Filopodial protrusion driven by density-dependent Ena-TOCA-1 interactions.

Filopodia are narrow actin-rich protrusions with important roles in neuronal development where membrane-binding adaptor proteins, such as I-BAR- and F-BAR-domain-containing proteins, have emerged as upstream regulators that link membrane interactions to actin regulators such as formins and proteins of the Ena/VASP family. Both the adaptors and their binding partners are part of diverse and redundant protein networks that can functionally compensate for each other. To explore the significance of the F-BAR domain-containing neuronal membrane adaptor TOCA-1 (also known as FNBP1L) in filopodia we performed a quantitative analysis of TOCA-1 and filopodial dynamics in Xenopus retinal ganglion cells, where Ena/VASP proteins have a native role in filopodial extension. Increasing the density of TOCA-1 enhances Ena/VASP protein binding in vitro, and an accumulation of TOCA-1, as well as its coincidence with Ena, correlates with filopodial protrusion in vivo. Two-colour single-molecule localisation microscopy of TOCA-1 and Ena supports their nanoscale association. TOCA-1 clusters promote filopodial protrusion and this depends on a functional TOCA-1 SH3 domain and activation of Cdc42, which we perturbed using the small-molecule inhibitor CASIN. We propose that TOCA-1 clusters act independently of membrane curvature to recruit and promote Ena activity for filopodial protrusion.

Extension of the Segatella copri complex to 13 species with distinct large extrachromosomal elements and associations with host conditions.

The Segatella copri (formerly Prevotella copri) complex (ScC) comprises taxa that are key members of the human gut microbiome. It was previously described to contain four distinct phylogenetic clades. Combining targeted isolation with large-scale metagenomic analysis, we defined 13 distinct Segatella copri-related species, expanding the ScC complex beyond four clades. Complete genome reconstruction of thirteen strains from seven species unveiled the presence of genetically diverse large circular extrachromosomal elements. These elements are consistently present in most ScC species, contributing to intra- and inter-species diversities. The nine species-level clades present in humans display striking differences in prevalence and intra-species genetic makeup across human populations. Based on a meta-analysis, we found reproducible associations between members of ScC and the male sex and positive correlations with lower visceral fat and favorable markers of cardiometabolic health. Our work uncovers genomic diversity within ScC, facilitating a better characterization of the human microbiome.

Filopodia In Vitro and In Vivo.

Filopodia are dynamic cell surface protrusions used for cell motility, pathogen infection, and tissue development. The molecular mechanisms determining how and where filopodia grow and retract need to integrate mechanical forces and membrane curvature with extracellular signaling and the broader state of the cytoskeleton. The involved actin regulatory machinery nucleates, elongates, and bundles actin filaments separately from the underlying actin cortex. The refined membrane and actin geometry of filopodia, importance of tissue context, high spatiotemporal resolution required, and high degree of redundancy all limit current models. New technologies are improving opportunities for functional insight, with reconstitution of filopodia in vitro from purified components, endogenous genetic modification, inducible perturbation systems, and the study of filopodia in multicellular environments. In this review, we explore recent advances in conceptual models of how filopodia form, the molecules involved in this process, and our latest understanding of filopodia in vitro and in vivo.

Alpha heating of indirect-drive layered implosions on the National Ignition Facility.

In order to understand how close current layered implosions in indirect-drive inertial confinement fusion are to ignition, it is necessary to measure the level of alpha heating present. To this end, pairs of experiments were performed that consisted of a low-yield tritium-hydrogen-deuterium (THD) layered implosion and a high-yield deuterium-tritium (DT) layered implosion to validate experimentally current simulation-based methods of determining yield amplification. The THD capsules were designed to reduce simultaneously DT neutron yield (alpha heating) and maintain hydrodynamic similarity with the higher yield DT capsules. The ratio of the yields measured in these experiments then allowed the alpha heating level of the DT layered implosions to be determined. The level of alpha heating inferred is consistent with fits to simulations expressed in terms of experimentally measurable quantities and enables us to infer the level of alpha heating in recent high-performing implosions.

Enhanced cultured diversity of the mouse gut microbiota enables custom-made synthetic communities.

