Complex subsurface hydrothermal fluid mixing at a submarine arc volcano supports distinct and highly diverse microbial communities.

Hydrothermally active submarine volcanoes are mineral-rich biological oases contributing significantly to chemical fluxes in the deep sea, yet little is known about the microbial communities inhabiting these systems. Here we investigate the diversity of microbial life in hydrothermal deposits and their metagenomics-inferred physiology in light of the geological history and resulting hydrothermal fluid paths in the subsurface of Brothers submarine volcano north of New Zealand on the southern Kermadec arc. From metagenome-assembled genomes we identified over 90 putative bacterial and archaeal genomic families and nearly 300 previously unknown genera, many potentially endemic to this submarine volcanic environment. While magmatically influenced hydrothermal systems on the volcanic resurgent cones of Brothers volcano harbor communities of thermoacidophiles and diverse members of the superphylum "DPANN," two distinct communities are associated with the caldera wall, likely shaped by two different types of hydrothermal circulation. The communities whose phylogenetic diversity primarily aligns with that of the cone sites and magmatically influenced hydrothermal systems elsewhere are characterized predominately by anaerobic metabolisms. These populations are probably maintained by fluids with greater magmatic inputs that have interacted with different (deeper) previously altered mineral assemblages. However, proximal (a few meters distant) communities with gene-inferred aerobic, microaerophilic, and anaerobic metabolisms are likely supported by shallower seawater-dominated circulation. Furthermore, mixing of fluids from these two distinct hydrothermal circulation systems may have an underlying imprint on the high microbial phylogenomic diversity. Collectively our results highlight the importance of considering geologic evolution and history of subsurface processes in studying microbial colonization and community dynamics in volcanic environments.

Validation of the Global Limb Anatomic Staging System in first-time lower extremity revascularization.

OBJECTIVE: The Global Limb Anatomic Staging System (GLASS) was developed as a new anatomic classification scheme to grade the severity of chronic limb threatening ischemia. We evaluated the ability of this anatomic grading system to determine major adverse limb events after lower extremity revascularization. METHODS: We performed a single-institutional retrospective review of 1060 consecutive patients who had undergone 1180 first-time open or endovascular revascularization procedures for chronic limb threatening ischemia from 2005 to 2014. Using the review of angiographic images, the limbs were classified as GLASS stage 1, 2, or 3. The primary composite outcome was reintervention, major amputation (below- or above-the-knee amputation), and/or restenosis (>3.5× step-up by duplex criteria) events (RAS). The secondary outcomes included all-cause mortality, failure to cross the lesion by endovascular methods, and a comparison between bypass vs endovascular intervention. Kaplan-Meier estimates were used to determine the event rates at 1 and 5 years, and Cox regression analysis was used to adjust for baseline differences among the GLASS stages. RESULTS: Of all patients undergoing first-time revascularization, imaging studies were available for 1180 procedures (91%) for GLASS grading. Of these procedures, 552 were open bypass (47%) and 628 were endovascular intervention (53%). Compared with GLASS stage 1 disease (n = 267, 23%), stage 2 (n = 367; 31%) and stage 3 (n = 546; 42%) disease were associated with a greater risk of RAS at 1 year (stage 1, 33% vs stage 2, 48% vs stage 3, 53%) and 5 years (stage 1, 45% [reference]; stage 2, 65%; hazard ratio [HR], 1.7; 95% confidence interval [CI], 1.3-2.2; P < .001; stage 3, 69%; HR, 2.3; 95% CI, 1.7-2.9; P < .001). These differences were mainly driven by reintervention and restenosis rather than by major amputation. The 5-year mortality was similar for stage 2 and 3 compared with stage 1 disease (stage 1, 40% [reference]; stage 2, 45%; HR, 1.1; 95% CI, 0.8-1.4; P = .69; stage 3, 49%; HR, 1.2; 95% CI, 1.0-1.6; P = .11). For all attempted endovascular interventions, failure to cross a target lesion increased with advancing GLASS stage (stage 1, 4.5% vs stage 2, 6.3% vs stage 3, 13.3%; P < .01). Compared with open bypass (n = 552; 46.8%), endovascular intervention (n = 628; 53.3%) was associated with a higher rate of 5-year RAS for GLASS stage 1 (49% vs 34%; HR, 1.9; 95% CI, [1.1-3.5; P = .03), stage 2 (69% vs 52%; HR, 1.7; 95% CI, 1.2-2.5; P < .01), and stage 3 (83% vs 61%; HR, 1.5; 95% CI, 1.2-2.0; P < .01) disease. CONCLUSIONS: For patients undergoing first-time lower extremity revascularization, the GLASS can be used to predict for reintervention and restenosis. Bypass resulted in better long-term outcomes compared with endovascular intervention for all GLASS stages.

