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Search indexed bioRxiv preprints in genomics, neuroscience, cell biology and bioinformatics. Read source abstracts and check manuscript versions; preprints are not peer reviewed.

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bioRxiv2018

Removing Contaminants from Metagenomic Databases

Metagenomic sequencing of patient samples is a very promising method for the diagnosis of human infections. Sequencing has the ability to capture all the DNA or RNA from pathogenic organisms in a human sample. However, complete and accurate characterization of the sequence, including identification of any pathogens, depends on the availability and quality of genomes for comparison. Thousands of genomes are now available, and as these numbers grow, the power of metagenomic sequencing for diagnosis should increase. However, recent studies have exposed the presence of contamination in published genomes, which when used for diagnosis increases the risk of falsely identifying the wrong pathogen.\n\nTo address this problem, we have developed a bioinformatics system for eliminating contamination as well as low-complexity genomic sequences in the draft genomes of eukaryotic pathogens. We applied this software to identify and remove human, bacterial, archaeal, and viral sequences present in a comprehensive database of all sequenced eukaryotic pathogen genomes. We also removed low-complexity genomic sequences, another source of false positives. Using this pipeline, we have produced a database of \"clean\" eukaryotic pathogen genomes for use with bioinformatics classification and analysis tools. We demonstrate that when attempting to find eukaryotic pathogens in metagenomic samples, the new database provides better sensitivity than one using the original genomes while offering a dramatic reduction in false positives.

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bioRxiv2018

GTFtools: a Python package for analyzing various modes of gene models

SummaryGene-centric bioinformatics studies frequently involve calculation or extraction of various features of genes such as gene ID mapping, GC content calculation and different types of gene lengths, through manipulation of gene models that are often annotated in GTF format and available from ENSEMBL or GENCODE database. Such computation is essential for subsequent analysis such as intron retention detection where independent introns may need to be identified, converting RNA-seq read counts to FPKM where gene length is required, and obtaining flanking regions around transcription start sites. However, to our knowledge, a software package that is dedicated to analyzing various modes of gene models directly from GTF file is not publicly available. In this work, GTFtools (implemented in Python and not dependent on any non-python third-party software), a stand-alone command-line software that provides a set of functions to analyze various modes of gene models, is provided for facilitating routine bioinformatics studies where information about gene models needs to be calculated.\n\nAvailabilityGTFtools is freely available at www.genemine.org/gtftools.php\n\nContacthongdong@csu.edu.cn.

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bioRxiv2018

MetaMap: An atlas of metatranscriptomic reads in human disease-related RNA-seq data

BackgroundWith the advent of the age of big data in bioinformatics, large volumes of data and high performance computing power enable researchers to perform re-analyses of publicly available datasets at an unprecedented scale. Ever more studies imply the microbiome in both normal human physiology and a wide range of diseases. RNA sequencing technology (RNA-seq) is commonly used to infer global eukaryotic gene expression patterns under defined conditions, including human disease-related contexts, but its generic nature also enables the detection of microbial and viral transcripts.\n\nFindingsWe developed a bioinformatic pipeline to screen existing human RNA-seq datasets for the presence of microbial and viral reads by re-inspecting the non-human-mapping read fraction. We validated this approach by recapitulating outcomes from 6 independent controlled infection experiments of cell line models and comparison with an alternative metatranscriptomic mapping strategy. We then applied the pipeline to close to 150 terabytes of publicly available raw RNA-seq data from >17,000 samples from >400 studies relevant to human disease using state-of-the-art high performance computing systems. The resulting data of this large-scale re-analysis are made available in the presented MetaMap resource.\n\nConclusionsOur results demonstrate that common human RNA-seq data, including those archived in public repositories, might contain valuable information to correlate microbial and viral detection patterns with diverse diseases. The presented MetaMap database thus provides a rich resource for hypothesis generation towards the role of the microbiome in human disease.

