Advancing Simultaneous Extraction and Sequential Single-Particle ICP-MS Analysis for Metallic Nanoparticle Mixtures in Plant Tissues.

Engineered nanoparticles (ENPs) have been increasingly used in agricultural operations, leading to an urgent need for robust methods to analyze co-occurring ENPs in plant tissues. In response, this study advanced the simultaneous extraction of coexisting silver, cerium oxide, and copper oxide ENPs in lettuce shoots and roots using macerozyme R-10 and analyzed them by single-particle inductively coupled plasma-mass spectrometry (ICP-MS). Additionally, the standard stock suspensions of the ENPs were stabilized with citrate, and the long-term stability (up to 5 months) was examined for the first time. The method performance results displayed satisfactory accuracies and precisions and achieved low particle concentration and particle size detection limits. Significantly, the oven drying process was proved not to impact the properties of the ENPs; therefore, oven-dried lettuce tissues were used in this study, which markedly expanded the applicability of this method. This robust methodology provides a timely approach to characterize and quantify multiple coexisting ENPs in plants.

Genomic full-length sequence of the novel HLA-B*56:94 allele.

HLA-B*56:94 differs from HLA-B*56:05:01 by two non-synonymous nucleotide substitutions in exon 1 and synonymous nucleotide substitutions in exon 1 and exon 2.

Genomic full-length sequence of the novel HLA-A*02:1100 allele.

HLA-A*02:1100 differs from HLA-A*02:01:01:01 by a single non-synonymous nucleotide substitution in codon 76 of exon 2.

Discovery of the HLA-DQB1*06:465 allele, a variant of HLA-DQB1*06:04:01:01.

HLA-DQB1*06:465 differs from HLA-DQB1*06:04:01:01 by a non-synonymous nucleotide substitution in codon 38.

The HLA-B*58:01:43 allele identified in a Korean individual awaiting hematopoietic stem cell transplant.

HLA-B*58:01:43 differs from HLA-B*58:01:01:01 by one synonymous nucleotide substitution in codon 197.

Amendment-driven soil health restoration through soil pH and microbial robustness in a Cd/Cu-combined acidic soil: A ten-year in-situ field experiment.

Soil health arguably depends on biodiversity and has received wide attention in heavy-metal (HM) contaminated farmland remediation in recent years. However, long-term effects and mechanisms of soil amendment remain poorly understood with respect to soil microbal community. In this in-situ field study, four soil amendments (attapulgite-At, apatite-Ap, montmorillonite-M, lime-L) at three rates were applied once only for ten years in a cadmium (Cd)-copper (Cu) contaminated paddy soil deprecated for over five years. Results showed that after ten years and in compared with CK (no amendment), total Cd concentration and its risk in plot soils were not altered by amendments (p > 0.05), but total Cu concentration and its risk were significantly increased by both Ap and L, especially the former, rather than At and M (p < 0.05), through increased soil pH and enhanced bacterial alpha diversity as well as plant community. Soil microbial communities were more affected by amendment type (30%) than dosage (11%), microbial network characteristics were dominated by rare taxa, and soil multifunctionality was improved in Ap- and L-amended soils. A structural equation model (SEM) indicated that 57.3% of soil multifunctionality variances were accounted for by soil pH (+0.696) and microbial network robustness (-0.301). Moreover, microbial robustness could be potentially used as an indicator of soil multifunctionality, and Ap could be optimized to improve soil health in combined with biomass removal. These findings would advance the understanding of soil microbial roles, especially its network robustness, on soil multifunctionality for the remediation of metal contaminated soils and metal control management strategies in acidic soils. ENVIRONMENTAL IMPLICATION: Farmland soil contamination by heavy metals (HMs) has been becoming a serious global environmental challenge. However, most studies have been conducted over the short term, leading to a gap in the long-term remediation efficiency and ecological benefits of soil amendments. For the successful deployment of immobilization technologies, it is critical to understand the long-term stability of the immobilized HMs and soil health. Our study, to the best of our knowlege, is the first to state the long-term effects and mechanisms of soil amendments on soil health and optimize an effective and eco-friendly amendment for long-term Cd/Cu immobilization.

GTP Signaling Links Metabolism, DNA Repair, and Responses to Genotoxic Stress.