Microbiome research needs comprehensive repositories of cultured bacteria from the intestine of mammalian hosts. We expanded the mouse intestinal bacterial collection (www.dsmz.de/miBC) to 212 strains, all publicly available and taxonomically described. This includes strain-level diversity, small-sized bacteria, and previously undescribed taxa (one family, 10 genera, and 39 species). This collection enabled metagenome-educated prediction of synthetic communities (SYNs) that capture key functional differences between microbiomes, notably identifying communities associated with either resistance or susceptibility to DSS-induced colitis. Additionally, nine species were used to amend the Oligo-Mouse Microbiota (OMM)12 model, yielding the OMM19.1 model. The added strains compensated for phenotype differences between OMM12 and specific pathogen-free mice, including body composition and immune cells in the intestine and associated lymphoid tissues. Ready-to-use OMM stocks are available for future studies. In conclusion, this work improves our knowledge of gut microbiota diversity in mice and enables functional studies via the modular use of isolates.

Three-dimensional hot-spot x-ray emission tomography from cryogenic deuterium-tritium direct-drive implosions on OMEGA.

A three-dimensional model of the hot-spot x-ray emission has been developed and applied to the study of low-mode drive asymmetries in direct-drive inertial confinement fusion implosions on OMEGA with cryogenic deuterium-tritium targets. The steady-state model assumes an optically thin plasma and the data from four x-ray diagnostics along quasi-orthogonal lines of sight are used to obtain a tomographic reconstruction of the hot spot. A quantitative analysis of the hot-spot shape is achieved by projecting the x-ray emission into the diagnostic planes and comparing this projection to the measurements. The model was validated with radiation-hydrodynamic simulations assuming a mode-2 laser illumination perturbation resulting in an elliptically shaped hot spot, which was accurately reconstructed by the model using synthetic x-ray images. This technique was applied to experimental data from implosions in polar-direct-drive illumination geometry with a deliberate laser-drive asymmetry, and the hot-spot emission was reconstructed using spherical-harmonic modes of up to ℓ = 3. A 10% stronger drive on the equator relative to that on the poles resulted in a prolate-shaped hot spot at stagnation with a large negative A2,0 coefficient of A2,0 = -0.47 ± 0.03, directly connecting the modal contribution of the hot-spot shape with the modal contribution in laser-drive asymmetry.

Diagnosis of the imploding shell asymmetry in polar-direct-drive deuterium-tritium cryogenic target implosions on OMEGA.

We discuss the analyses of gated, x-ray imaging data from polar-direct-drive experiments with cryogenically layered deuterium-tritium targets on the OMEGA laser. The in-flight shell asymmetries were diagnosed at various times during the implosion, which was caused by the beam pointing geometry and preimposed variations in the energy partition between the different groups of laser beams. The shape of the ablation surface during the acceleration phase of the implosion was measured along two different lines of sight, and a Legendre mode (ℓ-mode) decomposition was applied for modes of up to ten to investigate shell asymmetries. A clear causal relationship between the imposed beam imbalance and the shape of the in-flight shell asymmetries was observed. The imploded shell with a balanced energy ratio shows smaller values of the amplitudes of ℓ-mode 2 compared to that from implosions with an imbalanced ring energy ratio. The amplitudes of ℓ-modes 4 and 6 are the same within the measurement uncertainty with respect to the change in beam energy ratio.

Microbiome-based interventions to modulate gut ecology and the immune system.

The gut microbiome lies at the intersection between the environment and the host, with the ability to modify host responses to disease-relevant exposures and stimuli. This is evident in how enteric microbes interact with the immune system, e.g., supporting immune maturation in early life, affecting drug efficacy via modulation of immune responses, or influencing development of immune cell populations and their mediators. Many factors modulate gut ecosystem dynamics during daily life and we are just beginning to realise the therapeutic and prophylactic potential of microbiome-based interventions. These approaches vary in application, goal, and mechanisms of action. Some modify the entire community, such as nutritional approaches or faecal microbiota transplantation, while others, such as phage therapy, probiotics, and prebiotics, target specific taxa or strains. In this review, we assessed the experimental evidence for microbiome-based interventions, with a particular focus on their clinical relevance, ecological effects, and modulation of the immune system.

A taxonomic note on the genus Prevotella: Description of four novel genera and emended description of the genera Hallella and Xylanibacter.

The genus Prevotella comprises 55 species with validly published, and correct, names (at June 2021) that are phenotypically, ecologically and functionally diverse. This study used a range of comparative genome approaches (marker gene-based genome phylogeny, core genome phylogeny, average amino acid identity, percentage of conserved proteins and clade-specific marker genes) to identify large differences between the 53 species for which genomes are available, as well as two effectively published yet not validly named species and four novel species. These differences were consistent between the various analysis methods and justify the separation of Prevotella into multiple genera. While the distribution across 19 ecosystem types was unique for each species and inconsistent within clades, the functional repertoire based on the presence/absence of both PFAMs and CAZy families revealed distinct clustering based on the proposed genera. Based on the integration of all results, we propose the reclassification of species previously assigned to the genus Prevotella into seven genera, including four novel genera for which the names Segatella, Hoylesella, Leyella and Palleniella are proposed. In addition to the reclassification of Prevotella, this work describes four novel species, Hallella faecis, Xylanibacter rodentium, Xylanibacter muris, and Palleniella intestinalis.