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bioRxiv2018

VirTect: a computational method for detecting virus species from RNA-Seq and its application in head and neck squamous cell carcinoma

Next generation sequencing (NGS) provides an opportunity to detect viral species from RNA-seq data on human tissues, but existing computational approaches do not perform optimally on clinical samples. We developed a bioinformatics method called VirTect for detecting viruses in neoplastic human tissues using RNA-seq data. Here, we used VirTect to analyze RNA-seq data from 363 HNSCC (head and neck squamous cell carcinoma) patients and identified 22 HPV-induced HNSCCs. These predictions were validated by manual review of pathology reports on histopathologic specimens. Compared to two existing prediction methods, VirusFinder and VirusSeq, VirTect demonstrated superior performance with many fewer false positives and false negatives. The majority of HPV carcinogenesis studies thus far have been performed on cervical cancer and generalized to HNSCC. Our results suggest that HPV-induced HNSCC involves unique mechanisms of carcinogenesis, so understanding these molecular mechanisms will have a significant impact on therapeutic approaches and outcomes. In summary, VirTect can be an effective solution for the detection of viruses with NGS data, and can facilitate the clinicopathologic characterization of various types of cancers with broad applications for oncology.\n\nSignificance StatementWe developed a new bioinformatics tool, and reported the new inside of HPV carcinogenesis mechanism in HPV-induced head and neck squamous cell carcinoma (HNSCC). This novel bioin-formatics tool and the new knowledge of HPV-induced HNSCC will facilitate the development of target therapies for treating HNSCC.

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bioRxiv2018

SoS Notebook: An Interactive Multi-Language Data Analysis Environment

MotivationComplex bioinformatic data analysis workflows involving multiple scripts in different languages can be difficult to consolidate, share, and reproduce. An environment that streamlines the entire processes of data collection, analysis, visualization and reporting of such multi-language analyses is currently lacking.\n\nResultsWe developed Script of Scripts (SoS) Notebook, a web-based notebook environment that allows the use of multiple scripting language in a single notebook, with data flowing freely within and across languages. SoS Notebook enables researchers to perform sophisticated bioinformatic analysis using the most suitable tools for different parts of the workflow, without the limitations of a particular language or complications of cross-language communications.\n\nAvailabilitySoS Notebook is hosted at http://vatlab.github.io/SoS/ and is distributed under a BSD license.\n\nContactbpeng@mdanderson.org

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bioRxiv2018

An analysis of current state of the art software on nanopore metagenomic data

ContextOur insight into DNA is controlled through a process called sequencing. Until recently, it was only possible to sequence DNA into short strings called \"reads\". Nanopore is a new sequencing technology to produce significantly longer reads. Using nanopore sequencing, a single molecule of DNA can be sequenced without the need for time consuming PCR amplification (polymerase chain reaction is a technique used in molecular biology to amplify a single copy or a few copies of a segment of DNA across several orders of magnitude).\n\nAimsMetagenomics is the study of genetic material recovered from environmental samples. A research team from IBERS (Institute of Biological, Environmental & Rural Sciences) at Aberystwyth University have sampled metagenomes from a coal mine in South Wales using the Nanopore MinION and given initial taxonomic (classification of organisms) summaries of the contents of the microbial community.\n\nMethodsUsing various new software aimed for metagenomic data, we are interested to discover how well current bioinformatics software works with the data-set. We will conduct analysis and research into how well these new state of the art software works with this new long read data and try out some recent new developments for such analysis.\n\nResultsMost of the software we used worked very well: we gained understanding of the ACGT count and quality of the data. However some software for bioinformatics dont seem to work with nanopore data. Furthermore, we can conclude that low quality nanopore data may actually be quite average.

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bioRxiv2018

DIMEdb: an integrated database and web service for metabolite identification in direct infusion mass spectrometery

MotivationMetabolomics involves the characterisation, identification, and quantification of small molecules (metabolites) that act as the reaction intermediates of biological processes. Over the past few years, we have seen wide scale improvements in data processing, database, and statistical analysis tools. Direct infusion mass spectrometery (DIMS) is a widely used platform that is able to produce a global fingerprint of the metabolome, without the requirement of a prior chromatographic step - making it ideal for wide scale high-throughput metabolomics analysis. In spite of these developments, metabolite identification still remains a key bottleneck in untargeted mass spectrometry-based metabolomics studies. The first step of the metabolite identification task is to query masses against a metaboite database to get putative metabolite annotations. Each existing metabolite database differs in a number of aspects including coverage, format, and accessibility - often limiting the user to a rudimentary web interface. Manually combining multiple search results for a single experiment where there may be potentially hundreds of masses to investigate becomes an incredibly arduous task.\n\nResultsTo facilitate unified access to metabolite information we have created the Direct Infusion MEtabolite database (DIMEdb), a comprehensive web-based metabolite database that contains over 80,000 metabolites sourced from a number of renowned metabolite databases of which can be utilised in the analysis and annotation of DIMS data. To demostrate the efficacy of DIMEdb, a simple use case for metabolic identification is presented. DIMEdb aims to provide a single point of access to metabolite information, and hopefully facilitate the development of much needed bioinformatic tools.\n\nAvailabilityDIMEdb is freely available at https://dimedb.ibers.aber.ac.uk.\n\nContactkeo7@aber.ac.uk\n\nSupplementary informationSupplementary data are available at Bioinformatics online.