How cell metabolism regulates DNA repair is incompletely understood. Here, we define a GTP-mediated signaling cascade that links metabolism to DNA repair and has significant therapeutic implications. GTP, but not other nucleotides, regulates the activity of Rac1, a guanine nucleotide-binding protein, which promotes the dephosphorylation of serine 323 on Abl-interactor 1 (Abi-1) by protein phosphatase 5 (PP5). Dephosphorylated Abi-1, a protein previously not known to activate DNA repair, promotes nonhomologous end joining. In patients and mouse models of glioblastoma, Rac1 and dephosphorylated Abi-1 mediate DNA repair and resistance to standard-of-care genotoxic treatments. The GTP-Rac1-PP5-Abi-1 signaling axis is not limited to brain cancer, as GTP supplementation promotes DNA repair and Abi-1-S323 dephosphorylation in nonmalignant cells and protects mouse tissues from genotoxic insult. This unexpected ability of GTP to regulate DNA repair independently of deoxynucleotide pools has important implications for normal physiology and cancer treatment. SIGNIFICANCE: A newly described GTP-dependent signaling axis is an unexpected link between nucleotide metabolism and DNA repair. Disrupting this pathway can overcome cancer resistance to genotoxic therapy while augmenting it can mitigate genotoxic injury of normal tissues. This article is featured in Selected Articles from This Issue, p. 5.

Genomic full-length sequence of the novel HLA-DQB1*04:01:01:04 allele.

The sequence of HLA-DQB1*04:01:01:04 differs from HLA-DQB1*04:01:01:03 by four nucleotide deletion in intron 2.

The novel HLA-C allele, HLA-C*01:02:84 allele, identified in a solid organ transplantation donor.

HLA-C*01:02:84 differs from HLA-C*01:02:01:01 by one synonymous nucleotide substitution in codon 48.

Risks and benefits of anesthesia for combined pediatric procedures in the NIH undiagnosed diseases program.

PURPOSE: The NIH Undiagnosed Diseases Program (UDP) aims to provide diagnoses to patients who have previously received exhaustive evaluations yet remain undiagnosed. Patients undergo procedural anesthesia for deep phenotyping for analysis with genomic testing. METHODS: A retrospective chart review was performed to determine the safety and benefit of procedural anesthesia in pediatric patients in the UDP. Adverse perioperative events were classified as anesthesia-related complications or peri-procedural complications. The contribution of procedures performed under anesthesia to arriving at a diagnosis was also determined. RESULTS: From 2008 to 2020, 249 pediatric patients in the UDP underwent anesthesia for diagnostic procedures. The majority had a severe systemic disease (American Society for Anesthesiology status III, 79%) and/or a neurologic condition (91%). Perioperative events occurred in 45 patients; six of these were attributed to anesthesia. All patients recovered fully without sequelae. Nearly half of the 249 patients (49%) received a diagnosis, and almost all these diagnoses (88%) took advantage of information gleaned from procedures performed under anesthesia. CONCLUSIONS: The benefits of anesthesia involving multiple diagnostic procedures in a well-coordinated, multidisciplinary, research setting, such as in the pediatric UDP, outweigh the risks.

Local structural flexibility drives oligomorphism in computationally designed protein assemblies.

Many naturally occurring protein assemblies have dynamic structures that allow them to perform specialized functions. For example, clathrin coats adopt a wide variety of architectures to adapt to vesicular cargos of various sizes. Although computational methods for designing novel self-assembling proteins have advanced substantially over the past decade, most existing methods focus on designing static structures with high accuracy. Here we characterize the structures of three distinct computationally designed protein assemblies that each form multiple unanticipated architectures, and identify flexibility in specific regions of the subunits of each assembly as the source of structural diversity. Cryo-EM single-particle reconstructions and native mass spectrometry showed that only two distinct architectures were observed in two of the three cases, while we obtained six cryo-EM reconstructions that likely represent a subset of the architectures present in solution in the third case. Structural modeling and molecular dynamics simulations indicated that the surprising observation of a defined range of architectures, instead of non-specific aggregation, can be explained by constrained flexibility within the building blocks. Our results suggest that deliberate use of structural flexibility as a design principle will allow exploration of previously inaccessible structural and functional space in designed protein assemblies.

Magnetic resonance study of visceral, subcutaneous, liver and pancreas fat changes after 12 weeks intermittent fasting in obese participants with prediabetes.

