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Tracking Ebolavirus genomic drift with a resequencing microarray, verification with NGS

ABSTRACT: Tracking Ebolavirus genomic drift with a resequencing microarray, verification with NGS

PROVIDER: PRJEB43115 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	ERR5301994_1.fastq.gz	Fastqsanger.gz
	ERR5301994_2.fastq.gz	Fastqsanger.gz

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Tracking Ebolavirus genomic drift with a resequencing microarray, verification with NGS

Project description:This study evaluates a resequencing microarray designed to determine the sequence of the four major genes in the Ebolavirus genome, Nucleoprotein (NP), matrix protein (VP40), glycoprotein (GP) and polymerase (L). The array has the ability to determine the sequence of five species and strains: Zaire Ebolavirus (Mayinga and Makona), Bundibugyo Ebolavirus, Sudan Ebolavirus and Tai Forest Ebolavirus. Illumina Next Generation Sequencing verified the sequence of the Zaire Ebolavirus (Mayinga) sample.

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Tracking Ebolavirus genomic drift in a replicating virus with a resequencing microarray, verification with NGS 2

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Tracking Ebolavirus genomic drift in a replicating virus with a resequencing microarray, verification with NGS 2

Project description:This study demonstrates the ability of an Ebolavirus resequencing microarray to determine the sequence of the Zaire ebolavirus glycoprotein that has been engineered into the place of the surface protein of a recombinant vesicular stomatitis virus expressing green fluorescent protein (rVSV-EBOVgp-GFP). The rVSV-EBOVgp-GFP was cultured in VERO-E6 cells for three passages either in the presence of a monoclonal antibody that blocks infection (KZ52) or in control cultures with no antibody. Culture supernatant and cell lysate was collected before passaging and after each passage. RNA was extracted from each sample and the sequence of the Ebola glycoprotein in each sample was determined by the Ebola resequencing microarray. Illumina Next Generation Sequencing was performed on the initial virus stock, before passaging and on the third passage of the KZ52 antibody selected virus stock to validate the microarray sequence results.

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Project description:This study demonstrates the ability of a Ebolavirus resequencing microarray to determine the sequence of the Zaire ebolavirus glycoprotein that has been engineered into the place of the surface protein of a recombinant vesicular stomatitis virus expressing green fluorescent protein (rVSV-EBOVgp-GFP). The rVSV-EBOVgp-GFP was cultured in VERO-E6 cells for three passages either in the presence of a monoclonal antibody that blocks infection (KZ52) or in control cultures with no antibody. Culture supernatant and cell lysate was collected before passaging and after each passage. RNA was extracted from each sample and the sequence of the Ebola glycoprotein in each sample was determined by the Ebola resequencing microarray.

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Project description:Filoviruses are emerging pathogens that cause acute fever with high fatality rate and present a global public health threat. During the 2013-2016 Ebola virus outbreak, genome sequencing allowed the study of virus evolution, mutations affecting pathogenicity and infectivity, and tracing the viral spread. In 2018, early sequence identification of the Ebolavirus as EBOV in the Democratic Republic of the Congo supported the use of an Ebola virus vaccine. However, field-deployable sequencing methods are needed to enable a rapid public health response. Resequencing microarrays (RMA) are a targeted method to obtain genomic sequence on clinical specimens rapidly, and sensitively, overcoming the need for extensive bioinformatic analysis. This study presents the design and initial evaluation of an ebolavirus resequencing microarray (Ebolavirus-RMA) system for sequencing the major genomic regions of four Ebolaviruses that cause disease in humans. The design of the Ebolavirus-RMA system is described and evaluated by sequencing repository samples of three Ebolaviruses and two EBOV variants. The ability of the system to identify genetic drift in a replicating virus was achieved by sequencing the ebolavirus glycoprotein gene in a recombinant virus cultured under pressure from a neutralizing antibody. Comparison of the Ebolavirus-RMA results to the Genbank database sequence file with the accession number given for the source RNA and Ebolavirus-RMA results compared to Next Generation Sequence results of the same RNA samples showed up to 99% agreement.

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Correction: Tracking ebolavirus genomic drift with a resequencing microarray.

Project description:[This corrects the article DOI: 10.1371/journal.pone.0263732.].

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