A new species of Cincticostella Allen, 1971 (Ephemeroptera, Ephemerellidae) from Yunnan, China and establishment of a new species complex.

Cincticostellajianchuan sp. nov. from Dali Bai Autonomous Prefecture, Yunnan Province, China, is described based on chorionic structure, nymph, and winged stages. The new species is closely related to C.fusca (Kang & Yang, 1995), but it can be distinguished in the male imago stage by its mesonotum and penes morphology, coloration, and the forking point of the stem of MA+Rs on the forewing; in the nymph stage, it can be distinguished by the length of the posterolateral projections of abdominal segment IX and the setation of the abdominal terga. Compared to other congeners, nymphs and male imagoes of the new species and C.fusca share several morphological characteristics, such as a larger body, mesothorax with medially notched anterolateral projections, forefemur without a subapical band of transverse spines of the nymphs, the area between C, Sc and R1 of the forewings distinctly pigmented, and an apical sclerite on the ventral face of the penes of the male imagoes, supporting the proposition of a new species complex, the jianchuan complex. The systematics of Cincticostella and related genera are discussed briefly.

Neuronal DNA repair reveals strategies to influence CRISPR editing outcomes.

Genome editing is poised to revolutionize treatment of genetic diseases, but poor understanding and control of DNA repair outcomes hinders its therapeutic potential. DNA repair is especially understudied in nondividing cells like neurons, which must withstand decades of DNA damage without replicating. This lack of knowledge limits the efficiency and precision of genome editing in clinically relevant cells. To address this, we used induced pluripotent stem cells (iPSCs) and iPSC-derived neurons to examine how postmitotic human neurons repair Cas9-induced DNA damage. We discovered that neurons can take weeks to fully resolve this damage, compared to just days in isogenic iPSCs. Furthermore, Cas9-treated neurons upregulated unexpected DNA repair genes, including factors canonically associated with replication. Manipulating this response with chemical or genetic perturbations allowed us to direct neuronal repair toward desired editing outcomes. By studying DNA repair in postmitotic human cells, we uncovered unforeseen challenges and opportunities for precise therapeutic editing.

Conserved and repetitive motifs in an intrinsically disordered protein drive ⍺-carboxysome assembly.

All cyanobacteria and some chemoautotrophic bacteria fix CO2 into sugars using specialized proteinaceous compartments called carboxysomes. Carboxysomes enclose the enzymes Rubisco and carbonic anhydrase inside a layer of shell proteins to increase the CO2 concentration for efficient carbon fixation by Rubisco. In the ⍺-carboxysome lineage, a disordered and highly repetitive protein named CsoS2 is essential for carboxysome formation and function. Without it, the bacteria require high CO2 to grow. How does a protein predicted to be lacking structure serve as the architectural scaffold for such a vital cellular compartment? In this study, we identify key residues present in the repeats of CsoS2, VTG and Y, which are necessary for building functional ⍺-carboxysomes in vivo. These highly conserved and repetitive residues contribute to the multivalent binding interaction and phase separation behavior between CsoS2 and shell proteins. We also demonstrate 3-component reconstitution of CsoS2, Rubisco, and shell proteins into spherical condensates, and show the utility of reconstitution as a biochemical tool to study carboxysome biogenesis. The precise self-assembly of thousands of proteins is crucial for carboxysome formation, and understanding this process could enable their use in alternative biological hosts or industrial processes as effective tools to fix carbon.

Enhancement of Low Temperature Superionic Conductivity by Suppression of Li Site Ordering in Li7Si2-xGexS7I.

