Single-Cell Phenotyping of Extracellular Electron Transfer via Microdroplet Encapsulation.

Electroactive organisms contribute to metal cycling, pollutant removal, and other redox-driven environmental processes. Studying this phenomenon in high-throughput is challenging since extracellular reduction cannot easily be traced back to its cell of origin within a mixed population. Here, we describe the development of a microdroplet emulsion system to enrich EET-capable organisms. We validated our system using the model electroactive organism S. oneidensis and describe the tooling of a benchtop microfluidic system for oxygen-limited processes. We demonstrated enrichment of EET-capable phenotypes from a mixed wild-type and EET-knockout population. As a proof-of-concept application, bacteria were collected from iron sedimentation from Town Lake (Austin, TX) and subjected to microdroplet enrichment. We observed an increase in EET-capable organisms in the sorted population that was distinct when compared to a population enriched in a bulk culture more closely akin to traditional techniques for discovering EET-capable bacteria. Finally, two bacterial species, C. sakazakii and V. fessus not previously shown to be electroactive, were further cultured and characterized for their ability to reduce channel conductance in an organic electrochemical transistor (OECT) and to reduce soluble Fe(III). We characterized two bacterial species not previously shown to exhibit electrogenic behavior. Our results demonstrate the utility of a microdroplet emulsions for identifying putative EET-capable bacteria and how this technology can be leveraged in tandem with existing methods.

Effect of Sample Geometry on Graphitization of Polyacrylonitrile.

In this study, it is analyzed how sample geometry (spheres, nanofibers, or films) influences the graphitization behavior of polyacrylonitrile (PAN) molecules. The chemical bonding and changes in the composition of these three geometries are studied at the oxidation, carbonization, and graphitization stages via scanning electron microscopy (SEM), in situ thermogravimetric-infrared (TGA-IR) analysis, elemental analysis, Raman spectroscopy, and X-ray photoelectron spectroscopy (XPS). The influence of molecular alignment on the graphitization of the three sample geometries is investigated using synchrotron wide-angle X-ray diffraction (WAXD) and transmission electron microscopy (TEM). The effects of molecular alignment at different draw rates during spinning are explored in detail.

Growth of diamond in liquid metal at 1 atm pressure.

Natural diamonds were (and are) formed (thousands of million years ago) in the upper mantle of Earth in metallic melts at temperatures of 900-1,400 °C and at pressures of 5-6 GPa (refs. 1,2). Diamond is thermodynamically stable under high-pressure and high-temperature conditions as per the phase diagram of carbon3. Scientists at General Electric invented and used a high-pressure and high-temperature apparatus in 1955 to synthesize diamonds by using molten iron sulfide at about 7 GPa and 1,600 °C (refs. 4-6). There is an existing model that diamond can be grown using liquid metals only at both high pressure and high temperature7. Here we describe the growth of diamond crystals and polycrystalline diamond films with no seed particles using liquid metal but at 1 atm pressure and at 1,025 °C, breaking this pattern. Diamond grew in the subsurface of liquid metal composed of gallium, iron, nickel and silicon, by catalytic activation of methane and diffusion of carbon atoms into and within the subsurface regions. We found that the supersaturation of carbon in the liquid metal subsurface leads to the nucleation and growth of diamonds, with Si playing an important part in stabilizing tetravalently bonded carbon clusters that play a part in nucleation. Growth of (metastable) diamond in liquid metal at moderate temperature and 1 atm pressure opens many possibilities for further basic science studies and for the scaling of this type of growth.

Inflammatory Consequences: Hepatitis C Virus-Induced Inflammasome Activation and Pyroptosis.