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bioRxiv2018

Non-parametric and semi-parametric support estimation using SEquential RESampling random walks on biomolecular sequences

Non-parametric and semi-parametric resampling procedures are widely used to perform support estimation in computational biology and bioinformatics. Among the most widely used methods in this class is the standard bootstrap method, which consists of random sampling with replacement. While not requiring assumptions about any particular parametric model for resampling purposes, the bootstrap and related techniques assume that sites are independent and identically distributed (i.i.d.). The i.i.d. assumption can be an over-simplification for many problems in computational biology and bioinformatics. In particular, sequential dependence within biomolecular sequences is often an essential biological feature due to biochemical function, evolutionary processes such as recombination, and other factors.\n\nTo relax the simplifying i.i.d. assumption, we propose a new non-parametric/semi-parametric sequential resampling technique that generalizes \"Heads-or-Tails\" mirrored inputs, a simple but clever technique due to Landan and Graur. The generalized procedure takes the form of random walks along either aligned or unaligned biomolecular sequences. We refer to our new method as the SERES (or \"SEquential RESampling\") method.\n\nTo demonstrate the flexibility of the new technique, we apply SERES to two different applications - one involving aligned inputs and the other involving unaligned inputs. Using simulated and empirical data, we show that SERES-based support estimation yields comparable or typically better performance compared to state-of-the-art methods for both applications.

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bioRxiv2018

Reproducible genomics analysis pipelines with GNU Guix

In bioinformatics, as well as other computationally-intensive research fields, there is a need for workflows that can reliably produce consistent output, independent of the software environment or configuration settings of the machine on which they are executed. Indeed, this is essential for controlled comparison between different observations or for the wider dissemination of workflows. Providing this type of reproducibility, however, is often complicated by the need to accommodate the myriad dependencies included in a larger body of software, each of which generally come in various versions. Moreover, in many fields (bioinformatics being a prime example), these versions are subject to continual change due to rapidly evolving technologies, further complicating problems related to reproducibility. Here, we propose a principled approach for building analysis pipelines and managing their dependencies. As a case study to demonstrate the utility of our approach, we present a set of highly reproducible pipelines for the analysis of RNA-seq, ChIP-seq, Bisulfite-seq, and single-cell RNA-seq. All pipelines process raw experimental data, and generate reports containing publication-ready plots and figures, with interactive report elements and standard observables. Users may install these highly reproducible packages and apply them to their own datasets without any special computational expertise beyond the use of the command line. We hope such a toolkit will provide immediate benefit to laboratory workers wishing to process their own data sets or bioinformaticians seeking to automate all, or parts of, their analyses. In the long term, we hope our approach to reproducibility will serve as a blueprint for reproducible workflows in other areas. Our pipelines, along with their corresponding documentation and sample reports, are available at http://bioinformatics.mdc-berlin.de/pigx

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bioRxiv2018

HMP16SData: Efficient Access to the Human Microbiome Project through Bioconductor

Phase 1 of the NIH Human Microbiome Project (HMP) investigated 18 body subsites of 239 healthy American adults, to produce the first comprehensive reference for the composition and variation of the \"healthy\" human microbiome. Publicly-available data sets from amplicon sequencing of two 16S rRNA variable regions, with extensive controlled-access participant data, provide a reference for ongoing microbiome studies. However, utilization of these data sets can be hindered by the complex bioinformatic steps required to access, import, decrypt, and merge the various components in formats suitable for ecological and statistical analysis. The HMP16SData package provides count data for both 16S variable regions, integrated with phylogeny, taxonomy, public participant data, and controlled participant data for authorized researchers, using standard integrative Bioconductor data objects. By removing bioinformatic hurdles of data access and management, HMP16SData enables epidemiologists with only basic R skills to quickly analyze HMP data.

