An assessment of minimum sequence copy thresholds for identifying and reducing the prevalence of artefacts in dietary metabarcoding data

1. Metabarcoding provides a powerful tool for investigating biodiversity and trophic interactions, but the high sensitivity of this methodology makes it vulnerable to errors, resulting in artefacts in the final data. Metabarcoding studies thus often utilise minimum sequence copy thresholds (MSCTs) to remove artefacts that remain in datasets; however, there is no consensus on best practice for the use of MSCTs. 2. To mitigate erroneous reporting of results and inconsistencies, this study discusses and provides guidance for best-practice filtering of metabarcoding data for the ascertainment of conservative and accurate data. Several of the most commonly used MSCTs were applied to example datasets of Eurasian otter Lutra lutra and cereal crop spider (Araneae: Linyphiidae and Lycosidae) diets. 3. Changes in both the method and threshold value considerably affected the resultant data. Of the MSCTs tested, it was concluded that the optimal method for the examples given combined a sample-based threshold with removal of maximum taxon contamination, providing stringent filtering of artefacts while retaining target data. 4. Choice of threshold value differed between datasets due to variation in artefact abundance and sequencing depth, thus studies should employ controls (mock communities, negative controls with no DNA and unused MID tag combinations) to select threshold values appropriate for each individual study

DNA metabarcoding reveals introduced species predominate in the diet of a threatened endemic omnivore, Telfair's skink (Leiolopisma telfairii)

Introduced species can exert disproportionately negative effects on island ecosystems, but their potential role as food for native consumers is poorly studied. Telfair's skinks are endemic omnivores living on Round Island, Mauritius, a globally significant site of biodiversity conservation. We aimed to determine the dietary diversity and key trophic interactions of Telfair's skinks, whether introduced species are frequently consumed, and if diet composition changes seasonally between male and female skinks. We used DNA metabarcoding of skink fecal samples to identify animals (COI) and plants (ITS2) consumed by skinks. There were 389 dietary presence counts belonging to 77 dietary taxa found across the 73 Telfair's skink fecal samples. Introduced taxa were cumulatively consumed more frequently than other categories, accounting for 49.4% of all detections, compared to cryptogenic (20.6%), native (20.6%), and endemic taxa (9.5%). The most frequently consumed introduced species was the ant, Pheidole megacephala, present in 40% of samples. Blue latan palm, Latania loddigesii, was the most frequently consumed endemic species, present in 33% of samples but was only detected in the dry season, when fruits are produced. We found a strong seasonal difference in diet composition explained by the presence of certain plant species solely or primarily in one season and a marked increase in the consumption of animal prey in the dry season. Male and female skinks consumed several taxa at different frequencies. These results present a valuable perspective on the role of introduced species in the trophic network of their invaded ecosystem. Both native and introduced species provide nutritional resources for skinks, and this may have management implications in the context of species conservation and island restoration

Money spider dietary choice in pre‐ and post‐harvest cereal crops using metabarcoding

Money spiders (Linyphiidae) are an important component of conservation biological control in cereal crops, but they rely on alternative prey when pests are not abundant, such as between cropping cycles. To optimally benefit from these generalist predators, prey choice dynamics must first be understood. Money spiders and their locally available prey were collected from cereal crops 2 weeks pre‐ and post‐harvest. Spider gut DNA was amplified with two novel metabarcoding primer pairs designed for spider dietary analysis, and sequenced. The combined general and spider‐exclusion primers successfully identified prey from 15 families in the guts of the 46 linyphiid spiders screened, whilst avoiding amplification of Erigone spp. The primers show promise for application to the diets of other spider families such as Agelenidae and Pholcidae. Distinct invertebrate communities were identified pre‐ and post‐harvest, and changes in spider diet and, to a lesser extent, prey choice reflected this. Spiders were found to consume one another more than expected, indicating their propensity towards intraguild predation, but also consumed common pest families. Changes in spider prey choice may redress prey community changes to maintain a consistent dietary intake. Consistent provision of alternative prey via permanent refugia should be considered to sustain effective conservation biocontrol

Density-independent prey choice, taxonomy, life history, and web characteristics determine the diet and biocontrol potential of spiders (Linyphiidae and Lycosidae) in cereal crops