Bound on hot-spot mix in high-velocity, high-adiabat direct-drive cryogenic implosions based on comparison of absolute x-ray and neutron yields.

In laser-driven implosions for laboratory fusion, the comparison of hot-spot x-ray yield to neutron production can serve to infer hot-spot mix. For high-performance direct-drive implosions, this ratio depends sensitively on the degree of equilibration between the ion and electron fluids. A scaling for x-ray yield as a function of neutron yield and characteristic ion and electron hot-spot temperatures is developed on the basis of simulations with varying degrees of equilibration. We apply this model to hot-spot x-ray measurements of direct-drive cryogenic implosions typical of the direct-drive designs with best ignition metrics. The comparison of the measured x-ray and neutron yields indicates that hot-spot mix, if present, is below a sensitivity estimated as ∼2% by-atom mix of ablator plastic into the hot spot.

Documenting the short-tailed albatross (Phoebastria albatrus) clades historically present in British Columbia, Canada, through ancient DNA analysis of archaeological specimens.

The short-tailed albatross (Phoebastria albatrus) is a threatened seabird whose present-day range encompasses much of the North Pacific. Within this species, there are two genetic clades (Clades 1 and 2) that have distinctive morphologies and foraging ecologies. Due to a global population collapse in the late 19th and early 20th centuries, the frequency of these clades among the short-tailed albatross population that historically foraged off British Columbia, Canada, is unclear. To document the species' historical genetic structure in British Columbia, we applied ancient DNA (aDNA) analysis to 51 archaeological short-tailed albatross specimens from the Yuquot site (Borden site number: DjSp-1) that span the past four millennia. We obtained a 141 bp cytochrome b sequence from 43 of the 51 (84.3%) analyzed specimens. Analyses of these sequences indicate 40 of the specimens belong to Clade 1, while 2 belong to Clade 2. We also identified a single specimen with a novel cytochrome b haplotype. Our results indicate that during the past four millennia most of the short-tailed albatrosses foraging near Yuquot belonged to Clade 1, while individuals from other lineages made more limited use of the area. Comparisons with the results of previous aDNA analyses of archaeological albatrosses from Japanese sites suggest the distribution of Clades 1 and 2 differed. While both albatross clades foraged extensively in the Northwest Pacific, Clade 1 albatrosses appear to have foraged along the west coast of Vancouver Island to a greater extent. Due to their differing distributions, these clades may be exposed to different threats.

Four millennia of long-term individual foraging site fidelity in a highly migratory marine predator.

Theory and field studies suggest that long-term individual foraging site fidelity (IFSF) may be an important adaptation to competition from increasing population. However, the driving mechanisms and extent of long-term IFSF in wild populations of long-lived, migratory animals has been logistically difficult to study, with only a few confirmed instances. Temporal isotopic datasets can reveal long-term patterns in geographical foraging behaviour. We investigate the isotopic compositions of endangered short-tailed albatross (Phoebastria albatrus) over four millennia leading up to their near-extinction. Although not exhibited by short-tailed albatross today, we show past sub-populations displayed a high-degree of long-term IFSF, focusing on the same locations for hundreds of generations. This is the first large-scale evidence for the deep antiquity of long-term IFSF and suggests that it's density-driven. Globally, as populations of species like short-tailed albatross continue to recover from overexploitation, potential for resurgence of geographic specialization may increase exposure to localized hazards, requiring closer conservation monitoring.

Anaerobic single-cell dispensing facilitates the cultivation of human gut bacteria.

Cultivation via classical agar plate (CAP) approaches is widely used to study microbial communities, but they are time-consuming. An alternative approach is the application of single-cell dispensing (SCD), which allows high-throughput, label-free sorting of microscopic particles. We aimed to develop a new anaerobic SCD workflow to cultivate human gut bacteria and compared it with CAP using faecal communities on three rich culture media. We found that the SCD approach significantly decreased the experimental time to obtain pure cultures from 17 ± 4 to 5 ± 0 days, while the isolate diversity and relative abundance coverage were comparable for both approaches. We further tested the total captured fraction by sequencing the sorted bacteria directly after growth as bulk biomass from 2400 dispensed single cells without downstream identification of individual strains. In this approach, the cultured fraction increased from 35.2% to 52.2% for SCD, highlighting the potential for deeper cultivation projects from single samples. SCD-based cultivation also captured species not detected by sequencing (16 ± 5 per sample, including seven novel taxa). From this work, 82 human gut bacterial species across five phyla (Actinobacteriota, Bacteroidota, Desulfobacterota, Firmicutes and Proteobacteria) and 24 families were obtained, including the first cultured member of 11 novel genera and 10 novel species that were fully characterized taxonomically.