Ge4+ substitution into the recently discovered superionic conductor Li7Si2S7I is demonstrated by synthesis of Li7Si2-xGexS7I, where x ≤ 1.2. The anion packing and tetrahedral silicon location of Li7Si2S7I are retained upon substitution. Single crystal X-ray diffraction shows that substitution of larger Ge4+ for Si4+ expands the unit cell volume and further increases Li+ site disorder, such that Li7Si0.88Ge1.12S7I has one Li+ site more (sixteen in total) than Li7Si2S7I. The ionic conductivity of Li7Si0.8Ge1.2S7I (x = 1.2) at 303 K is 1.02(3) × 10-2 S cm-1 with low activation energies for Li+ transport demonstrated over a wide temperature range by AC impedance and 7Li NMR spectroscopy. All sixteen Li+ sites remain occupied to temperatures as low as 30 K in Li7Si0.88Ge1.12S7I as a result of the structural expansion. This differs from Li7Si2S7I, where the partial Li+ site ordering observed below room temperature reduces the ionic conductivity. The suppression of Li+ site depopulation by Ge4+ substitution retains the high mobility to temperatures as low as 200 K, yielding low temperature performance comparable with state-of-the-art Li ion conducting materials.

Adverse event comparison between glucagon-like peptide-1 receptor agonists and other antiobesity medications following bariatric surgery.

AIM: To compare the incidence of adverse events (AEs) related to antiobesity medications (AOMs; glucagon-like peptide-1 receptor agonists [GLP-1RAs] vs. non-GLP-1RAs) after bariatric surgery. METHODS: This single-centre retrospective cohort included patients (aged 16-65 years) who had undergone laparoscopic Roux-en-Y gastric bypass or sleeve gastrectomy (cohort entry date) and initiated AOMs. Participants were categorized as users of US Food and Drug Administration (FDA)-approved, off-label, or GLP-1RA AOMs if documented as receiving the medication on or after cohort entry date. Non-GLP-1RA AOMs were phentermine, orlistat, topiramate, canagliflozin, dapagliflozin, empagliflozin, naltrexone, bupropion/naltrexone and phentermine/topiramate. GLP-1RA AOMs included: semaglutide, dulaglutide, exenatide and liraglutide. The primary outcome was AE incidence. Logistic regression was used to determine the association of AOM exposure with AEs. RESULTS: We identified 599 patients meeting our inclusion criteria, 83% of whom were female. Their median (interquartile range [IQR]) age was 47.8 (40.9-55.4) years. The median duration of surgery to AOM exposure was 30 months. GLP-1RAs use was not associated with higher odds of AEs: adjusted odds ratio (aOR) 1.1 (95% confidence interval [CI] 0.5-2.6) and aOR 1.1 (95% CI 0.6-2.3) for GLP-1RA versus FDA-approved and off-label AOM use, respectively. AOM initiation ≥12 months after surgery was associated with lower risk of AEs compared to <12 months (aOR 0.01 [95% CI 0.0-0.01]; p < 0.001). CONCLUSION: Our results showed that GLP-1RA AOMs were not associated with an increased risk of AEs compared to non-GLP-1RA AOMs in patients who had previously undergone bariatric surgery. Prospective studies are needed to identify the optimal timeframe for GLP-1RA initiation.

Rising Perfluorocyclobutane (PFC-318, c-C4F8) Emissions in China from 2011 to 2020 Inferred from Atmospheric Observations.

Global atmospheric emissions of perfluorocyclobutane (c-C4F8, PFC-318), a potent greenhouse gas, have increased rapidly in recent years. Combining atmospheric observations made at nine Chinese sites with a Lagrangian dispersion model-based Bayesian inversion technique, we show that PFC-318 emissions in China grew by approximately 70% from 2011 to 2020, rising from 0.65 (0.54-0.72) Gg year-1 in 2011 to 1.12 (1.05-1.19) Gg year-1 in 2020. The PFC-318 emission increase from China played a substantial role in the overall increase in global emissions during the study period, contributing 58% to the global total emission increase. This growth predominantly originated in eastern China. The regions with high emissions of PFC-318 in China overlap with areas densely populated with polytetrafluoroethylene (PTFE) factories, implying that fluoropolymer factories are important sources of PFC-318 emissions in China. Our investigation reveals an emission factor of approximately 3.02 g of byproduct PFC-318 emissions per kg of hydrochlorofluorocarbon-22 (HCFC-22) feedstock use in the production of tetrafluoroethylene (TFE) (for PTFE production) and hexafluoropropylene (HFP) if we assume all HCFC-22 produced for feedstock uses in China are pyrolyzed to produce PTFE and HFP. Further facility-level sampling and analysis are needed for a more precise evaluation of emissions from these factories.