Wound location is independently associated with adverse outcomes following first-time revascularization for tissue loss.

OBJECTIVE: Few studies adequately evaluate the impact of wound location on patient outcomes after lower extremity revascularization. Consequently, we evaluated the relationship between lower extremity wound location and long-term outcomes. METHODS: We reviewed all patients at our institution undergoing any first-time open surgical bypass or percutaneous transluminal angioplasty with or without stenting for tissue loss between 2005 and 2014. We categorized wounds into three distinct groups: forefoot (ie, toes and metatarsal heads), midfoot (ie, dorsal, plantar, lateral, medial surfaces excluding toes, metatarsal heads, or heel), and heel. Limbs with multiple wounds were excluded from analyses. We compared rates of perioperative complications, wound healing, reintervention, limb salvage, amputation-free survival, and survival using χ2, Kaplan-Meier, and Cox regression analyses. RESULTS: Of 2869 infrainguinal revascularizations from 2005 to 2014, 1126 underwent a first-time revascularization for tissue loss, of which 253 patients had multiple wounds, 197 had wounds proximal to the ankle, 100 had unreliable wound information, and 576 (forefoot, n = 397; midfoot, n = 61; heel, n = 118) fit our criteria and had a single foot wound with reliable information regarding wound specifics. Patients with forefoot, midfoot, and heel wounds had similar rates of coronary artery disease, hypertension, diabetes, and smoking history (all P > .05). Conversely, there were significant differences in patient age (71 vs 69 vs 70 years), prevalence of gangrene (41% vs 5% vs 21%), and dialysis dependence (18% vs 17% vs 30%) (all P < .05). There were no statistically significant differences in perioperative mortality (1.3% vs 4.9% vs 4.2%; P = .06) or postoperative complications among the three groups. Between forefoot, midfoot, and heel wounds, there were significant differences in unadjusted 6-month rates of complete wound healing (69% vs 64% vs 53%), 3-year rates of amputation-free survival (54% vs 57% vs 35%), and survival (61% vs 72% vs 41%) (all P < .05). After adjustment, compared with forefoot wounds, heel wounds were associated with higher rates of incomplete 6-month wound healing (hazard ratio [HR], 1.6; 95% confidence interval [CI], 1.1-2.]), major amputation or mortality (HR, 1.7; 95% CI, 1.1-2.7), and all-cause mortality (HR, 1.8; 95% CI, 1.1-3.0), but not major amputation alone (HR, 2.1; 95% CI, 0.9-4.5). In open surgical bypass-first patients, heel wounds were solely associated with an increased risk of all-cause mortality (HR, 1.7; 95% CI, 1.1-2.8), whereas heel wounds in percutaneous transluminal angioplasty-first patients were associated with an increased risk of incomplete wound healing (HR, 2.2; 95% CI, 1.3-3.7), major amputation or mortality (HR, 2.3; 95% CI, 1.1-5.4), and all-cause mortality (HR, 2.8; 95% CI, 1.1-7.2). CONCLUSIONS: Heel wounds confer considerably higher short- and long-term morbidity and mortality compared with midfoot or forefoot wounds in patients undergoing any first-time lower extremity revascularization.

Genomic comparison of deep-sea hydrothermal genera related to Aeropyrum, Thermodiscus and Caldisphaera, and proposed emended description of the family Acidilobaceae.

Deep-sea hydrothermal vents host archaeal and bacterial thermophilic communities, including taxonomically and functionally diverse Thermoproteota. Despite their prevalence in high-temperature submarine communities, Thermoproteota are chronically under-represented in genomic databases and issues have emerged regarding their nomenclature, particularly within the Aeropyrum-Thermodiscus-Caldisphaera. To resolve some of these problems, we identified 47 metagenome-assembled genomes (MAGs) within this clade, from 20 previously published deep-sea hydrothermal vent and submarine volcano metagenomes, and 24 MAGs from public databases. Using phylogenomic analysis, Genome Taxonomy Database Toolkit (GTDB-Tk) taxonomic assessment, 16S rRNA gene phylogeny, average amino acid identity (AAI) and functional gene patterns, we re-evaluated of the taxonomy of the Aeropyrum-Thermodiscus-Caldisphaera. At least nine genus-level clades were identified with two or more MAGs. In accordance with SeqCode requirements and recommendations, we propose names for three novel genera, viz. Tiamatella incendiivivens, Hestiella acidicharens and Calypsonella navitae. A fourth genus was also identified related to Thermodiscus maritimus, for which no available sequenced genome exists. We propose the novel species Thermodiscus eudorianus to describe our high-quality Thermodiscus MAG, which represents the type genome for the genus. All three novel genera and T. eudorianus are likely anaerobic heterotrophs, capable of fermenting protein-rich carbon sources, while some Tiamatella, Calypsonella and T. eudorianus may also reduce polysulfides, thiosulfate, sulfur and/or selenite, and the likely acidophile, Hestiella, may reduce nitrate and/or perchlorate. Based on phylogenomic evidence, we also propose the family Acidilobaceae be amended to include Caldisphaera, Aeropyrum, Thermodiscus and Stetteria and the novel genera described here.