Hepatitis C virus (HCV), despite the availability of effective direct-acting antivirals (DAAs) that clear the virus from >95% of individuals treated, continues to cause significant health care burden due to disease progression that can lead to fibrosis, cirrhosis, and/or hepatocellular carcinoma. The fact that some people who are treated with DAAs still go on to develop worsening liver disease warrants further study into the immunopathogenesis of HCV. Many viral infections, including HCV, have been associated with activation of the inflammasome/pyroptosis pathway. This inflammatory cell death pathway ultimately results in cell lysis and release of inflammatory cytokines, IL-18 and IL-1ß. This review will report on studies that investigated HCV and inflammasome activation/pyroptosis. This includes clinical in vivo data showing elevated pyroptosis-associated cytokines in the blood of individuals living with HCV, studies of genetic associations of pyroptosis-related genes and development of liver disease, and in vitro studies aimed at understanding the mechanism of pyroptosis induced by HCV. Finally, we discuss major gaps in understanding and outstanding questions that remain in the field of HCV-induced pyroptosis.

Potential routes of plastics biotransformation involving novel plastizymes revealed by global multi-omic analysis of plastic associated microbes.

Despite increasing efforts across various disciplines, the fate, transport, and impact of synthetic plastics on the environment and public health remain poorly understood. To better elucidate the microbial ecology of plastic waste and its potential for biotransformation, we conducted a large-scale analysis of all publicly available meta-omic studies investigating plastics (n = 27) in the environment. Notably, we observed low prevalence of known plastic degraders throughout most environments, except for substantial enrichment in riverine systems. This indicates rivers may be a highly promising environment for discovery of novel plastic bioremediation products. Ocean samples associated with degrading plastics showed clear differentiation from non-degrading polymers, showing enrichment of novel putative biodegrading taxa in the degraded samples. Regarding plastisphere pathogenicity, we observed significant enrichment of antimicrobial resistance genes on plastics but not of virulence factors. Additionally, we report a co-occurrence network analysis of 10 + million proteins associated with the plastisphere. This analysis revealed a localized sub-region enriched with known and putative plastizymes-these may be useful for deeper investigation of nature's ability to biodegrade man-made plastics. Finally, the combined data from our meta-analysis was used to construct a publicly available database, the Plastics Meta-omic Database (PMDB)-accessible at plasticmdb.org. These data should aid in the integrated exploration of the microbial plastisphere and facilitate research efforts investigating the fate and bioremediation potential of environmental plastic waste.

3D microprinting of inorganic porous materials by chemical linking-induced solidification of nanocrystals.

Three-dimensional (3D) microprinting is considered a next-generation manufacturing process for the production of microscale components; however, the narrow range of suitable materials, which include mainly polymers, is a critical issue that limits the application of this process to functional inorganic materials. Herein, we develop a generalised microscale 3D printing method for the production of purely inorganic nanocrystal-based porous materials. Our process is designed to solidify all-inorganic nanocrystals via immediate dispersibility control and surface linking-induced interconnection in the nonsolvent linker bath and thereby creates multibranched gel networks. The process works with various inorganic materials, including metals, semiconductors, magnets, oxides, and multi-materials, not requiring organic binders or stereolithographic equipment. Filaments with a diameter of sub-10 µm are printed into designed complex 3D microarchitectures, which exhibit full nanocrystal functionality and high specific surface areas as well as hierarchical porous structures. This approach provides the platform technology for designing functional inorganics-based porous materials.

Protocol for the electrochemical polymerization of polyhydrocarbons from chlorinated methanes and an analysis of its polymerization reaction pathway.

The mass production of polyhydrocarbons (PHCs) is achieved through electrochemical polymerization and its synthetic pathway is investigated using a combination of electrochemical techniques, NMR, and Fourier transform infrared (FTIR) spectroscopy. Here, we present a protocol for the electrochemical polymerization of PHCs from chlorinated methanes and an analysis of its polymerization reaction pathway. We describe steps for large-scale synthesis and characterization of PHCs and studying electrochemical polymerization reactions using cyclic voltammetry and chronoamperometry techniques along with NMR and FTIR spectroscopy using isotope-labeled reactants. For complete details on the use and execution of this protocol, please refer to Seo et al.,1 Lee et al.,2 and Seo et al.3.

Direct Electrochemical Functionalization of Graphene Grown on Cu Including the Reaction Rate Dependence on the Cu Facet Type.