The TOTAL trial for weight management participation: A randomized controlled trial protocol.

BACKGROUND: The three types of evidence-based treatment options for adults with overweight and obesity - behavioral weight management, anti-obesity medications (AOM), and bariatric surgery - are underutilized in the Veterans Health Administration (VHA) system. Our objective in this manuscript is to describe the study protocol for an adequately powered randomized controlled trial (RCT) of a behavioral intervention: TOTAL (Teaching Obesity Treatment Options to Adult Learners) to increase patient uptake of obesity treatment. METHODS: In this multi-site, parallel, RCT, eligible Veterans with a body mass index [BMI] ≥ 27 who had not received obesity treatment within the past 12 months were randomly assigned to TOTAL or usual care. TOTAL involves watching an 18-min video that highlights obesity health risks, pros/cons of all three evidence-based obesity treatments, and expected treatment outcomes. It also includes motivational sessions delivered via televideo at 2 weeks, 6 months, and 12 months after the video (target n = 494 participants). The primary outcome is initiation of behavioral weight management treatment within 18 months of randomization. Secondary outcomes include sustained behavioral weight management treatment, initiation of AOM, bariatric surgery referral, and weight change across 18 months. CONCLUSION: TOTAL, which seeks to increase delivery of weight management treatment within the largest integrated health system in the U.S., combines patient education with motivational interviewing components. If efficacious in this trial, further evaluation of intervention effectiveness and implementation throughout the VHA and other healthcare systems would be warranted.

High Throughput Phosphoglycolate Phosphatase Activity Assay Using Crude Leaf Extract and Recombinant Enzyme to Determine Kinetic Parameters Km and Vmax Using a Microplate Reader.

Determining enzyme activities involved in photorespiration, either in a crude plant tissue extract or in a preparation of a recombinant enzyme, is time-consuming, especially when large number of samples need to be processed. This chapter presents a phosphoglycolate phosphatase (PGLP) activity assay that is adapted for use in a 96-well microplate format. The microplate format for the assay requires fewer enzymes and reagents and allows rapid and less expensive measurement of PGLP enzyme activity. The small volume of reaction mix in a 96-well microplate format enables the determination of PGLP enzyme activity for screening many plant samples, multiple enzyme activities using the same protein extract, and/or identifying kinetic parameters for a recombinant enzyme. To assist in preparing assay reagents, we also present an R Shiny buffer preparation app for PGLP and other photorespiratory enzyme activities and a Km and Vmax calculation app.

Measuring and Quantifying Characteristics of the Post-illumination Burst.

Leaf-level gas exchange enables accurate measurements of net CO2 assimilation in the light, as well as CO2 respiration in the dark. Net positive CO2 assimilation in the light indicates that the gain of carbon by photosynthesis offsets the photorespiratory loss of CO2 and respiration of CO2 in the light (RL), while the CO2 respired in the dark is mainly attributed to respiration in the dark (RD). Measuring the CO2 release specifically from photorespiration in the light is challenging since net CO2 assimilation involves three concurrent processes (the velocity of rubisco carboxylation; vc, velocity of rubisco oxygenation; vo, and RL). However, by employing a rapid light-dark transient, it is possible to transiently measure some of the CO2 release from photorespiration without the background of vc-based assimilation in the dark. This method is commonly known as the post-illumination CO2 burst (PIB) and results in a "burst" of CO2 immediately after the transition to the dark. This burst can be quantitatively characterized using several approaches. Here, we describe how to set up a PIB measurement and provide some guidelines on how to analyze and interpret the data obtained using a PIB analysis application developed in R.

