Single metabarcoding multiplex captures community‐level freshwater biodiversity and beyond

Cost‐effective and accurate quantification of biodiversity is important for biodiversity conservation, resource management, and forecasting. Traditional monitoring approaches have relied on direct observations, remote sensing, and mark‐recapture techniques, providing insights into species ecology and the impact of pollution and climate change on indicator species. However, these techniques are typically low throughput, expensive, and can be invasive. In addition, they cannot detect cryptic diversity and are biased toward species that leave identifiable remains. DNA‐based methods, such as metabarcoding or single marker gene assays, have enabled high throughput screening of a wide range of taxonomic groups, including ones without well‐preserved remains. When compared with traditional techniques, these approaches have high throughput, can resolve cryptic diversity, do not require taxonomic specialist skills, and are non‐invasive. However, although they are comparatively cheaper than traditional approaches, they are expensive when applied at the community‐level as single marker assays are amplified and sequenced independently. Multilocus approaches in which multiple gene markers are amplified in a single reaction are desirable to deliver community‐level assessments in a cost‐effective manner. Yet, they are uncommon because of technical challenges that may lead to biases in downstream analyses, such as index hopping and unbalanced representation of taxonomic groups. Here, we developed a highly multiplexed protocol that combines the early pooling of marker genes that target broad taxonomic groups and taxon‐specific markers in a single tube reaction. This step is followed by the pooling of up to 384 samples per locus (N = 15,636 samples) with unique dual‐indexed sequencing adapters in a single sequencing run. This approach dramatically reduces the costs of community‐level biodiversity quantification and lowers the need for input DNA without compromising output quality. We optimized the multiplex assay on lake freshwater sediment samples and benchmarked the assay on samples from other environmental matrices, demonstrating its direct application to the river and marine communities

Multigenerational exposures of Daphnia magna to pristine and aged silver nanoparticles:epigenetic changes and phenotypical ageing related effects

Engineered nanoparticles (NPs) undergo physical, chemical, and biological transformation after environmental release, resulting in different properties of the ‘aged’ versus ‘pristine’ forms. While many studies have investigated the ecotoxicological effects of silver (Ag) NPs, the majority focus on ‘pristine’ Ag NPs in simple exposure media, rather than investigating realistic environmental exposure scenarios with transformed NPs. Here, we systematically evaluate the effects of ‘pristine’ and ‘aged’ Ag NPs with different surface coatings on Daphnia magna over four generations, comparing continuous exposure versus parental only exposure to assess recovery potential for three generations. Biological endpoints including survival, growth and reproduction and genetic effects associated with Ag NP exposure were investigated. Parental exposure to ‘pristine’ Ag NPs had an inhibitory effect on reproduction, induced expression of antioxidant stress related genes and reduced survival. Pristine Ag NPs also induced morphological changes including tail losses and lipid accumulation associated with aging phenotypes in the heart, abdomen and abdominal claw. These effects were epigenetic remaining two generations post-maternal exposure (F2 and F3). Exposure to identical Ag NPs (same concentrations) aged for 6 months in environmentally realistic water containing natural organic matter showed considerably reduced toxicological effects in continuously exposed generations and to the recovery generations

Spatial estimation of groundwater quality, hydrogeochemical investigation, and health impacts of shallow groundwater in Kabul city, Afghanistan

The management of groundwater in densely populated areas with no centralized water treatment is critical for the prevention of diseases and maintaining sanitation. Here, we determine the bacteriological and chemical characteristics of groundwater in Kabul city, a resource that 4.1 million individuals rely on. Groundwater samples were analyzed from 41 newly established piezometric wells across Kabul, and data were compared with the last detailed study, undertaken in 2007, to understand contamination trends in an area that has undergone significant development and social changes. Piper diagrams, Gibbs diagrams, correlation analysis, and bivariate plots examine the hydrogeochemical and natural occurring processes of groundwater. The average concentration of cations followed the order Na+  > Mg2+  > Ca2+  > K+, and anions HCO3− > NO3− > Cl− > SO42− > F with Gibbs diagrams indicating mainly rock-weathering influence groundwater chemistry. An increase in nitrate (NO3−) and E. coli indicates anthropogenic activities impacting the shallow groundwater quality, with significantly elevated nitrate (over 50 mg/L) and E. coli (up to 250 CFU/100 mL). The increasing presence of E. coli and NO3− in the shallow groundwater of Kabul city in turn suggests problematic links to the prevalence of waterborne diseases. Additionally, the water quality index (WQI) was used to assess groundwater quality, and rank its suitability for drinking purposes. The WQI analysis showed that less than 35% of shallow groundwater samples had good water quality. The findings of this study are crucial for the development and sustainable management of groundwater in the city. In short term, we propose interventions such as point-of-use (POU) water purification which may offer temporary respite for waterborne disease prevention. Kabul city requires immediate attention to developing sustainable groundwater management policies, expansion of the water supply network, groundwater quality monitoring, and wastewater management

Remasking of Candida albicans β-Glucan in Response to Environmental pH Is Regulated by Quorum Sensing

Candida albicans is a commensal yeast of the human gut which is tolerated by the immune system but has the potential to become an opportunistic pathogen. One way in which C. albicans achieves this duality is through concealing or exposing cell wall pathogen-associated molecular patterns (PAMPs) in response to host-derived environment cues (pH, hypoxia, and lactate). This cell wall remodeling allows C. albicans to evade or hyperactivate the host’s innate immune responses, leading to disease. Previously, we showed that adaptation of C. albicans to acidic environments, conditions encountered during colonization of the female reproductive tract, induces significant cell wall remodeling resulting in the exposure of two key fungal PAMPs (β-glucan and chitin). Here, we report that this pH-dependent cell wall remodeling is time dependent, with the initial change in pH driving cell wall unmasking, which is then remasked at later time points. Remasking of β-glucan was mediated via the cell density-dependent fungal quorum sensing molecule farnesol, while chitin remasking was mediated via a small, heat-stable, nonproteinaceous secreted molecule(s). Transcript profiling identified a core set of 42 genes significantly regulated by pH over time and identified the transcription factor Efg1 as a regulator of chitin exposure through regulation of CHT2. This dynamic cell wall remodeling influenced innate immune recognition of C. albicans, suggesting that during infection, C. albicans can manipulate the host innate immune responses

The RNA helicase UPF1 associates with mRNAs co-transcriptionally and is required for the release of mRNAs from gene loci

UPF1 is an RNA helicase that is required for efficient nonsense-mediated mRNA decay (NMD) in eukaryotes, and the predominant view is that UPF1 mainly operates on the 3'UTRs of mRNAs that are directed for NMD in the cytoplasm. Here we offer evidence, obtained from Drosophila, that UPF1 constantly moves between the nucleus and cytoplasm by a mechanism that requires its RNA helicase activity. UPF1 is associated, genome-wide, with nascent RNAs at most of the active Pol II transcription sites and at some Pol III-transcribed genes, as demonstrated microscopically on the polytene chromosomes of salivary glands and by ChIP-seq analysis in S2 cells. Intron recognition seems to interfere with association and translocation of UPF1 on nascent pre-mRNAs, and cells depleted of UPF1 show defects in the release of mRNAs from transcription sites and mRNA export from the nucleus

Total haemoglobin content

Total haemoglobin content measured in µmol/L in the 30 genotypes of Daphnia magna studied here at the control temperature (20C) and after exposure to hyper-thermal stress (30C). Genotype ID, population ID, average Hb content across three replicates and log2 fold change of total Hb content between the two experimental temperatures are shown. The Hb data are From Cambronero et al. 2017