Naturalizing laboratory mice by housing in a farmyard-type habitat confers protection against colorectal carcinogenesis.

Living in a farm environment in proximity to animals is associated with reduced risk of developing allergies and asthma, and has been suggested to protect against other diseases, such as inflammatory bowel disease and cancer. Despite epidemiological evidence, experimental disease models that recapitulate such environments are needed to understand the underlying mechanisms. In this study, we show that feralizing conventional inbred mice by continuous exposure to a livestock farmyard-type environment conferred protection toward colorectal carcinogenesis. Two independent experimental approaches for colorectal cancer induction were used; spontaneous (Apc Min/+ mice on an A/J background) or chemical (AOM/DSS). In contrast to conventionally reared laboratory mice, the feralized mouse gut microbiota structure remained stable and resistant to mutagen- and colitis-induced neoplasia. Moreover, the feralized mice exhibited signs of a more mature immunophenotype, indicated by increased expression of NK and T-cell maturation markers, and a more potent IFN-γ response to stimuli. In our study, hygienically born and raised mice subsequently feralized post-weaning were protected to a similar level as life-long exposed mice, although the greatest effect was seen upon neonatal exposure. Collectively, we show protective implications of a farmyard-type environment on colorectal cancer development and demonstrate the utility of a novel animal modeling approach that recapitulates realistic disease responses in a naturalized mammal.

Indigenous sex-selective salmon harvesting demonstrates pre-contact marine resource management in Burrard Inlet, British Columbia, Canada.

To gain insight into pre-contact Coast Salish fishing practices, we used new palaeogenetic analytical techniques to assign sex identifications to salmonid bones from four archaeological sites in Burrard Inlet (Tsleil-Waut), British Columbia, Canada, dating between about 2300-1000 BP (ca. 400 BCE-CE 1200). Our results indicate that male chum salmon (Oncorhynchus keta) were preferentially targeted at two of the four sampled archaeological sites. Because a single male salmon can mate with several females, selectively harvesting male salmon can increase a fishery's maximum sustainable harvest. We suggest such selective harvesting of visually distinctive male spawning chum salmon was a common practice, most effectively undertaken at wooden weirs spanning small salmon rivers and streams. We argue that this selective harvesting of males is indicative of an ancient and probably geographically widespread practice for ensuring sustainable salmon populations. The archaeological data presented here confirms earlier ethnographic accounts describing the selective harvest of male salmon.

Upregulation of Anti-Oxidative Stress Response Improves Metabolic Changes in L-Selectin-Deficient Mice but Does Not Prevent NAFLD Progression or Fecal Microbiota Shifts.

(1) Background: Non-alcoholic fatty liver disease (NAFLD) is a growing global health problem. NAFLD progression involves a complex interplay of imbalanced inflammatory cell populations and inflammatory signals such as reactive oxygen species and cytokines. These signals can derive from the liver itself but also from adipose tissue or be mediated via changes in the gut microbiome. We analyzed the effects of a simultaneous migration blockade caused by L-selectin-deficiency and an enhancement of the anti-oxidative stress response triggered by hepatocytic Kelch-like ECH-associated protein 1 (Keap1) deletion on NAFLD progression. (2) Methods: L-selectin-deficient mice (Lsel-/-Keap1flx/flx) and littermates with selective hepatic Keap1 deletion (Lsel-/-Keap1Δhepa) were compared in a 24-week Western-style diet (WD) model. (3) Results: Lsel-/-Keap1Δhepa mice exhibited increased expression of erythroid 2-related factor 2 (Nrf2) target genes in the liver, decreased body weight, reduced epidydimal white adipose tissue with decreased immune cell frequencies, and improved glucose response when compared to their Lsel-/-Keap1flx/flx littermates. Although WD feeding caused drastic changes in fecal microbiota profiles with decreased microbial diversity, no genotype-dependent shifts were observed. (4) Conclusions: Upregulation of the anti-oxidative stress response improves metabolic changes in L-selectin-deficient mice but does not prevent NAFLD progression and shifts in the gut microbiota.

MiMiC: a bioinformatic approach for generation of synthetic communities from metagenomes.