Electron Capture vs Transfer Dissociation for Site Determination of Tryptic Peptide Tyrosine Sulfation: Direct Detection of Fibrinogen Sulfation Sites and Identification of Novel Isobaric Interferences.

Tyrosine sulfation, an understudied but crucial post-translational modification, cannot be directly detected in conventional nanoflow liquid chromatography-tandem mass spectrometry (nanoLC-MS/MS) due to the extreme sulfate lability. Here, we report the detection of sulfate-retaining fragments from LC-electron capture dissociation (ECD) and nanoLC-electron transfer higher energy collision dissociation (EThcD). Sulfopeptide candidates were identified by Proteome Discoverer and MSFragger analysis of nanoLC-HCD MS/MS data and added to inclusion lists for LC-ECD or nanoLC-EThcD MS/MS. When this approach failed, targeted LC-ECD with fixed m/z isolation windows was performed. For the plasma protein fibrinogen, the known pyroglutamylated sulfopeptide QFPTDYDEGQDDRPK from the beta chain N-terminus was identified despite a complete lack of sulfate-containing fragment ions. The peptide QVGVEHHVEIEYD from the gamma-B chain C-terminus was also identified as sulfated or phosphorylated. This sulfopeptide is not annotated in Uniprot but was previously reported. MSFragger further identified a cysteine-containing peptide from the middle of the gamma chain as sulfated and deamidated. NanoLC-EThcD and LC-ECD MS/MS confirmed the two former sulfopeptides via sulfate-retaining fragment ions, whereas an unexpected fragmentation pattern was observed for the third sulfopeptide candidate. Manual interpretation of the LC-ECD spectrum revealed two additional isobaric identifications: a trisulfide-linked cysteinyl-glycine or a carbamidomethyl-dithiothreiotol covalent adduct. Synthesis of such adducts confirmed the latter identity.

Selective Gold Precipitation by a Tertiary Diamide Driven by Thermodynamic Control.

The simple diamide ligand L was previously shown to selectively precipitate gold from acidic solutions typical of e-waste leach streams, with precipitation of gallium, iron, tin, and platinum possible under more forcing conditions. Herein, we report direct competition experiments to afford the order of selectivity. Thermal analysis indicates that the gold-, gallium-, and iron-containing precipitates present as the most thermodynamically stable structures at room temperature, while the tin-containing structure does not. Computational modeling established that the precipitation process is thermodynamically driven, with ion exchange calculations matching the observed experimental selectivity ordering. Calculations also show that the stretched ligand conformation seen in the X-ray crystal structure of the gold-containing precipitate is more strained than in the structures of the other metal precipitates, indicating that intermolecular interactions likely dictate the selectivity ordering. This was confirmed through a combination of Hirshfeld, noncovalent interaction (NCI), and quantum theory of atoms in molecules (QTAIM) analyses, which highlight favorable halogen···halogen contacts between metalates and pseudo-anagostic C-H···metal interactions in the crystal structure of the gold-containing precipitate.

Synthesis, Structure, and Properties of CuBiSeCl2: A Chalcohalide Material with Low Thermal Conductivity.

Mixed anion halide-chalcogenide materials have recently attracted attention for a variety of applications, owing to their desirable optoelectronic properties. We report the synthesis of a previously unreported mixed-metal chalcohalide material, CuBiSeCl2 (Pnma), accessed through a simple, low-temperature solid-state route. The physical structure is characterized through single-crystal X-ray diffraction and reveals significant Cu displacement within the CuSe2Cl4 octahedra. The electronic structure of CuBiSeCl2 is investigated computationally, which indicates highly anisotropic charge carrier effective masses, and by experimental verification using X-ray photoelectron spectroscopy, which reveals a valence band dominated by Cu orbitals. The band gap is measured to be 1.33(2) eV, a suitable value for solar absorption applications. The electronic and thermal properties, including resistivity, Seebeck coefficient, thermal conductivity, and heat capacity, are also measured, and it is found that CuBiSeCl2 exhibits a low room temperature thermal conductivity of 0.27(4) W K-1 m-1, realized through modifications to the phonon landscape through increased bonding anisotropy.