We present an electrochemical method to functionalize single-crystal graphene grown on copper foils with a (111) surface orientation by chemical vapor deposition (CVD). Graphene on Cu(111) is functionalized with 4-iodoaniline by applying a constant negative potential, and the degree of functionalization depends on the applied potential and reaction time. Our approach stands out from previous methods due to its transfer-free method, which enables more precise and efficient functionalization of single-crystal graphene. We report the suggested effects of the Cu substrate facet by comparing the reactivity of graphene on Cu(111) and Cu(115). The electrochemical reaction rate changes dramatically at the potential threshold for each facet. Kelvin probe force microscopy was used to measure the work function, and the difference in onset potentials of the electrochemical reaction on these two different facets are explained in terms of the difference in work function values. Density functional theory and Monte Carlo calculations were used to calculate the work function of graphene and the thermodynamic stability of the aniline functionalized graphene on these two facets. This study provides a deeper understanding of the electrochemical behavior of graphene (including single-crystal graphene) on Cu(111) and Cu(115). It also serves as a basis for further study of a broad range of reagents and thus functional groups and of the role of metal substrate beneath graphene.

Characteristics of Vaccine- and Infection-Induced Systemic IgA Anti-SARS-CoV-2 Spike Responses.

Mucosal IgA is widely accepted as providing protection against respiratory infections, but stimulation of mucosal immunity, collection of mucosal samples and measurement of mucosal IgA can be problematic. The relationship between mucosal and circulating IgA responses is unclear, however, whole blood is readily collected and circulating antigen-specific IgA easily measured. We measured circulating IgA against SARS-CoV-2 spike (S) to investigate vaccine- and infection-induced production and correlation with protection. Circulating IgA against ancestral (Wuhan-Hu-1) and Omicron (BA.1) S proteins was measured at different time points in a total of 143 subjects with varied backgrounds of vaccination and infection. Intramuscular vaccination induced circulating anti-SARS-CoV-2 S IgA. Subjects with higher levels of vaccine-induced IgA against SARS-CoV-2 S (p = 0.0333) or receptor binding domain (RBD) (p = 0.0266) were less likely to experience an Omicron breakthrough infection. The same associations did not hold for circulating IgG anti-SARS-CoV-2 S levels. Breakthrough infection following two vaccinations generated stronger IgA anti-SARS-CoV-2 S responses (p = 0.0002) than third vaccinations but did not selectively increase circulating IgA against Omicron over ancestral S, indicating immune imprinting of circulating IgA responses. Circulating IgA against SARS-CoV-2 S following breakthrough infection remained higher than vaccine-induced levels for over 150 days. In conclusion, intramuscular mRNA vaccination induces circulating IgA against SARS-CoV-2 S, and higher levels are associated with protection from breakthrough infection. Vaccination with ancestral S enacts imprinting within circulating IgA responses that become apparent after breakthrough infection with Omicron. Breakthrough infection generates stronger and more durable circulating IgA responses against SARS-CoV-2 S than vaccination alone.

Addressing the psychology of weight loss and maintenance: A feasibility study of the Skills for weight loss and Maintenance weight management programme.

OBJECTIVES: Building on prior theory, we aimed to evaluate the feasibility and acceptability of integrating novel, weight loss maintenance strategies into existing weight management programmes. We also piloted recruitment and data collection procedures for future research. DESIGN: Two phases of action research nested within a single-arm feasibility study. The intervention was refined between phases using feedback from intervention fidelity analysis and qualitative exploration of patient and provider experiences. Changes in outcomes were assessed up to 18 months post-baseline. METHODS: One hundred adults with a mean body mass index of 37 kg/m2 were offered the Skills for weight loss and Maintenance (SkiM) intervention. This included existing weight management programme content and additional weight loss maintenance techniques delivered fortnightly for 6 months in local community centres to groups of 11-15 people. RESULTS: Of the 100 participants, 65%, 58% and 56% provided data at 7, 12 and 18 months. Across both phases, the mean initial weight loss was 4.2 kg (95% CI: 2.4-5.9) and 3.1 kg at 18 months (95% CI: .8-5.5). In Phase 2, we observed better weight loss maintenance (.5 kg [13.2%] regain from 7 to 18 months, vs. 1.7 kg [36.2%] in Phase 1). Variation in outcomes, high early dropout rates and qualitative feedback indicated that, although delivery of the intervention and trial procedures was feasible and acceptable, there was scope to refine the intervention to engage a wider range of participants. Intervention fidelity was acceptable, particularly in Phase 2. CONCLUSIONS: The SkiM intervention seems promising, but more research is needed to improve recruitment and retention prior to further evaluation.

