as they contribute to the dissemination of antibiotic resistance. As the number of newly sequenced bacterial genomes is growing a comprehensive resource is required which will allow to browse and filter the available plasmids, and to perform sequence analyses. Here, we present PLSDB, a resource containing 13 789 plasmid records collected from the NCBI nucleotide database. The web server provides an interactive view of all obtained plasmids with additional meta information such as sequence characteristics, sample-related information and taxonomy. Moreover, nucleotide sequence data can be uploaded to search for short nucleotide sequences (e.g. specific genes) in the plasmids, to compare a given plasmid to the records in the collection or to determine whether a sample contains one or multiple of the known plasmids (containment analysis). The resource is freely accessible under https://ccb-microbe.cs.uni-saarland.de/plsdb/.
Significant effort has been devoted to discovering microRNA (miRNA) disease biomarkers. In particular, miRNAs in whole blood or specific blood components are candidates for improving the diagnosis of ...diseases, including life-threatening pathologies. This review covers the challenges crucial for the translation of miRNAs in body fluids (circulating miRNAs) from a research setting into a clinical care scenario. First, we discuss the specificity of miRNA biomarkers for the diagnosis of a disease. While single miRNAs such as miR-20a, miR-21, miR-155, and miR-126 are frequently not disease specific, miRNA signatures that consist of a plurality of different miRNAs may help to improve differentiation between pathologies. Second, we discuss the degree of reproducibility and highlight selected validation studies. While single miRNA markers are often confirmed by independent studies, miRNA signatures are less frequently verified. Third, we address challenges to the profiling of miRNAs in high-throughput settings and we discuss the appropriateness of various analytical platforms and bioinformatics towards a clinical application of miRNAs. Finally, we shed light on the suitability of enriched miRNA sources, e.g. fractionation of body fluids for extracellular vesicles such as exosomes or blood cells, to develop miRNA signatures. With an increasing number of verified miRNA signatures and with the advance of matured medium-throughput approaches in clinical settings, specific miRNA markers are increasingly likely to contribute to human healthcare.
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EMUNI, FIS, FZAB, GEOZS, GIS, IJS, IMTLJ, KILJ, KISLJ, MFDPS, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, SBMB, SBNM, UKNU, UL, UM, UPUK, VKSCE, ZAGLJ
Abstract
While the number of human miRNA candidates continuously increases, only a few of them are completely characterized and experimentally validated. Toward determining the total number of true ...miRNAs, we employed a combined in silico high- and experimental low-throughput validation strategy. We collected 28 866 human small RNA sequencing data sets containing 363.7 billion sequencing reads and excluded falsely annotated and low quality data. Our high-throughput analysis identified 65% of 24 127 mature miRNA candidates as likely false-positives. Using northern blotting, we experimentally validated miRBase entries and novel miRNA candidates. By exogenous overexpression of 108 precursors that encode 205 mature miRNAs, we confirmed 68.5% of the miRBase entries with the confirmation rate going up to 94.4% for the high-confidence entries and 18.3% of the novel miRNA candidates. Analyzing endogenous miRNAs, we verified the expression of 8 miRNAs in 12 different human cell lines. In total, we extrapolated 2300 true human mature miRNAs, 1115 of which are currently annotated in miRBase V22. The experimentally validated miRNAs will contribute to revising targetomes hypothesized by utilizing falsely annotated miRNAs.
We present a human miRNA tissue atlas by determining the abundance of 1997 miRNAs in 61 tissue biopsies of different organs from two individuals collected post-mortem. One thousand three hundred ...sixty-four miRNAs were discovered in at least one tissue, 143 were present in each tissue. To define the distribution of miRNAs, we utilized a tissue specificity index (TSI). The majority of miRNAs (82.9%) fell in a middle TSI range i.e. were neither specific for single tissues (TSI > 0.85) nor housekeeping miRNAs (TSI < 0.5). Nonetheless, we observed many different miRNAs and miRNA families that were predominantly expressed in certain tissues. Clustering of miRNA abundances revealed that tissues like several areas of the brain clustered together. Considering -3p and -5p mature forms we observed miR-150 with different tissue specificity. Analysis of additional lung and prostate biopsies indicated that inter-organism variability was significantly lower than inter-organ variability. Tissue-specific differences between the miRNA patterns appeared not to be significantly altered by storage as shown for heart and lung tissue. MiRNAs TSI values of human tissues were significantly (P = 10(-8)) correlated with those of rats; miRNAs that were highly abundant in certain human tissues were likewise abundant in according rat tissues. We implemented a web-based repository enabling scientists to access and browse the data (https://ccb-web.cs.uni-saarland.de/tissueatlas).