Tuning the pore chemistry of Zr-MOFs for efficient metal ion capture from complex streams.

Metal-organic frameworks (MOFs) have shown promise for adsorptive separations of metal ions. Herein, MOFs based on highly stable Zr(IV) building units were systematically functionalized with targeted metal binding groups. Through competitive adsorption studies, it was shown that the selectivity for different metal ions was directly tunable through functional group chemistry.

An adapted protocol to derive microglia from stem cells and its application in the study of CSF1R-related disorders.

BACKGROUND: Induced pluripotent stem cell-derived microglia (iMGL) represent an excellent tool in studying microglial function in health and disease. Yet, since differentiation and survival of iMGL are highly reliant on colony-stimulating factor 1 receptor (CSF1R) signaling, it is difficult to use iMGL to study microglial dysfunction associated with pathogenic defects in CSF1R. METHODS: Serial modifications to an existing iMGL protocol were made, including but not limited to changes in growth factor combination to drive microglial differentiation, until successful derivation of microglia-like cells from an adult-onset leukoencephalopathy with axonal spheroids and pigmented glia (ALSP) patient carrying a c.2350G > A (p.V784M) CSF1R variant. Using healthy control lines, the quality of the new iMGL protocol was validated through cell yield assessment, measurement of microglia marker expression, transcriptomic comparison to primary microglia, and evaluation of inflammatory and phagocytic activities. Similarly, molecular and functional characterization of the ALSP patient-derived iMGL was carried out in comparison to healthy control iMGL. RESULTS: The newly devised protocol allowed the generation of iMGL with enhanced transcriptomic similarity to cultured primary human microglia and with higher scavenging and inflammatory competence at ~ threefold greater yield compared to the original protocol. Using this protocol, decreased CSF1R autophosphorylation and cell surface expression was observed in iMGL derived from the ALSP patient compared to those derived from healthy controls. Additionally, ALSP patient-derived iMGL presented a migratory defect accompanying a temporal reduction in purinergic receptor P2Y12 (P2RY12) expression, a heightened capacity to internalize myelin, as well as heightened inflammatory response to Pam3CSK4. Poor P2RY12 expression was confirmed to be a consequence of CSF1R haploinsufficiency, as this feature was also observed following CSF1R knockdown or inhibition in mature control iMGL, and in CSF1RWT/KO and CSF1RWT/E633K iMGL compared to their respective isogenic controls. CONCLUSIONS: We optimized a pre-existing iMGL protocol, generating a powerful tool to study microglial involvement in human neurological diseases. Using the optimized protocol, we have generated for the first time iMGL from an ALSP patient carrying a pathogenic CSF1R variant, with preliminary characterization pointing toward functional alterations in migratory, phagocytic and inflammatory activities.

Standardized indolent systemic mastocytosis evaluations across a healthcare system: implications for screening accuracy.

Timely diagnosis of systemic mastocytosis (SM) remains challenging due to care heterogeneity. We implemented a standardized approach for SM screening and diagnosis utilizing a novel healthcare system-wide international screening registry. A retrospective analysis assessed rates of SM, cutaneous mastocytosis (CM), and molecular diagnoses before and two years after care standardization. Accuracy of individual and combined SM screening tests - basal serum tryptase (BST) ≥11.5 and ≥20.0 ng/mL, REMA ≥2, monomorphic maculopapular CM, and elevated BST based upon tryptase genotype - was analyzed. Tryptase genotyping and high-sensitivity KIT p.D816V testing increased substantially two years following care standardization. SM diagnoses doubled from 47 to 94 and KIT p.D816V molecular diagnoses increased from 24 to 79. Mean BST and KIT p.D816V variant allele frequency (VAF) values were significantly lower in patients diagnosed after standardization. Hereditary-alpha tryptasemia prevalence was increased in SM prior to care standardization at 4/30 (13.3%) but reflected the general population prevalence two years later at 5/76 (6.6%). Elevated BST based upon genotype and BST ≥11.5 ng/mL had the highest sensitivities at 84.2% and 88.3%, respectively. Presence of monomorphic MPCM, elevated BST based upon tryptase genotype, and the combination of REMA ≥2 with elevated BST based upon tryptase genotype had specificities >90%. BST >20.0 ng/mL had low sensitivity and specificity and was not required to establish any indolent SM diagnosis. Care standardization increased SM diagnosis rates, particularly in patients with low BSTs. Stratifying BST based upon genotype had the best overall sensitivity and specificity of any indolent SM screening test and improved the REMA score specificity.