Growth Mechanism and Kinetics of Diamond in Liquid Gallium from Quantum Mechanics Molecular Dynamics Simulations.

Ruoff and co-workers recently demonstrated low-temperature (1193 K) homoepitaxial diamond growth from liquid gallium solvent. To develop an atomistic mechanism for diamond growth underlying this remarkable demonstration, we carried out density functional theory-based molecular dynamics (DFT-MD) simulations to examine the mechanism of single-crystal diamond growth on various low-index crystallographic diamond surfaces (100), (110), and (111) in liquid Ga with CH4. We find that carbon linear chains form in liquid Ga and then react with the growing diamond surface, leading first to the formation of carbon rings on the surface and then initiation of diamond growth. Our simulations find faster growth on the (110) surface than on the (100) or (111) surfaces, suggesting the (110) surface as a plausible growth surface in liquid Ga. For (110) surface growth, we predict the optimum growth temperature to be ∼1300 K, arising from a balance between the kinetics of forming carbon chains dissolved in Ga and the stability of carbon rings on the growing surface. We find that the rate-determining step for diamond growth is dehydrogenation of the growing hydrogenated (110) surface of diamond. Inspired by the recent experimental studies by Ruoff and co-workers demonstrating that Si accelerates diamond growth in Ga, we show that addition of Si into liquid Ga significantly increases the rate of dehydrogenating the growing surface. Extrapolating from the DFT-MD predicted rates at 2800 to 3500 K, we predict the growth rate at the experimental growth temperature of 1193 K, leading to rates in reasonable agreement with the experiment. These fundamental mechanisms should provide guidance in optimizing low-temperature diamond growth.

Maintaining soldier musculoskeletal health using personalised digital humans, wearables and/or computer vision.

OBJECTIVES: The physical demands of military service place soldiers at risk of musculoskeletal injuries and are major concerns for military capability. This paper outlines the development new training technologies to prevent and manage these injuries. DESIGN: Narrative review. METHODS: Technologies suitable for integration into next-generation training devices were examined. We considered the capability of technologies to target tissue level mechanics, provide appropriate real-time feedback, and their useability in-the-field. RESULTS: Musculoskeletal tissues' health depends on their functional mechanical environment experienced in military activities, training and rehabilitation. These environments result from the interactions between tissue motion, loading, biology, and morphology. Maintaining health of and/or repairing joint tissues requires targeting the "ideal" in vivo tissue mechanics (i.e., loading and strain), which may be enabled by real-time biofeedback. Recent research has shown that these biofeedback technologies are possible by integrating a patient's personalised digital twin and wireless wearable devices. Personalised digital twins are personalised neuromusculoskeletal rigid body and finite element models that work in real-time by code optimisation and artificial intelligence. Model personalisation is crucial in obtaining physically and physiologically valid predictions. CONCLUSIONS: Recent work has shown that laboratory-quality biomechanical measurements and modelling can be performed outside the laboratory with a small number of wearable sensors or computer vision methods. The next stage is to combine these technologies into well-designed easy to use products.

Cellular Immune Responses to SARS-CoV-2 in Exposed Seronegative Individuals.

