Biology subjects

Hiebert, J.

Publications and source records attributed to Hiebert, J..

3 recordsLinked to original sources

bioRxiv2026

Measles Whole Genome Sequencing by an Illumina Tiled Amplification Method

Measles virus remains a significant global health threat, and despite the availability of an effective vaccine, measles cases continue to increase worldwide in recent years. Genomic surveillance has become an essential tool for monitoring virus circulation and investigating outbreaks. Here, we describe a wet-laboratory method for whole-genome sequencing of measles virus using a tiled amplicon approach and Illumina sequencing technology. A previously published Oxford Nanopore-based tiled primer scheme was adapted to include both circulating measles genotypes and for use on the Illumina platform. Two Illumina library preparation kits, Illumina DNA Prep (IDP) and Nextera XT (XT), were evaluated for performance. The IDP kit demonstrated more complete genomes and consistent genome coverage compared with XT. Using quantified reference genomes, the limit of detection was determined to be 10,000 genome copies for genotype B3 and D8. Sequence accuracy was evaluated using previously characterized clinical samples and showed high concordance. This method provides a reliable and sensitive approach for measles virus whole-genome sequencing using Illumina platforms and is suitable for genomic surveillance applications.

genomics↗

bioRxiv2026

Evaluation of MeaSeq: comprehensive analysis and reporting of measles virus whole genome sequences.

Although vaccine-preventable, measles virus (MeV) continues to pose a significant public health challenge, with a substantial resurgence of cases worldwide. As whole-genome sequencing (WGS) becomes increasingly affordable and routinely adopted in public health laboratories, reliable and accessible analysis of next-generation sequencing (NGS) data is critical for outbreak investigation and molecular surveillance. Here, we present MeaSeq, a fast, user-friendly, open-source bioinformatics pipeline for MeV analysis using Illumina or Oxford Nanopore Technologies (ONT) NGS data. MeaSeq performs quality control assessments, consensus genome assembly and variant detection, optional genotype-specific reference selection, Distinct Sequence Identifier (DSId) assignment via user-provided databases or hashing, sub-consensus variant visualization, genome quality assessment, and standardized HTML reporting. We compared the performance of MeaSeq on NGS data generated from multiple sequencing platforms and targeted enrichment strategies against gold-standard Sanger data, reference genomes, and publicly available comparative data. This validation demonstrates that MeaSeq provides an accurate, reproducible, and accessible solution for routine MeV WGS analysis, supporting genomic surveillance and outbreak response workflows in public health and research settings. Impact StatementThe recent surge in measles cases worldwide, causing several countries to lose their measles elimination status, underscores the urgent need for effective and accessible genomic surveillance. Our manuscript introduces MeaSeq, a comprehensive and open-source bioinformatics pipeline specifically designed for analyzing MeV NGS data. MeaSeq includes MeV specific analyses such as genotype prediction from sequencing reads with optional genotype-specific reference selection; DSId assignment; quality control checks such as genome rule-of-six divisibility and gene CDS validation; subconsensus nucleotide analysis with mixed-site highlighting; and genomic plotting. By leveraging NGS technology, our pipeline can facilitate the identification of transmission chains and may provide critical insights into the dynamics of MeV outbreaks. This information is essential for public health officials and researchers to implement targeted interventions and optimize vaccine strategies. Additionally, the open-source nature of MeaSeq fosters collaboration and innovation within the scientific measles community along with providing access to a wider range of researchers. Data SummaryThe MeaSeq pipeline code is available on GitHub (https://github.com/phac-nml/measeq). Comparative datasets of publicly available WGS data were accessed through the NCBI Sequence Read Archive under the following BioProjects: PRJNA869081 (https://www.ncbi.nlm.nih.gov/bioproject/PRJNA869081) PRJNA480551 (https://www.ncbi.nlm.nih.gov/bioproject/PRJNA480551) PRJNA1017431 (https://www.ncbi.nlm.nih.gov/bioproject/PRJNA1017431) PRJNA1241325 (https://www.ncbi.nlm.nih.gov/bioproject/PRJNA1241325) PRJNA1174053 (https://www.ncbi.nlm.nih.gov/bioproject/PRJNA1174053) PRJNA1293457 (https://www.ncbi.nlm.nih.gov/bioproject/PRJNA1293457) PRJNA843031 (https://www.ncbi.nlm.nih.gov/bioproject/PRJNA843031) Whole-genome sequences were included in the validation analysis if they consisted of paired-end data (Illumina) and achieved [≥]95% genome completeness following trimming of the 5' and 3' untranslated regions (UTRs). This criterion ensured sufficient genome coverage for robust validation while allowing for limited missing data arising from regions of low sequencing depth or amplicon dropout. A complete list of sequences included in the validation, along with their accession numbers, is provided in Supplementary Table 1.

bioinformatics↗

bioRxiv2019

Simultaneous Dual Optical Mapping of the Heart Using a Single Excitation, Single Camera Approach

BackgroundOptical mapping of transmembrane voltage and intracellular calcium is a powerful tool for investigating cardiac physiology and pathophysiology. However, simultaneous dual mapping of two fluorescent probes remains technically challenging. We introduce a novel, easy-to-use approach that requires a path splitter, single camera and excitation light to simultaneously acquire voltage and calcium signals from whole heart preparations, which can be applied to other physiological models - including neurons and isolated cardiomyocytes.\n\nResultsComplementary probes were selected that could be excited with a single wavelength light source. Langendorff-perfused hearts (rat, swine) were stained and imaged using a sCMOS camera outfitted with an optical path splitter to simultaneously acquire two emission fields at high spatial and temporal resolution. Voltage (RH237) and calcium (Rhod2) signals were acquired concurrently on a single sensor, resulting in two 384x256 images at 814 frames per second. At this frame rate, the signal-to-noise ratio was 47 (RH237) and 85 (Rhod2). Imaging experiments were performed on small rodent hearts, as well as larger pig hearts with sufficient optical signals. In separate experiments, each dye was used independently to assess crosstalk and demonstrate signal specificity. Additionally, the effect of ryanodine on myocardial calcium transients was validated - with no measurable effect on the amplitude of optical action potentials. To demonstrate spatial resolution, ventricular tachycardia was induced -resulting in the novel finding that spatially discordant calcium alternans can be present in different regions of the heart, even when electrical alternans remain concordant. The described system excels in providing a wide field of view and high spatiotemporal resolution for a variety of cardiac preparations.\n\nConclusionsWe report the first multiparametric mapping system that simultaneously acquires calcium and voltage signals from cardiac preparations, using a path splitter, single camera and excitation light. This approach eliminates the need for multiple cameras, excitation light patterning or frame interleaving. These features can aid in the adoption of dual mapping technology by the broader cardiovascular research community, and decrease the barrier of entry into panoramic heart imaging, as it reduces the number of required cameras.

bioengineering↗