In many research disciplines, hypothesis tests are applied to evaluate whether findings are statistically significant or could be explained by chance. The Wilcoxon-Mann-Whitney （WMW） test is among ...the most popular hypothesis tests in medicine and life science to analyze if two groups of samples are equally distributed. This nonparametric statistical homogeneity test is commonly applied in molecular diagnosis. Generally, the solution of the WMW test takes a high combinatorial effort for large sample cohorts containing a significant number of ties. Hence, P value is frequently approximated by a normal distribution. We developed EDISON-WMW, a new approach to calcu- late the exact permutation of the two-tailed unpaired WMW test without any corrections required and allowing for ties. The method relies on dynamic programing to solve the combinatorial problem of the WMW test efficiently. Beyond a straightforward implementation of the algorithm, we pre- sented different optimization strategies and developed a parallel solution. Using our program, the exact P value for large cohorts containing more than 1000 samples with ties can be calculated within minutes. We demonstrate the performance of this novel approach on randomly-generated data, benchmark it against 13 other commonly-applied approaches and moreover evaluate molec- ular biomarkers for lung carcinoma and chronic obstructive pulmonary disease （COPD）. We foundthat approximated P values were generally higher than the exact solution provided by EDISON- WMW. Importantly, the algorithm can also be applied to high-throughput omics datasets, where hundreds or thousands of features are included. To provide easy access to the multi-threaded version of EDISON-WMW, a web-based solution of our algorithm is freely available at http：// www.ccb.uni-saarland.de/software/wtest/.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Objective To determine whether microRNAs are differentially expressed in men with normal versus impaired spermatogenesis, and to find a biomarker for accurate diagnosis of male infertility. Design ...Microarray with real-time polymerase chain reaction (RT-PCR) validation. Setting University research and clinical institutes. Patient(s) Male partner of selected couples (n = 27) who were undergoing assisted reproduction techniques for infertility treatment. Intervention(s) None. Main Outcome Measure(s) Statistically significantly altered microRNA expression profiles in normozoospermic versus asthenozoospermic and oligoasthenozoospermic men. Result(s) There were 50 miRNAs up-regulated and 27 miRNAs down-regulated in asthenozoospermic males. In oligoasthenozoospermic males, 42 miRNAs were up-regulated and 44 miRNAs down-regulated when compared with normozoospermic males. The miRNAs that exhibited the highest fold changes and area under the receiver operating characteristic curve were miR-34b, miR-122, and miR-1973 in samples from asthenozoospermic men and miR-34b, miR-34b*, miR-15b, miR-34c-5p, miR-122, miR-449a, miR-1973, miR-16, and miR-19a in samples from oligoasthenozoospermic men. Furthermore, quantitative RT-PCR assays on specific miRNAs, including miR-141, miR-200a, miR-122, miR-34b, miR-34c-5p, and miR-16, yielded results that were largely consistent with the microarray data. Conclusion(s) Our results reveal an extended number of miRNAs that were differentially expressed in asthenozoospermic and oligoasthenozoospermic males compared with normozoospermic males. These data provide evidence for analysis of miRNA profiles as a future diagnosing tool for male infertility.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Information on miRNA targeting genes is growing rapidly. For high-throughput experiments, but also for targeted analyses of few genes or miRNAs, easy analysis with concise representation of results ...facilitates the work of life scientists. We developed miRTargetLink, a tool for automating respective analysis procedures that are frequently applied. Input of the web-based solution is either a single gene or single miRNA, but also sets of genes or miRNAs, can be entered. Validated and predicted targets are extracted from databases and an interaction network is presented. Users can select whether predicted targets, experimentally validated targets with strong or weak evidence, or combinations of those are considered. Central genes or miRNAs are highlighted and users can navigate through the network interactively. To discover the most relevant biochemical processes influenced by the target network, gene set analysis and miRNA set analysis are integrated. As a showcase for miRTargetLink, we analyze targets of five cardiac miRNAs. miRTargetLink is freely available without restrictions at www.ccb.uni-saarland.de/mirtargetlink.
