Biology subjects

Brown, D. M.

Publications and source records attributed to Brown, D. M..

3 recordsLinked to original sources

bioRxiv2018

Programmatic Detection of Diploid-Triploid Mixoploidy via Whole Genome Sequencing

PurposeMixoploidy is a type of mosaicism where an organism is a mixture of cells with different numbers of chromosomes. There are a broad range of phenotypes associated with mixoploidy that vary greatly depending on the fraction of cells that are non-diploid, their chromosome number, their distribution, and presumably the specific variation present in the patient. Clinical detection of mixoploidy is important for diagnosis.\n\nMethodsWe developed a method to detect mixoploidy from clinical whole genome sequencing (WGS) data through the identification of excess of variant calls centered on unusual B-allele frequencies. Our method isolates the signal from these variants using trio calls and then solves a basic linear equation to estimate levels of diploid-triploid mixoploidy within the sample.\n\nResultsWe show that our method reflects the results from a cytogenetic test. We provide examples detailing how our method has been used to identify diploid-triploid mixoploid individuals from within the NIH Undiagnosed Diseases Network. We present confirmatory findings obtained by clinical cytogenetic testing and show that our method can be used to identify the diploid-triploid ratio in these cases.\n\nConclusionWGS data from patients with rare diseases can be used to identify mixoploid individuals. Individuals with certain characteristics as discussed should be tested for mixoploidy as part of standard clinical pipeline procedures. Scripts that perform this calculation are publicly available at https://github.com/HudsonAlpha/mixoviz.

genomics↗

bioRxiv2018

Contemporary circulating enterovirus D68 strains show differential viral entry and replication in human neuronal cells

Historically, enterovirus D68 (EV-D68) has primarily been associated with respiratory illnesses. However, in the summers of 2014 and 2016 EV-D68 outbreaks coincided with a spike in polio-like acute flaccid myelitis/paralysis (AFM/AFP) cases. This raised concerns that the EV-D68 virus could be the causative agent of AFM during these recent outbreaks. To assess the neurotropic capacity of EV-D68, we explored the use of the neuroblastoma-derived neuronal cell line, SH-SY5Y, as a tissue culture model to determine if differential infection permissibility is observed for different EV-D68 strains. In contrast to HeLa and A549 cells, which support viral infection of all EV-D68 strains tested, SH-SY5Y cells only supported infection by a subset of contemporary EV-D68 strains, including members from the 2014 outbreak. Viral replication and infectivity in SH-SY5Y was assessed using four different assays - infectious virus production, cytopathic effects, cellular ATP release, and VP1 capsid protein production - with similar results. Similar differential neurotropism was also observed in differentiated SH-SY5Y cells, primary human neuron cultures, and a mouse paralysis model. Using the SH-SY5Y cell culture model, we determined that barriers to viral entry was at least partly responsible for the differential infectivity phenotype, since transfection of genomic RNA into SH-SY5Y generated virions for all EV-D68 isolates, but only a single round of replication was observed from strains which could not directly infect SH-SY5Y. In addition to supporting virus replication and other functional studies, this cell culture model may help confirm epidemiological associations between EV-D68 strains and AFM and allow for the rapid identification of emerging neurotropic strains.\n\nAuthor SummarySince the outbreak during the summer of 2014, EV-D68 has been linked to a type of limb paralysis referred to as acute flaccid myelitis (AFM), with evidence mounting for the causal link of EV-D68 to AFM. Among these AFM cases, concurrent EV-D68 infection was confirmed in several independent epidemiological clusters in four continents. In this report, we describe a neuronal cell culture model (SH-SY5Y cells) where only a subset of contemporary 2014 outbreak strains of EV-D68 show infectivity in neuronal cells, or neurotropism, based on four different assays of viral replication and infection. We further confirmed the observed difference in neurotropism in vitro using primary human neuron cell cultures and in vivo with a mouse paralysis model. Using the SH-SY5Y cell model, we determined that a barrier to viral entry is at least partly responsible for neurotropism. SH-SY5Y cells may be useful in determining if specific EV-D68 genetic determinants are associated with neuropathogenesis, and replication in this cell line could be used as rapid screening tool for identifying neurotropic EV-D68 strains. This may assist with better understanding of pathogenesis and epidemiology, and with the development of potential therapies.

microbiology↗

bioRxiv2017

Analysis of the Aedes albopictus C6/36 genome provides insight into cell line adaptations to in vitro viral propagation

BackgroundThe 50-year old Aedes albopictus C6/36 cell line is a resource for the detection, amplification, and analysis of mosquito-borne viruses including Zika, dengue, and chikungunya. The cell line is derived from an unknown number of larvae from an unspecified strain of Aedes albopictus mosquitoes. Toward improved utility of the cell line for research in virus transmission, we present an annotated assembly of the C6/36 genome.\n\nResultsThe C6/36 genome assembly has the largest contig N50 (3.3 Mbp) of any mosquito assembly, presents the sequences of both haplotypes for most of the diploid genome, reveals independent null mutations in both alleles of the Dicer locus, and indicates a male-specific genome. Gene annotation was computed with publicly available mosquito transcript sequences. Gene expression data from cell line RNA sequence identified enrichment of growth-related pathways and conspicuous deficiency in aquaporins and inward rectifier K+ channels. As a test of utility, RNA sequence data from Zika-infected cells was mapped to the C6/36 genome and transcriptome assemblies. Host subtraction reduced the data set by 89%, enabling faster characterization of non-host reads.\n\nConclusionsThe C6/36 genome sequence and annotation should enable additional uses of the cell line to study arbovirus vector interactions and interventions aimed at restricting the spread of human disease.

genomics↗