A Formative Evaluation of an Online Meal Kit and Grocery Platform for Supplemental Nutrition Assistance Program Recipients.

OBJECTIVE: To assess the barriers and facilitators to online food purchasing through a meal kit and grocery shopping website titled NY SNAP Express among Supplemental Nutrition Assistance Program (SNAP) beneficiaries. METHODS: A purposive sample of SNAP-eligible adults residing in New York State participated in interviews guided by the Capabilities, Opportunities, Motivations, and Behaviors Model. RESULTS: Barriers to online food purchasing among participants (n = 32) include physiological and health conditions, the weight of food, technology, language, the price of foods, transportation challenges, the stigma associated with SNAP, and concerns regarding the quantity and quality of meal kits. Facilitators include health and nutrition improvements, knowledge and skills, saving money, culturally relevant meals, increased efficiency in food purchases and preparation, and customization. CONCLUSIONS AND IMPLICATIONS: Online platforms such as NY SNAP Express have the potential to increase nutritious food access and resources among SNAP recipients; however, improvements are necessary to meet the needs of its audience.

Sleep slow-wave oscillations trigger seizures in a genetic epilepsy model of Dravet syndrome.

Sleep is the preferential period when epileptic spike-wave discharges appear in human epileptic patients, including genetic epileptic seizures such as Dravet syndrome with multiple mutations including SCN1A mutation and GABAA receptor γ2 subunit Gabrg2Q390X mutation in patients, which presents more severe epileptic symptoms in female patients than male patients. However, the seizure onset mechanism during sleep still remains unknown. Our previous work has shown that the sleep-like state-dependent homeostatic synaptic potentiation can trigger epileptic spike-wave discharges in one transgenic heterozygous Gabrg2+/Q390X knock-in mouse model.1 Here, using this heterozygous knock-in mouse model, we hypothesized that slow-wave oscillations themselves in vivo could trigger epileptic seizures. We found that epileptic spike-wave discharges in heterozygous Gabrg2+/Q390X knock-in mice exhibited preferential incidence during non-rapid eye movement sleep period, accompanied by motor immobility/facial myoclonus/vibrissal twitching and more frequent spike-wave discharge incidence appeared in female heterozygous knock-in mice than male heterozygous knock-in mice. Optogenetically induced slow-wave oscillations in vivo significantly increased epileptic spike-wave discharge incidence in heterozygous Gabrg2+/Q390X knock-in mice with longer duration of non-rapid eye movement sleep or quiet-wakeful states. Furthermore, suppression of slow-wave oscillation-related homeostatic synaptic potentiation by 4-(diethylamino)-benzaldehyde injection (i.p.) greatly attenuated spike-wave discharge incidence in heterozygous knock-in mice, suggesting that slow-wave oscillations in vivo did trigger seizure activity in heterozygous knock-in mice. Meanwhile, sleep spindle generation in wild-type littermates and heterozygous Gabrg2+/Q390X knock-in mice involved the slow-wave oscillation-related homeostatic synaptic potentiation that also contributed to epileptic spike-wave discharge generation in heterozygous Gabrg2+/Q390X knock-in mice. In addition, EEG spectral power of delta frequency (0.1-4 Hz) during non-rapid eye movement sleep was significantly larger in female heterozygous Gabrg2+/Q390X knock-in mice than that in male heterozygous Gabrg2+/Q390X knock-in mice, which likely contributes to the gender difference in seizure incidence during non-rapid eye movement sleep/quiet-wake states of human patients. Overall, all these results indicate that slow-wave oscillations in vivo trigger the seizure onset in heterozygous Gabrg2+/Q390X knock-in mice, preferentially during non-rapid eye movement sleep period and likely generate the sex difference in seizure incidence between male and female heterozygous Gabrg2+/Q390X knock-in mice.