Some SARS-CoV-2-exposed individuals develop immunity without overt infection. We identified 11 individuals who were negative by nucleic acid testing during prolonged close contact and with no serological diagnosis of infection. As this could reflect natural immunity, cross-reactive immunity from previous coronavirus exposure, abortive infection due to de novo immune responses, or other factors, our objective was to characterize immunity against SARS-CoV-2 in these individuals. Blood was processed into plasma and peripheral blood mononuclear cells (PBMC) and screened for IgG, IgA, and IgM antibodies (Ab) against SARS-CoV-2 and common ß-coronaviruses OC43 and HKU1. Receptor blocking activity and interferon-alpha (IFN-α) in plasma were also measured. Circulating T cells against SARS-CoV-2 were enumerated and CD4+ and CD8+ T cell responses discriminated after in vitro stimulation. Exposed uninfected individuals were seronegative against SARS-CoV-2 spike (S) and selectively reactive against OC43 nucleocapsid protein (N), suggesting common ß-coronavirus exposure induced Ab cross-reactive against SARS-CoV-2 N. There was no evidence of protection from circulating angiotensin-converting enzyme (ACE2) or IFN-α. Six individuals had T cell responses against SARS-CoV-2, with four involving CD4+ and CD8+ T cells. We found no evidence of protection from SARS-CoV-2 through innate immunity or immunity induced by common ß-coronaviruses. Cellular immune responses against SARS-CoV-2 were associated with time since exposure, suggesting that rapid cellular responses may contain SARS-CoV-2 infection below the thresholds required for a humoral response.

Imprinted Anti-Hemagglutinin and Anti-Neuraminidase Antibody Responses after Childhood Infections of A(H1N1) and A(H1N1)pdm09 Influenza Viruses.

Immune imprinting is a driver known to shape the anti-hemagglutinin (HA) antibody landscape of individuals born within the same birth cohort. With the HA and neuraminidase (NA) proteins evolving at different rates under immune selection pressures, anti-HA and anti-NA antibody responses since childhood influenza virus infections have not been evaluated in parallel at the individual level. This is partly due to the limited knowledge of changes in NA antigenicity, as seasonal influenza vaccines have focused on generating neutralizing anti-HA antibodies against HA antigenic variants. Here, we systematically characterized the NA antigenic variants of seasonal A(H1N1) viruses from 1977 to 1991 and completed the antigenic profile of N1 NAs from 1977 to 2015. We identified that NA proteins of A/USSR/90/77, A/Singapore/06/86, and A/Texas/36/91 were antigenically distinct and mapped N386K as a key determinant of the NA antigenic change from A/USSR/90/77 to A/Singapore/06/86. With comprehensive panels of HA and NA antigenic variants of A(H1N1) and A(H1N1)pdm09 viruses, we determined hemagglutinin inhibition (HI) and neuraminidase inhibition (NI) antibodies from 130 subjects born between 1950 and 2015. Age-dependent imprinting was observed for both anti-HA and anti-NA antibodies, with the peak HI and NI titers predominantly detected from subjects at 4 to 12 years old during the year of initial virus isolation, except the age-independent anti-HA antibody response against A(H1N1)pdm09 viruses. More participants possessed antibodies that reacted to multiple antigenically distinct NA proteins than those with antibodies that reacted to multiple antigenically distinct HA proteins. Our results support the need to include NA proteins in seasonal influenza vaccine preparations. IMPORTANCE Seasonal influenza vaccines have aimed to generate neutralizing anti-HA antibodies for protection since licensure. More recently, anti-NA antibodies have been established as an additional correlate of protection. While HA and NA antigenic changes occurred discordantly, the anti-HA and anti-NA antibody profiles have rarely been analyzed in parallel at the individual level, due to the limited knowledge on NA antigenic changes. By characterizing NA antigenic changes of A(H1N1) viruses, we determined the anti-HA and anti-NA antibody landscape against antigenically distinct A(H1N1) and A(H1N1)pdm09 viruses using sera of 130 subjects born between 1950 and 2015. We observed age-dependent imprinting of both anti-HA and anti-NA antibodies against strains circulated during the first decade of life. A total of 67.7% (88/130) and 90% (117/130) of participants developed cross-reactive antibodies to multiple HA and NA antigens at titers ≥1:40. With slower NA antigenic changes and cross-reactive anti-NA antibody responses, including NA protein in influenza vaccine preparation may enhance vaccine efficacy.