Environmental and host-associated microbial communities are complex ecosystems, of which many members are still unknown. Hence, it is challenging to study community dynamics and important to create model systems of reduced complexity that mimic major community functions. Therefore, we developed MiMiC, a computational approach for data-driven design of simplified communities from shotgun metagenomes. We first built a comprehensive database of species-level bacterial and archaeal genomes (n = 22 627) consisting of binary (presence/absence) vectors of protein families (Pfam = 17 929). MiMiC predicts the composition of minimal consortia using an iterative scoring system based on maximal match-to-mismatch ratios between this database and the Pfam binary vector of any input metagenome. Pfam vectorization retained enough resolution to distinguish metagenomic profiles between six environmental and host-derived microbial communities (n = 937). The calculated number of species per minimal community ranged between 5 and 11, with MiMiC selected communities better recapitulating the functional repertoire of the original samples than randomly selected species. The inferred minimal communities retained habitat-specific features and were substantially different from communities consisting of most abundant members. The use of a mixture of known microbes revealed the ability to select 23 of 25 target species from the entire genome database. MiMiC is open source and available at https://github.com/ClavelLab/MiMiC.

Early-Life Immune System Maturation in Chickens Using a Synthetic Community of Cultured Gut Bacteria.

The gut microbiome is crucial for both maturation of the immune system and colonization resistance against enteric pathogens. Although chicken are important domesticated animals, the impact of their gut microbiome on the immune system is understudied. Therefore, we investigated the effect of microbiome-based interventions on host mucosal immune responses. Increased levels of IgA and IgY were observed in chickens exposed to maternal feces after hatching compared with strict hygienic conditions. This was accompanied by increased gut bacterial diversity as assessed by 16S rRNA gene amplicon sequencing. Cultivation work allowed the establishment of a collection of 43 bacterial species spanning 4 phyla and 19 families, including the first cultured members of 3 novel genera and 4 novel species that were taxonomically described. This resource is available at www.dsmz.de/chibac A synthetic community consisting of nine phylogenetically diverse and dominant species from this collection was designed and found to be moderately efficient in boosting immunoglobulin levels when provided to chickens early in life.IMPORTANCE The immune system plays a crucial role in sustaining animal health. Its development is markedly influenced by early microbial colonization of the gastrointestinal tract. As chicken are fully dependent on environmental microbes after hatching, extensive hygienic measures in production facilities are detrimental to the microbiota, resulting in low colonization resistance against pathogens. To combat enteric infections, antibiotics are frequently used, which aggravates the issue by altering gut microbiota colonization. Intervention strategies based on cultured gut bacteria are proposed to influence immune responses in chicken.

Recent advances in culture-based gut microbiome research.

Gut microbes affect the physiology of their hosts. Studying their diversity and functions is thus of utmost importance as it will open new avenues towards the discovery of new biomolecules and the treatment of diseases. Gut microbiome research is currently boosted by the unification of metagenomics, which has dominated the field in the last two decades, and cultivation, which is experiencing a renaissance. Each of these approaches has advantages and drawbacks that can be overcome if used synergistically. In this brief article, we summarize recent literature and own studies on the cultivation of gut microbes, provide a succinct status quo of cultured fractions and collections of isolates, and give short opinions on challenges and next steps to take.

Handling of spurious sequences affects the outcome of high-throughput 16S rRNA gene amplicon profiling.

16S rRNA gene amplicon sequencing is a popular approach for studying microbiomes. However, some basic concepts have still not been investigated comprehensively. We studied the occurrence of spurious sequences using defined microbial communities based on data either from the literature or generated in three sequencing facilities and analyzed via both operational taxonomic units (OTUs) and amplicon sequence variants (ASVs) approaches. OTU clustering and singleton removal, a commonly used approach, delivered approximately 50% (mock communities) to 80% (gnotobiotic mice) spurious taxa. The fraction of spurious taxa was generally lower based on ASV analysis, but varied depending on the gene region targeted and the barcoding system used. A relative abundance of 0.25% was found as an effective threshold below which the analysis of spurious taxa can be prevented to a large extent in both OTU- and ASV-based analysis approaches. Using this cutoff improved the reproducibility of analysis, i.e., variation in richness estimates was reduced by 38% compared with singleton filtering using six human fecal samples across seven sequencing runs. Beta-diversity analysis of human fecal communities was markedly affected by both the filtering strategy and the type of phylogenetic distances used for comparison, highlighting the importance of carefully analyzing data before drawing conclusions on microbiome changes. In summary, handling of artifact sequences during bioinformatic processing of 16S rRNA gene amplicon data requires careful attention to avoid the generation of misleading findings. We propose the concept of effective richness to facilitate the comparison of alpha-diversity across studies.