Biology subjects

Jenkins, L.

Publications and source records attributed to Jenkins, L..

3 recordsLinked to original sources

bioRxiv2018

Gene fitness landscape of group A streptococcus during necrotizing myositis

Necrotizing fasciitis and myositis are devastating infections characterized by high mortality. Group A streptococcus (GAS) is a common cause of these infections, but the molecular pathogenesis is poorly understood. We report a genome-wide analysis using serotype M1 and M28 strains that identified novel GAS genes contributing to necrotizing myositis in nonhuman primates (NHP), a clinically relevant model. Using transposon directed insertion-site sequencing (TraDIS) we identified 126 and 116 GAS genes required for infection by serotype M1 and M28 organisms, respectively. For both M1 and M28 strains, more than 25% of the GAS genes required for necrotizing myositis encode known or putative transporters. Thirteen GAS transporters contributed to both M1 and M28 strain fitness in NHP myositis, including putative importers for amino acids, carbohydrates, and vitamins, and exporters for toxins, quorum sensing peptides, and uncharacterized molecules. Targeted deletion of genes encoding five transporters confirmed that each isogenic mutant strain was significantly impaired in causing necrotizing myositis in NHPs. qRT-PCR analysis showed that these five genes are expressed in infected NHP and human skeletal muscle. Certain substrate-binding lipoproteins of these transporters, such as Spy0271 and Spy1728, were previously documented to be surface-exposed, suggesting that our findings have translational research implications.

microbiology↗

bioRxiv2018

Rapid Paediatric Sequencing (RaPS): Comprehensive real-life workflow for rapid diagnosis of critically ill children

BackgroundRare genetic conditions are frequent risk factors for, or direct causes of, organ failure requiring paediatric intensive care unit (PICU) support. Such conditions are frequently suspected but unidentified at PICU admission. Compassionate and effective care is greatly assisted by definitive diagnostic information. There is therefore a need to provide a rapid genetic diagnosis to inform clinical management.\n\nTo date, Whole Genome Sequencing (WGS) approaches have proved successful in diagnosing a proportion of children with rare diseases, but results may take months to report or require the use of equipment and practices not compatible with a clinical diagnostic setting. We describe an end-to-end workflow for the use of rapid WGS for diagnosis in critically ill children in a UK National Health Service (NHS) diagnostic setting.\n\nMethodsWe sought to establish a multidisciplinary Rapid Paediatric Sequencing (RaPS) team for case selection, trio WGS, a rapid bioinformatics pipeline for sequence analysis and a phased analysis and reporting system to prioritise genes with a high likelihood of being causal. Our workflow was iteratively developed prospectively during the analysis of the first 10 children and applied to the following 14 to assess its utility.\n\nFindingsTrio WGS in 24 critically ill children led to a molecular diagnosis in ten (42%) through the identification of causative genetic variants. In three of these ten individuals (30%) the diagnostic result had an immediate impact on the individuals clinical management. For the last 14 trios, the shortest time taken to reach a provisional diagnosis was four days (median 7 days).\n\nInterpretationRapid WGS can be used to diagnose and inform management of critically ill children using widely available off the shelf products within the constraints of an NHS clinical diagnostic setting. We provide a robust workflow that will inform and facilitate the rollout of rapid genome sequencing in the NHS and other healthcare systems globally.\n\nFundingThe study was funded by NIHR GOSH/UCL BRC: ormbrc-2012-1

genomics↗

bioRxiv2017

C11orf70 mutations causing primary ciliary dyskinesia disrupt a conserved step in the intraflagellar transport-dependent assembly of multiple axonemal dyneins

Primary ciliary dyskinesia (PCD) is a genetically and phenotypically heterogeneous disorder characterized by destructive respiratory disease and laterality abnormalities due to randomised left-right body asymmetry. PCD is mostly caused by mutations affecting components of the core axoneme structure of motile cilia that are essential for cilia movement. In addition, there is a growing group of PCD genes that encode proteins essential for the assembly of the ciliary dynein motors and the active transport process that delivers them from their cytoplasmic assembly site into the axoneme. We screened a cohort of affected individuals for disease-causing mutations using a targeted next generation sequencing panel and identified 2 unrelated families (3 affected children) with mutations in the uncharacterized C11orf70 gene. The affected children share a consistent PCD phenotype from early life with laterality defects and immotile respiratory cilia displaying combined loss of inner and outer dynein arms (IDA+ODA). Phylogenetic analysis shows C11orf70 is highly conserved, distributed across species similarly to proteins involved in the intraflagellar transport (IFT)-dependant assembly of axonemal dyneins. Paramecium C11orf70 RNAi knockdown led to combined loss of ciliary IDA+ODA with reduced cilia beating and swim velocity. Fluorescently tagged C11orf70 in Paramecium and Chlamydomonas localises mainly in the cytoplasm with a small amount in the ciliary component, its abundance in the axoneme being IFT-dependant. During ciliogenesis, C11orf70 accumulates at the ciliary tips in a similar distribution to the IFT-B protein IFT46. In summary, C11orf70 is essential for IFT-dependant assembly of dynein arms and C11orf70 mutations cause defective cilia motility and PCD.

genetics↗