A Novel Model for Xenograft Right Ventricle to Pulmonary Artery Conduit.

The last 40 years have shown dramatic improvement in outcomes for neonatal cardiac surgery for a spectrum of congenital heart disease diagnoses. With more patients surviving into adulthood, the long-term impact of initial management strategies of these patients has come into focus. This is particularly true for patients with pediatric heart valve disease. Many patients born with right ventricular to pulmonary artery (RVPA) discontinuity require placement of a valved conduit in the neonatal period. Valved conduit options are limited in this patient population due to patient size and inability to respond to somatic growth. Genetically engineered porcine (GEP) donors may offer a xenograft conduit alternative that can grow with the patient. We have developed a model utilizing GEP donor RVPA conduits placed in infantile nonhuman primate (NHP) recipients. Our recipient is maintained on single-drug immunosuppression and demonstrates no evidence of pulmonary valve insufficiency or stenosis during short-term follow-up. Further studies and long-term outcomes are necessary to determine the utility of this technology in human application.

A map of the rubisco biochemical landscape.

Rubisco is the primary CO2 fixing enzyme of the biosphere yet has slow kinetics. The roles of evolution and chemical mechanism in constraining the sequence landscape of rubisco remain debated. In order to map sequence to function, we developed a massively parallel assay for rubisco using an engineered E. coli where enzyme function is coupled to growth. By assaying >99% of single amino acid mutants across CO2 concentrations, we inferred enzyme velocity and CO2 affinity for thousands of substitutions. We identified many highly conserved positions that tolerate mutation and rare mutations that improve CO2 affinity. These data suggest that non-trivial kinetic improvements are readily accessible and provide a comprehensive sequence-to-function mapping for enzyme engineering efforts.

Exploring Social Support Dynamics After Bariatric Surgery: Insights From Patients and Providers.

INTRODUCTION: Weight loss after bariatric surgery is impacted by several factors, and social support is one of them. Our objective was to characterize patient and provider perceptions about social support after bariatric surgery. METHODS: We reported a secondary analysis of qualitative data acquired from semi-structured interviews conducted from January-November 2020 with bariatric surgery patients and providers. Participants included primary care providers, health psychologists, registered dietitians, bariatric surgeons, and patients with at least 1 y of follow-up after their bariatric procedure. Interview guides were designed using a hybrid of Andersen's Behavioral Model of Health Services and Torain's Framework for Surgical Disparities. Using directed content analysis, study team members generated codes, which were categorized into themes about social support pertaining to dietary habits, physical activity, and follow-up care. RESULTS: Forty-five participants were interviewed, including 24 patients (83% female; 79% White; mean age 50.6 ± 10.7 y) and 21 providers (six primary care providers, four health psychologists, five registered dieticians, and six bariatric surgeons). We identified four themes relating to social support affecting weight loss after surgery: (1) family involvement in helping patients adjust to the bariatric diet, (2) engagement in activities with partners/friends, (3) help with transportation to appointments, and (4) life stressors experienced by patients within their social relationships. CONCLUSIONS: Continued assessment of interpersonal factors after bariatric surgery is essential for weight loss maintenance. Providers can contribute by reinforcing the facilitators of social support and making referrals that may help patients overcome barriers to social support for sustained weight loss after surgery.