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bioRxiv2018

PRESTO, a new tool for integrating large-scale -omics data and discovering disease-specific signatures

BackgroundCohesive visualization and interpretation of hyperdimensional, large-scale -omics data is an ongoing challenge, particularly for biologists and clinicians involved in current highly complex sequencing studies. Multivariate studies are often better suited towards non-linear network analysis than differential expression testing. Here, we present PRESTO, a PREdictive Stochastic neighbor embedding Tool for Omics, which allows unsupervised dimensionality reduction of multivariate data matrices with thousands of subjects or conditions. PRESTO is intuitively integrated into an interactive user interface that helps to visualize the multidimensional patterns in genome-wide transcriptomic data from basic science and clinical studies.\n\nResultsPRESTO was tested with multiple input omics platforms, including microarray and proteomics from both mouse and human clinical datasets. PRESTO can analyze up to tens of thousands of genes and shows no increase in processing time with a large number of samples or patients. In complex datasets, such as those with multiple time points, several patient groups, or diverse mouse strains, PRESTO outperformed conventional methods. Core co-expressed gene networks were intuitively grouped in clusters, or gates, after dimensionality reduction and remained consistent across users. Networks were identified and assigned to physiological and pathological functions that cannot be gleaned from conventional bioinformatics analyses. PRESTO detected gene networks from the natural variations among mouse macrophages and human blood leukocytes. We applied PRESTO to clinical transcriptomic and proteomic data from large patient cohorts and detected disease-defining signatures in antibody-mediated kidney transplant rejection, renal cell carcinoma, and relapsing acute myeloid leukemia (AML). In AML, PRESTO confirmed a previously described gene signature and found a new signature of 10 genes that is highly predictive of patient outcome.\n\nConclusionsPRESTO offers an important integration of powerful bioinformatics tools with an interactive user interface that increases data analysis accessibility beyond bioinformaticians and coders. Here, we show that PRESTO out performs conventional methods, such as DE analysis, in multi-dimensional datasets and can identify biologically relevant co-expression gene networks. In paired samples or time points, co-expression networks could be compared for insight into longitudinal regulatory mechanisms. Additionally, PRESTO identified disease-specific signatures in clinical datasets with highly significant diagnostic and prognostic potential.

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bioRxiv2018

CRISPRCloud2: A cloud-based platform for deconvoluting CRISPR screen data

The simplicity and cost-effectiveness of CRISPR technology have made high-throughput pooled screening approaches available to many. However, the large amount of sequencing data derived from these studies yields often unwieldy datasets requiring considerable bioinformatic resources to deconvolute data; a feature which is simply not accessible to many wet labs. To address these needs, we have developed a cloud-based webtool CRISPRCloud2 that provides a state-of-the-art accuracy in mapping short reads to CRISPR library, a powerful statistical test that aggregates information across multiple sgRNAs targeting the same gene, a user-friendly data visualization and query interface, as well as easy linking to other CRISPR tools and bioinformatics resources for target prioritization. CRISPRCloud2 is a one-stop shop for labs analyzing CRISPR screen data.

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bioRxiv2018

SupportNet: a novel incremental learning framework through deep learning and support data

MotivationIn most biological data sets, the amount of data is regularly growing and the number of classes is continuously increasing. To deal with the new data from the new classes, one approach is to train a classification model, e.g., a deep learning model, from scratch based on both old and new data. This approach is highly computationally costly and the extracted features are likely very different from the ones extracted by the model trained on the old data alone, which leads to poor model robustness. Another approach is to fine tune the trained model from the old data on the new data. However, this approach often does not have the ability to learn new knowledge without forgetting the previously learned knowledge, which is known as the catastrophic forgetting problem. To our knowledge, this problem has not been studied in the field of bioinformatics despite its existence in many bioinformatic problems.\n\nResultsHere we propose a novel method, SupportNet, to solve the catastrophic forgetting problem efficiently and effectively. SupportNet combines the strength of deep learning and support vector machine (SVM), where SVM is used to identify the support data from the old data, which are fed to the deep learning model together with the new data for further training so that the model can review the essential information of the old data when learning the new information. Two powerful consolidation regularizers are applied to ensure the robustness of the learned model. Comprehensive experiments on various tasks, including enzyme function prediction, subcellular structure classification and breast tumor classification, show that SupportNet drastically outperforms the state-of-the-art incremental learning methods and reaches similar performance as the deep learning model trained from scratch on both old and new data.\n\nAvailabilityOur program is accessible at: https://github.com/lykaust15/SupportNet.