Global patterns of diversity and metabolism of microbial communities in deep-sea hydrothermal vent deposits.

BACKGROUND: When deep-sea hydrothermal fluids mix with cold oxygenated fluids, minerals precipitate out of solution and form hydrothermal deposits. These actively venting deep-sea hydrothermal deposits support a rich diversity of thermophilic microorganisms which are involved in a range of carbon, sulfur, nitrogen, and hydrogen metabolisms. Global patterns of thermophilic microbial diversity in deep-sea hydrothermal ecosystems have illustrated the strong connectivity between geological processes and microbial colonization, but little is known about the genomic diversity and physiological potential of these novel taxa. Here we explore this genomic diversity in 42 metagenomes from four deep-sea hydrothermal vent fields and a deep-sea volcano collected from 2004 to 2018 and document their potential implications in biogeochemical cycles. RESULTS: Our dataset represents 3635 metagenome-assembled genomes encompassing 511 novel and recently identified genera from deep-sea hydrothermal settings. Some of the novel bacterial (107) and archaeal genera (30) that were recently reported from the deep-sea Brothers volcano were also detected at the deep-sea hydrothermal vent fields, while 99 bacterial and 54 archaeal genera were endemic to the deep-sea Brothers volcano deposits. We report some of the first examples of medium- (≥ 50% complete, ≤ 10% contaminated) to high-quality (> 90% complete, < 5% contaminated) MAGs from phyla and families never previously identified, or poorly sampled, from deep-sea hydrothermal environments. We greatly expand the novel diversity of Thermoproteia, Patescibacteria (Candidate Phyla Radiation, CPR), and Chloroflexota found at deep-sea hydrothermal vents and identify a small sampling of two potentially novel phyla, designated JALSQH01 and JALWCF01. Metabolic pathway analysis of metagenomes provides insights into the prevalent carbon, nitrogen, sulfur, and hydrogen metabolic processes across all sites and illustrates sulfur and nitrogen metabolic "handoffs" in community interactions. We confirm that Campylobacteria and Gammaproteobacteria occupy similar ecological guilds but their prevalence in a particular site is driven by shifts in the geochemical environment. CONCLUSION: Our study of globally distributed hydrothermal vent deposits provides a significant expansion of microbial genomic diversity associated with hydrothermal vent deposits and highlights the metabolic adaptation of taxonomic guilds. Collectively, our results illustrate the importance of comparative biodiversity studies in establishing patterns of shared phylogenetic diversity and physiological ecology, while providing many targets for enrichment and cultivation of novel and endemic taxa. Video Abstract.

Artificial sleep-like up/down-states induce synaptic plasticity in cortical neurons from mouse brain slices.

During non-rapid eye movement (NREM) sleep, cortical neuron activity alternates between a depolarized (firing, up-state) and a hyperpolarized state (down-state) coinciding with delta electroencephalogram (EEG) slow-wave oscillation (SWO, 0. 5-4 Hz) in vivo. Recently, we have found that artificial sleep-like up/down-states can potentiate synaptic strength in layer V cortical neurons ex vivo. Using mouse coronal brain slices, whole cell voltage-clamp recordings were made from layer V cortical pyramidal neurons to record spontaneous excitatory synaptic currents (sEPSCs) and inhibitory synaptic currents (sIPSCs). Artificial sleep-like up/down-states (as SWOs, 0.5 Hz, 10 min, current clamp mode) were induced by injecting sinusoidal currents into layer V cortical neurons. Baseline pre-SWO recordings were recorded for 5 min and post-SWO recordings for at least 25-30 min. Compared to pre-SWO sEPSCs or sIPSCs, post-SWO sEPSCs or sIPSCs in layer V cortical neurons exhibited significantly larger amplitudes and a higher frequency for 30 min. This finding suggests that both sEPSCs and sIPSCs could be potentiated in layer V cortical neurons by the low-level activity of SWOs, and sEPSCs and sIPSCs maintained a balance in layer V cortical neurons during pre- and post-SWO periods. Overall, this study presents an ex vivo method to show SWO's ability to induce synaptic plasticity in layer V cortical neurons, which may underlie sleep-related synaptic potentiation for sleep-related memory consolidation in vivo.

An essential role for tungsten in the ecology and evolution of a previously uncultivated lineage of anaerobic, thermophilic Archaea.