Spiders are among the dominant invertebrate predators in agricultural systems and are significant regulators of insect pests. The precise dynamics of biocontrol of pests in the field are, however, poorly understood. This study investigates how density-independent prey choice, taxonomy, life stage, sex, and web characteristics affect spider diet and biocontrol. We collected spiders in four genera of Linyphiidae (i.e., Bathyphantes, Erigone, Tenuiphantes, and Microlinyphia), and individuals from the Lycosidae genus Pardosa, and their proximate prey communities from barley fields in Wales, UK between April and September 2018. We analyzed the gut contents of 300 individual spiders using DNA metabarcoding. From the 300 spiders screened, 89 prey taxa were identified from 45 families, including a wide range of pests and predators. Thrips were the dominant prey, present in over a third of the spiders sampled, but a type IV functional response appears to reduce their predation at peak abundances. Spider diets significantly differed based on web characteristics, but this depended on the genus and sex of the spider and it was not the principal separating factor in the trophic niches of linyphiids and lycosids. Diets significantly differed between spider genera and life stages, reflected in different propensities for intraguild predation and pest predation. Adult spiders predated a greater diversity of other predators, and juveniles predated a greater diversity of pests. Overall, Tenuiphantes spp. and Bathyphantes spp. exhibited the greatest individual potential for biocontrol of the greatest diversity of pest genera. The greater trophic niche complementarity of Pardosa spp. and Erigone spp., however, suggests that their complementary predation of different pests might be of greater overall benefit to biocontrol. Sustainable agriculture should aim to optimize conditions throughout the cropping cycle for effective biocontrol, prioritizing provision for a diversity of spiders which predate a complementary diversity of pest species

Overview of the Proton-coupled MCT (SLC16A) Family of Transporters: Characterization, Function and Role in the Transport of the Drug of Abuse γ-Hydroxybutyric Acid

The transport of monocarboxylates, such as lactate and pyruvate, is mediated by the SLC16A family of proton-linked membrane transport proteins known as monocarboxylate transporters (MCTs). Fourteen MCT-related genes have been identified in mammals and of these seven MCTs have been functionally characterized. Despite their sequence homology, only MCT1–4 have been demonstrated to be proton-dependent transporters of monocarboxylic acids. MCT6, MCT8 and MCT10 have been demonstrated to transport diuretics, thyroid hormones and aromatic amino acids, respectively. MCT1–4 vary in their regulation, tissue distribution and substrate/inhibitor specificity with MCT1 being the most extensively characterized isoform. Emerging evidence suggests that in addition to endogenous substrates, MCTs are involved in the transport of pharmaceutical agents, including γ-hydroxybuytrate (GHB), 3-hydroxy-3-methyl-glutaryl-coenzyme A reductase inhibitors (statins), salicylic acid, and bumetanide. MCTs are expressed in a wide range of tissues including the liver, intestine, kidney and brain, and as such they have the potential to impact a number of processes contributing to the disposition of xenobiotic substrates. GHB has been extensively studied as a pharmaceutical substrate of MCTs; the renal clearance of GHB is dose-dependent with saturation of MCT-mediated reabsorption at high doses. Concomitant administration of GHB and l-lactate to rats results in an approximately two-fold increase in GHB renal clearance suggesting that inhibition of MCT1-mediated reabsorption of GHB may be an effective strategy for increasing renal and total GHB elimination in overdose situations. Further studies are required to more clearly define the role of MCTs on drug disposition and the potential for MCT-mediated detoxification strategies in GHB overdose

Investigating the specificity of peptide adsorption on gold using molecular dynamics simulations

We report all-atom molecular dynamics simulations following adsorption of gold-binding and non-gold-binding peptides on gold surfaces modeled with dispersive interactions. We examine the dependence of adsorption on both identity of the amino acids and mobility of the peptides. Within the limitations of the approach, results indicate that when the peptides are solvated, adsorption requires both configurational changes and local flexibility of individual amino acids. This is achieved when peptides consist mostly of random coils or when their secondary structural motifs (helices, sheets) are short and connected by flexible hinges. In the absence of solvent, only affinity for the surface is required: mobility is not important. In combination, these results suggest the barrier to adsorption presented by displacement of water molecules requires conformational sampling enabled through mobility.Fundação para a Ciência e a Tecnologia (FCT) – Programa Operacional “Ciência , Tecnologia, Inovação” – SFRH/BPD/20555/2004/0GV

Exercise training increases mitochondrial content and ex vivo mitochondrial function similarly in patients with type 2 diabetes and in control individuals