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IZUM, KILJ, NUK, PILJ, PNG, SAZU, UL, UM, UPUK
Metagenomics-based studies of mixed microbial communities are impacting biotechnology, life sciences and medicine. Computational binning of metagenomic data is a powerful approach for the ...culture-independent recovery of population-resolved genomic sequences, i.e. from individual or closely related, constituent microorganisms. Existing binning solutions often require a priori characterized reference genomes and/or dedicated compute resources. Extending currently available reference-independent binning tools, we developed the BusyBee Web server for the automated deconvolution of metagenomic data into population-level genomic bins using assembled contigs (Illumina) or long reads (Pacific Biosciences, Oxford Nanopore Technologies). A reversible compression step as well as bootstrapped supervised binning enable quick turnaround times. The binning results are represented in interactive 2D scatterplots. Moreover, bin quality estimates, taxonomic annotations and annotations of antibiotic resistance genes are computed and visualized. Ground truth-based benchmarks of BusyBee Web demonstrate comparably high performance to state-of-the-art binning solutions for assembled contigs and markedly improved performance for long reads (median F1 scores: 70.02-95.21%). Furthermore, the applicability to real-world metagenomic datasets is shown. In conclusion, our reference-independent approach automatically bins assembled contigs or long reads, exhibits high sensitivity and precision, enables intuitive inspection of the results, and only requires FASTA-formatted input. The web-based application is freely accessible at: https://ccb-microbe.cs.uni-saarland.de/busybee.
Small non-coding RNAs, especially microRNAs, are discussed as promising biomarkers for a substantial number of human pathologies. A broad understanding in which solid tissues, cell types or body ...fluids a microRNA is expressed helps also to understand and to improve the suitability of miRNAs as non- or minimally-invasive disease markers. We recently reported the Human miRNA Tissue Atlas (
http://www.ccb.uni-saarland.de/tissueatlas
) containing 105 miRNA profiles of 31 organs from 2 corpses. We subsequently added miRNA profiles measured by others and us using the same array technology as for the first version of the Human miRNA Tissue Atlas. The latter profiles stem from 163 solid organs including lung, prostate and gastric tissue, from 253 whole blood samples and 66 fractioned blood cell isolates, from body fluids including 72 serum samples, 278 plasma samples, 29 urine samples, and 16 saliva samples and from different collection and storage conditions. While most miRNAs are ubiquitous abundant in solid tissues and whole blood, we also identified miRNAs that are rather specific for tissues. Our web-based repository now hosting 982 full miRNomes all of which are measured by the same microarray technology. The knowledge of these variant abundances of miRNAs in solid tissues, in whole blood and in other body fluids is essential to judge the value of miRNAs as biomarker.
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BFBNIB, GIS, IJS, KISLJ, NUK, PNG, UL, UM, UPUK
In many research disciplines, ordered lists are compared. One example is to compare a subset of all significant genes or proteins in a primary study to those in a replication study. Often, the top of ...the lists are compared using Venn diagrams, ore more precisely Euler diagrams (set diagrams showing logical relations between a finite collection of different sets). If different cohort sizes, different techniques or algorithms for evaluation were applied, a direct comparison of significant genes with a fixed threshold can however be misleading and approaches comparing lists would be more appropriate.
We developed DynaVenn, a web-based tool that incrementally creates all possible subsets from two or three ordered lists and computes for each combination a p-value for the overlap. Respectively, dynamic Venn diagrams are generated as graphical representations. Additionally an animation is generated showing how the most significant overlap is reached by backtracking. We demonstrate the improved performance of DynaVenn over an arbitrary cut-off approach on an Alzheimer's Disease biomarker set.
DynaVenn combines the calculation of the most significant overlap of different cohorts with an intuitive visualization of the results. It is freely available as a web service at http://www.ccb.uni-saarland.de/dynavenn.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
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