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bioRxiv2018

Floating search methodology for combining classification models for site recognition in DNA sequences

Recognition of the functional sites of genes, such as translation initiation sites, donor and acceptor splice sites and stop codons, is a relevant part of many current problems in bioinformatics. Recognition of the functional sites of genes is also a fundamental step in gene structure predictions in the most powerful programs. The best approaches to this type of recognition use sophisticated classifiers, such as support vector machines. However, with the rapid accumulation of sequence data, methods for combining many sources of evidence are necessary as it is unlikely that a single classifier can solve this type of problem with the best possible performance.\n\nA major issue is that the number of possible models to combine is large and the use of all of these models is impractical. In this paper, we present a framework that is based on floating search for combining as many classifiers as needed for the recognition of any functional sites of a gene. The methodology can be used for the recognition of translation initiation sites, donor and acceptor splice sites and stop codons. Furthermore, we can combine any number of classifiers that are trained on any species. The method is also scalable to large datasets, as is shown in experiments in which the whole human genome is used. The method is also applicable to other recognition tasks.\n\nWe present experiments on the recognition of these four functional sites in the human genome, which is used as the target genome, and use another 20 species as sources of evidence. The proposed methodology shows significant improvement over state-of-the-art methods for use in a thorough evaluation process. The proposed method is also able to improve heuristic selection of species to be used as sources of evidence as the search finds the most useful datasets.\n\nAuthor summaryIn this paper we present a methodology for combining many sources of information to recognize some of the most important functional sites in a genomic sequence. The functional sites of the sequences, such as, translation start sites, translation initiation sites, acceptor and donor splice sites and stop codons, play a very relevant role in many Bioinformatics tasks. Their accurate recognition is an important task by itself and also as part of gene structure prediction programs.\n\nOur approach uses a methodology usually termed in Computer Science as \"floating search\". This is a powerful heuristics applicable when the cost of evaluating each possible solution is high. The methodology is applied to the recognition of four different functional sites in the human genome using as additional sources of evidence the annotated genomes of other twenty different species.\n\nThe results show an advantage of the proposed method and also challenge the standard assumption of using only genomes not very close and not very far from the human to improve the recognition of functional sites in the human genome.

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bioRxiv2018

RBV: Read balance validator, a tool for prioritising copy number variations in germline conditions

BackgroundThe popularisation and decreased cost of genome resequencing has resulted in an increased use in molecular diagnostics. While there are a number of established and high quality bioinfomatic tools for identifying small genetic variants including single nucleotide variants and indels, currently there is no established standard for the detection of copy number variants (CNVs) from sequence data. The requirement for CNV detection from high throughput sequencing has resulted in the development of a large number of software packages. These tools typically utilise the sequence data characteristics: read depth, split reads, read pairs, and assembly-based techniques. However the additional source of information from read balance, defined as relative proportion of reads of each allele at each position, has been underutilised in the existing applications.\n\nResultsWe present Read Balance Validator (RBV), a bioinformatic tool which uses read balance for prioritisation and validation of putative CNVs. The software simultaneously interrogates nominated regions for the presence of deletions or multiplications, and can differentiate larger CNVs from diploid regions. Additionally, the utility of RBV to test for inheritance of CNVs is demonstrated in this report.\n\nConclusionsRBV is a CNV validation and prioritisation bioinformatic tool for both genome and exome sequencing available as a python package from https://github.com/whitneywhitford/RBV

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bioRxiv2018

Characterizing molecular flexibility by combining lRMSD measures

The root mean square deviation (RMSD) and the least RMSD are two widely used similarity measures in structural bioinformatics. Yet, they stem from global comparisons, possibly obliterating locally conserved motifs. We correct these limitations with the so-called combined RMSD, which mixes independent lRMSD measures, each computed with its own rigid motion. The combined RMSD can be used to compare (quaternary) structures based on motifs defined from the sequence (domains, SSE), or to compare structures based on structural motifs yielded by local structural alignment methods.\n\nWe illustrate the benefits of combined RMSD over the usual RMSD on three problems, namely (i) the analysis of conformational changes based on combined RMSD of rigid structural motifs (case study: a class II fusion protein), (ii) the calculation of structural phylogenies (case study: class II fusion proteins), and (iii) the assignment of quaternary structures for hemoglobin. Using these, we argue that the combined RMSD is a tool a choice to perform positive and negative discrimination of degree of freedom, with applications to the design of move sets and collective coordinates.\n\nCombined RMSD are available within the Structural Bioinformatics Library (http://sbl.inria.fr).

