NLRP3 (NALP3, Cryopyrin) Facilitates In Vivo Caspase-1 Activation, Necrosis, and HMGB1 Release via Inflammasome-Dependent and -Independent Pathways

Bacterial infection elicits a range of beneficial as well as detrimental host inflammatory responses. Key among these responses are macrophage/monocyte necrosis, release of the pro-inflammatory factor high-mobility group box 1 protein (HMGB1), and induction of the cytokine IL-1. While the control of IL-1β has been well-studied, processes that control macrophage cell death and HMGB-1 release in animals are poorly understood. This study utilizes Klebsiella pneumonia as a model organism since it elicits all three responses in vivo. The regulation of these responses is studied in the context of the inflammasome components, NLRP3 and ASC, which are important for caspase-1 activation and IL-1β release. Using a pulmonary infection model that reflects human infection, we show that K. pneumonia-induced mouse macrophage necrosis, HMGB-1 and IL-1β release are dependent on NLRP3 and ASC. K. pneumoniae infection of mice lacking Nlrp3 results in decreased lung inflammation and reduced survival relative to control indicating the overall protective role of this gene. Macrophage/monocyte necrosis and HMGB1 release are controlled independently of caspase-1 suggesting that the former two responses are separable from inflammasome-associated functions. These results provide critical in vivo validation that the physiologic role of NLRP3 and ASC is not limited to inflammasome formation

Optimising species detection probability and sampling effort in lake fish eDNA surveys

Environmental DNA (eDNA) metabarcoding is transforming biodiversity monitoring in aquatic environments. Such an approach has been developed and deployed for monitoring lake fish communities in Great Britain, where the method has repeatedly shown a comparable or better performance than conventional approaches. Previous analyses indicated that 20 water samples per lake are sufficient to reliably estimate fish species richness, but it is unclear how reduced eDNA sampling effort affects richness, or other biodiversity estimates and metrics. As the number of samples strongly influences the cost of monitoring programmes, it is essential that sampling effort is optimised for a specific monitoring objective. The aim of this project was to explore the effect of reduced eDNA sampling effort on biodiversity metrics (namely species richness and community composition) using algorithmic and statistical resampling techniques of a data set from 101 lakes, covering a wide spectrum of lake types and ecological quality. The results showed that reliable estimation of lake fish species richness could, in fact, usually be achieved with a much lower number of samples. For example, in almost 90% of lakes, 95% of complete fish richness could be detected with only 10 water samples, regardless of lake area. Similarly, other measures of alpha and beta-diversity were not greatly affected by a reduction in sample size from 20 to 10 samples. We also found that there is no significant difference in detected species richness between shoreline and offshore sampling transects, allowing for simplified field logistics. This could potentially allow the effective sampling of a larger number of lakes within a given monitoring budget. However, rare species were more often missed with fewer samples, with potential implications for monitoring of invasive or endangered species. These results should inform the design of eDNA sampling strategies, so that these can be optimised to achieve specific monitoring goals

Retrospective evaluation of whole exome and genome mutation calls in 746 cancer samples

Funder: NCI U24CA211006Abstract: The Cancer Genome Atlas (TCGA) and International Cancer Genome Consortium (ICGC) curated consensus somatic mutation calls using whole exome sequencing (WES) and whole genome sequencing (WGS), respectively. Here, as part of the ICGC/TCGA Pan-Cancer Analysis of Whole Genomes (PCAWG) Consortium, which aggregated whole genome sequencing data from 2,658 cancers across 38 tumour types, we compare WES and WGS side-by-side from 746 TCGA samples, finding that ~80% of mutations overlap in covered exonic regions. We estimate that low variant allele fraction (VAF < 15%) and clonal heterogeneity contribute up to 68% of private WGS mutations and 71% of private WES mutations. We observe that ~30% of private WGS mutations trace to mutations identified by a single variant caller in WES consensus efforts. WGS captures both ~50% more variation in exonic regions and un-observed mutations in loci with variable GC-content. Together, our analysis highlights technological divergences between two reproducible somatic variant detection efforts

Developing DNA Based Methods for Environmental Regulation and Management- UK Insights

DNA assessments are revolutionising biomonitoring opportunities across the globe, including the monitoring of rare and invasive species, creating biodiversity inventories, and developing pollution diagnostic and ecosystem resilience assessment methods. To date pollution and ecosystem resilience assessments have been based on assessing the diversity of familiar taxonomic groups but the introduction of DNA based methods will significantly increase the opportunities to exploit groups not previously used for this work.Environmental regulators and managers can derive many benefits from the adoption of these methods, such as improved understanding of environmental conditions, cost effective sample processing, overcoming taxonomic bottlenecks, either through shortages in trained taxonomists or utilising biota with challenging taxonomies. In addition to creating diversity based metrics DNA monitoring also allows for the assessment of functional attributes such as those that support important ecosystem services. The UK has been an early adopter of this technology and this paper will explore how the alignment of scientific advances have coincided with operational needs to create a fertile arena for the development of DNA based assessment methods that will be used in environmental regulation and management. Development projects advanced in the UK will be examined to identify the common and specific issues associated with them that have led to early engagement and adoption

Water Framework Directive Intercalibration Technical Report: Northern Lake Fish fauna ecological assessment methods

One of the key actions identified by the Water Framework Directive (WFD; 2000/60/EC) is to develop ecological assessment tools and carry out a European intercalibration (IC) exercise. The aim of the Intercalibration is to ensure that the values assigned by each Member State to the good ecological class boundaries are consistent with the Directive’s generic description of these boundaries and comparable to the boundaries proposed by other MS. In total, 83 lake assessment methods were submitted for the 2nd phase of the WFD intercalibration (2008-2012) and 62 intercalibrated and included in the EC Decision on Intercalibration (EC 2013). The intercalibration was carried out in the 13 Lake Geographical Intercalibration Groups according to the ecoregion and biological quality element. In this report we describe how the intercalibration exercise has been carried out in the Northern Lake Fish fauna group.JRC.H.1-Water Resource

Optimising species detection probability and sampling effort in lake fish eDNA surveys

Environmental DNA (eDNA) metabarcoding is transforming biodiversity monitoring in aquatic environments where the method has repeatedly shown comparable or better performance than conventional approaches to fish monitoring. This method has been developed and deployed, primarily using shoreline sampling during the winter months, across 101 lakes in Great Britain alone, covering a wide spectrum of lake types and ecological quality. Previous analyses on a subset of these lakes indicated that 20 water samples per lake are sufficient to reliably estimate fish species richness, but it is unclear how reduced eDNA sampling effort affects richness, or other biodiversity estimates and metrics. As the number of samples strongly influences the cost of monitoring programmes, it is essential that sampling effort is optimised for a specific monitoring objective. The aim of this project was to explore the effect of reduced eDNA sampling effort on biodiversity metrics (namely species richness and community composition) using algorithmic and statistical resampling techniques. The results showed that reliable estimation of lake fish species richness could in fact usually be achieved with a much lower number of samples. For example, in almost 90% of lakes, 95% of complete fish richness could be detected with only 10 water samples, regardless of lake area. Similarly other measures of alpha and beta-diversity were not greatly affected by a reduction in sample size from 20 to 10 samples. We also found that there is no significant difference in detected species richness between shoreline and offshore sampling transects, allowing for simplified field logistics. This could potentially allow the effective sampling of a larger number of lakes within a given monitoring budget. However, rare species were more often missed with fewer samples, with potential implications for monitoring of invasive or endangered species. These results should inform the design of eDNA sampling strategies, so that these can be optimised to achieve specific monitoring goals