AIMS/HYPOTHESIS: We previously showed that type 2 diabetic patients are characterised by compromised intrinsic mitochondrial function. Here, we examined if exercise training could increase intrinsic mitochondrial function in diabetic patients compared with control individuals. METHODS: Fifteen male type 2 diabetic patients and 14 male control individuals matched for age, BMI and [Formula: see text] enrolled in a 12 week exercise intervention programme. Ex vivo mitochondrial function was assessed by high-resolution respirometry in permeabilised muscle fibres from vastus lateralis muscle. Before and after training, insulin-stimulated glucose disposal was examined during a hyperinsulinaemic-euglycaemic clamp. RESULTS: Diabetic patients had intrinsically lower ADP-stimulated state 3 respiration and lower carbonyl cyanide 4-(trifluoro-methoxy)phenylhydrazone (FCCP)-induced maximal oxidative respiration, both on glutamate and on glutamate and succinate, and in the presence of palmitoyl-carnitine (p < 0.05). After training, diabetic patients and control individuals showed increased state 3 respiration on the previously mentioned substrates (p < 0.05); however, an increase in FCCP-induced maximal oxidative respiration was observed only in diabetic patients (p < 0.05). The increase in mitochondrial respiration was accompanied by a 30% increase in mitochondrial content upon training (p < 0.01). After adjustment for mitochondrial density, state 3 and FCCP-induced maximal oxidative respiration were similar between groups after training. Improvements in mitochondrial respiration were paralleled by improvements in insulin-stimulated glucose disposal in diabetic patients, with a tendency for this in control individuals. CONCLUSIONS/INTERPRETATION: We confirmed lower intrinsic mitochondrial function in diabetic patients compared with control individuals. Diabetic patients increased their mitochondrial content to the same extent as control individuals and had similar intrinsic mitochondrial function, which occurred parallel with improved insulin sensitivity

The RCSB Protein Data Bank: redesigned web site and web services

The RCSB Protein Data Bank (RCSB PDB) web site (http://www.pdb.org) has been redesigned to increase usability and to cater to a larger and more diverse user base. This article describes key enhancements and new features that fall into the following categories: (i) query and analysis tools for chemical structure searching, query refinement, tabulation and export of query results; (ii) web site customization and new structure alerts; (iii) pair-wise and representative protein structure alignments; (iv) visualization of large assemblies; (v) integration of structural data with the open access literature and binding affinity data; and (vi) web services and web widgets to facilitate integration of PDB data and tools with other resources. These improvements enable a range of new possibilities to analyze and understand structure data. The next generation of the RCSB PDB web site, as described here, provides a rich resource for research and education

Empathy: a review of the concept

The inconsistent definition of empathy has had a negative impact on both research and practice. The aim of this article is to review and critically appraise a range of definitions of empathy and, through considered analysis, to develop a new conceptualisation. From the examination of 43 discrete definitions, 8 themes relating to the nature of empathy emerged: “distinguishing empathy from other concepts”; “cognitive or affective?”; “congruent or incongruent?”; “subject to other stimuli?”; “self/other distinction or merging?”; “trait or state influences?”; “has a behavioural outcome?”; and “automatic or controlled?” The relevance and validity of each theme is assessed and a new conceptualisation of empathy is offered. The benefits of employing a more consistent and complete definition of empathy are discussed

Two novel human cytomegalovirus NK cell evasion functions target MICA for lysosomal degradation

NKG2D plays a major role in controlling immune responses through the regulation of natural killer (NK) cells, αβ and γδ T-cell function. This activating receptor recognizes eight distinct ligands (the MHC Class I polypeptide-related sequences (MIC) A andB, and UL16-binding proteins (ULBP)1–6) induced by cellular stress to promote recognition cells perturbed by malignant transformation or microbial infection. Studies into human cytomegalovirus (HCMV) have aided both the identification and characterization of NKG2D ligands (NKG2DLs). HCMV immediate early (IE) gene up regulates NKGDLs, and we now describe the differential activation of ULBP2 and MICA/B by IE1 and IE2 respectively. Despite activation by IE functions, HCMV effectively suppressed cell surface expression of NKGDLs through both the early and late phases of infection. The immune evasion functions UL16, UL142, and microRNA(miR)-UL112 are known to target NKG2DLs. While infection with a UL16 deletion mutant caused the expected increase in MICB and ULBP2 cell surface expression, deletion of UL142 did not have a similar impact on its target, MICA. We therefore performed a systematic screen of the viral genome to search of addition functions that targeted MICA. US18 and US20 were identified as novel NK cell evasion functions capable of acting independently to promote MICA degradation by lysosomal degradation. The most dramatic effect on MICA expression was achieved when US18 and US20 acted in concert. US18 and US20 are the first members of the US12 gene family to have been assigned a function. The US12 family has 10 members encoded sequentially through US12–US21; a genetic arrangement, which is suggestive of an ‘accordion’ expansion of an ancestral gene in response to a selective pressure. This expansion must have be an ancient event as the whole family is conserved across simian cytomegaloviruses from old world monkeys. The evolutionary benefit bestowed by the combinatorial effect of US18 and US20 on MICA may have contributed to sustaining the US12 gene family