Trace metals have been an important ingredient for life throughout Earth's history. Here, we describe the genome-guided cultivation of a member of the elusive archaeal lineage Caldarchaeales (syn. Aigarchaeota), Wolframiiraptor gerlachensis, and its growth dependence on tungsten. A metagenome-assembled genome (MAG) of W. gerlachensis encodes putative tungsten membrane transport systems, as well as pathways for anaerobic oxidation of sugars probably mediated by tungsten-dependent ferredoxin oxidoreductases that are expressed during growth. Catalyzed reporter deposition-fluorescence in-situ hybridization (CARD-FISH) and nanoscale secondary ion mass spectrometry (nanoSIMS) show that W. gerlachensis preferentially assimilates xylose. Phylogenetic analyses of 78 high-quality Wolframiiraptoraceae MAGs from terrestrial and marine hydrothermal systems suggest that tungsten-associated enzymes were present in the last common ancestor of extant Wolframiiraptoraceae. Our observations imply a crucial role for tungsten-dependent metabolism in the origin and evolution of this lineage, and hint at a relic metabolic dependence on this trace metal in early anaerobic thermophiles.

Deep-sea hydrothermal vent metagenome-assembled genomes provide insight into the phylum Nanoarchaeota.

Ectosymbiotic Nanoarchaeota live on the surface of diverse archaeal hosts. Despite being broadly distributed in global geothermal systems, only three Nanoarchaeota have been successfully co-cultivated with their hosts, and until now no nanoarchaeotal cultures or genomes have been described from deep-sea hydrothermal vents. We recovered three nanoarchaeotal metagenome-assembled genomes (MAGs) from deep-sea hydrothermal vent sites at the Eastern Lau Spreading Center (M10-121), Guaymas Basin (Gua-46) and the Mid-Cayman Rise (MC-1). Based on average amino acid identity analysis, M10-121 is a novel species in the candidate genus Nanoclepta, while the other two MAGs represent novel genera in the Nanoarchaeota. Like previously sequenced Nanoarchaeota, each MAG encodes at least one split protein-coding gene. The MAGs also contain a mosaic of key nanoarchaeotal features, including CRISPR repeat regions and marker genes for gluconeogenesis and archaeal flagella. MC-1 also encodes the pentose bisphosphate pathway, which may allow the nanoarchaeote to bypass several steps in glycolysis and produce ATP.

A new symbiotic nanoarchaeote (Candidatus Nanoclepta minutus) and its host (Zestosphaera tikiterensis gen. nov., sp. nov.) from a New Zealand hot spring.

Three thermophilic Nanoarchaeota-Crenarchaeota symbiotic systems have been described. We obtained another stable anaerobic enrichment culture at 80°C, pH 6.0 from a New Zealand hot spring. The nanoarchaeote (Ncl-1) and its host (NZ3T) were isolated in co-culture and their genomes assembled. The small (∼200nm) flagellated cocci were often attached to larger cocci. Based on 16S rRNA gene similarity (88.4%) and average amino acid identity (52%), Ncl-1 is closely related to Candidatus Nanopusillus acidilobi. Their genomes both encode for archaeal flagella and partial glycolysis and gluconeogenesis pathways, but lack ATP synthase genes. Like Nanoarchaeum equitans, Ncl-1 has a CRISPR-Cas system. Ncl-1 also relies on its crenarchaeotal host for most of its biosynthetic needs. The host NZ3T was isolated and grows on proteinaceous substrates but not on sugars, alcohols, or fatty acids. NZ3T requires thiosulfate and grows best at 82°C, pH 6.0. NZ3T is most closely related to the Desulfurococcaceae, Ignisphaera aggregans (∼92% 16S rRNA gene sequence similarity, 45% AAI). Based on phylogenetic, physiological and genomic data, Ncl-1 and NZ3T represent novel genera in the Nanoarchaeota and the Desulfurococcaceae, respectively, with the proposed names Candidatus Nanoclepta minutus and Zestosphaera tikiterensis gen. nov., sp. nov., type strain NZ3T (=DSMZ 107634T=OCM 1213T).

Draft Genome Sequence of a Dictyoglomus sp. from an Enrichment Culture of a New Zealand Geothermal Spring.

A draft genome of a novel Dictyoglomus sp., NZ13-RE01, was obtained from a New Zealand hot spring enrichment culture. The 1,927,012-bp genome is similar in both size and G+C content to other Dictyoglomus spp. Like its relatives, Dictyoglomus sp. NZ13-RE01 encodes many genes involved in complex carbohydrate metabolism.