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bioRxiv2018

The Integrated Rapid Infectious Disease Analysis (IRIDA) Platform

Whole genome sequencing (WGS) is a powerful tool for public health infectious disease investigations owing to its higher resolution, greater efficiency, and cost-effectiveness over traditional genotyping methods. Implementation of WGS in routine public health microbiology laboratories is impeded by a lack of user-friendly automated and semi-automated pipelines, restrictive jurisdictional data sharing policies, and the proliferation of non-interoperable analytical and reporting systems. To address these issues, we developed the Integrated Rapid Infectious Disease Analysis (IRIDA) platform (irida.ca), a user-friendly, decentralized, open-source bioinformatics and analytical web platform to support real-time infectious disease outbreak investigations using WGS data. Instances can be independently installed on local high-performance computing infrastructure, enabling private and secure data management and analyses according to organizational policies and governance. IRIDAs data management capabilities enable secure upload, storage and sharing of all WGS data and metadata. The core platform currently includes pipelines for quality control, assembly, annotation, variant detection, phylogenetic analysis, in silico serotyping, multi-locus sequence typing, and genome distance calculation. Analysis pipeline results can be visualized within the platform through dynamic line lists and integrated phylogenomic clustering for research and discovery, and for enhancing decision-making support and hypothesis generation in epidemiological investigations. Communication and data exchange between instances are provided through customizable access controls. IRIDA complements centralized systems, empowering local analytics and visualizations for genomics-based microbial pathogen investigations. IRIDA is currently transforming the Canadian public health ecosystem and is freely available at https://github.com/phac-nml/irida and www.irida.ca.\n\nImpact StatementWhole genome sequencing (WGS) is revolutionizing infectious disease analysis and surveillance due to its cost effectiveness, utility, and improved analytical power. To date, no \"one-size-fits-all\" genomics platform has been universally adopted, owing to differences in national (and regional) health information systems, data sharing policies, computational infrastructures, lack of interoperability and prohibitive costs. The Integrated Rapid Infectious Disease Analysis (IRIDA) platform is a user-friendly, decentralized, open-source bioinformatics and analytical web platform developed to support real-time infectious disease outbreak investigations using WGS data. IRIDA empowers public health, regulatory and clinical microbiology laboratory personnel to better incorporate WGS technology into routine operations by shielding them from the computational and analytical complexities of big data genomics. IRIDA is now routinely used as part of a validated suite of tools to support outbreak investigations in Canada. While IRIDA was designed to serve the needs of the Canadian public health system, it is generally applicable to any public health and multi-jurisdictional environment. IRIDA enables localized analyses but provides mechanisms and standard outputs to enable data sharing. This approach can help overcome pervasive challenges in real-time global infectious disease surveillance, investigation and control, resulting in faster responses, and ultimately, better public health outcomes.\n\nDATA SUMMARYO_LIData used to generate some of the figures in this manuscript can be found in the NCBI BioProject PRJNA305824.\nC_LI

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bioRxiv2018

GranatumX: A community engaging and flexible software environment for single-cell analysis

We present GranatumX, a next-generation software environment for single-cell data analysis. GranatumX is inspired by the interactive web tool Granatum. It enables biologists to access the latest single-cell bioinformatics methods in a web-based graphical environment. It also offers software developers the opportunity to rapidly promote their own tools with others in customizable pipelines. The architecture of GranatumX allows for easy inclusion of plugin modules, named Gboxes, that wrap around bioinformatics tools written in various programming languages and on various platforms. GranatumX can be run on the cloud or private servers and generate reproducible results. It is a community-engaging, flexible, and evolving software ecosystem for scRNA-Seq analysis, connecting developers with bench scientists. GranatumX is freely accessible at http://garmiregroup.org/granatumx/app.

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