BACKGROUND: It is not clear whether there are differences in proportions of fat loss from visceral:subcutaneous depots by probiotic supplementation, ethnicity or sex during weight loss; or whether visceral/pancreatic fat depot changes are related to changes in HbA1c. Our objective is to investigate whether weight loss from different fat depots is related to these factors during weight loss achieved by intermittent fasting. METHOD: Prediabetes participants on 5:2 intermittent fasting were randomized 1:1 to either daily probiotic or placebo for 12 weeks. Twenty-four patients had magnetic resonance imaging data at baseline and 12 weeks. RESULTS: After 12 weeks of intermittent fasting, subcutaneous fat (%) changed from 35.9 ± 3.1 to 34.4 ± 3.2, visceral fat (%) from 15.8 ± 1.3 to 14.8 ± 1.2, liver fat (%) from 8.7 ± 0.8 to 7.5 ± 0.7 and pancreatic fat (%) from 7.7 ± 0.5 to 6.5 ± 0.5 (all p < 0.001). Changes in weight, HbA1c, SAT, VAT, LF and PF did not differ significantly between probiotic and placebo groups. CONCLUSION: Overall weight loss was correlated with fat loss from subcutaneous depots. Losses from different fat depots did not correlate with changes in HbA1c or differ by probiotic supplementation, ethnicity or sex.

GTP signaling links metabolism, DNA repair, and responses to genotoxic stress.

How cell metabolism regulates DNA repair is incompletely understood. Here, we define a GTP-mediated signaling cascade that links metabolism to DNA repair and has significant therapeutic implications. GTP, but not other nucleotides, regulates the activity of Rac1, a G protein, that promotes the dephosphorylation of serine 323 on Abl-interactor 1 (Abi-1) by protein phosphatase 5 (PP5). Dephosphorylated Abi-1, a protein previously not known to activate DNA repair, promotes non-homologous end joining. In patients and mouse models of glioblastoma, Rac1 and dephosphorylated Abi-1 mediate DNA repair and resistance to standard of care genotoxic treatments. The GTP-Rac1-PP5-Abi-1 signaling axis is not limited to brain cancer, as GTP supplementation promotes DNA repair and Abi-1-S323 dephosphorylation in non-malignant cells and protects mouse tissues from genotoxic insult. This unexpected ability of GTP to regulate DNA repair independently of deoxynucleotide pools has important implications for normal physiology and cancer treatment.

Improving the secretion of designed protein assemblies through negative design of cryptic transmembrane domains.

Computationally designed protein nanoparticles have recently emerged as a promising platform for the development of new vaccines and biologics. For many applications, secretion of designed nanoparticles from eukaryotic cells would be advantageous, but in practice, they often secrete poorly. Here we show that designed hydrophobic interfaces that drive nanoparticle assembly are often predicted to form cryptic transmembrane domains, suggesting that interaction with the membrane insertion machinery could limit efficient secretion. We develop a general computational protocol, the Degreaser, to design away cryptic transmembrane domains without sacrificing protein stability. The retroactive application of the Degreaser to previously designed nanoparticle components and nanoparticles considerably improves secretion, and modular integration of the Degreaser into design pipelines results in new nanoparticles that secrete as robustly as naturally occurring protein assemblies. Both the Degreaser protocol and the nanoparticles we describe may be broadly useful in biotechnological applications.

Improving earthworm quality and complex metal removal from water by adding aquatic plant residues to cattle manure.

Vermicomposting is an economical and environmentally friendly process. However, related knowledge of vermicomposting aquatic plant residues (APRs), earthworm quality, and mechanisms for metal removal from water is still lacking. Nelumbo and Oenanthe javanica residues and their mixture were treated with Eisenia foetida and cattle manure for 45 days. Compared with the control comprising only cattle manure, addition of the APR mixture improved earthworm quality, mainly for low crude ash, high alkaloid compounds and different fat compositions in the Nelumbo residue and the balanced protein proportion of the APR mixture. All the vermicompost especial O. javanica residue added (VO) played efficient roles in removing metals from water initially containing 2.0 mg Cu L-1 and 8.0 mg Zn L-1. There were higher removal efficiencies (Ers) at the dosage of 4 g L-1 with a small microbial contribution. VO significantly increased Ers, which could be from the decrease of phylum Firmicutes (especial Bacteroides) abundance, stronger CH2, C = O, and CH, the addition of COOH groups, and higher organic matter and total phosphorus contents. The combination of VO and Hippuris vulgaris was optimized as an ecological and economical method for treating complex-metal polluted water. Moreover, our study widened the route for APR reuse.

Identification of the novel HLA-A*11:423 allele by sequencing-based typing.

HLA-A*11:423 differs from HLA-A*11:01:01:01 by a non-synonymous nucleotide substitution in codon 170, changing Arginine to Lysine.

Effects of conservation buffer systems on adsorption of fluorescent-labeled Escherichia coli.

The magnitude of bacterial transport through runoff into surface water or infiltration into groundwater is influenced by the adsorption processes in soil. The objective of this study was to evaluate fluorescent-labeled Escherichia coli (E. coli) adsorption by soil under agroforestry buffer (AB), grass buffer (GB), and row crop (RC) management. Adsorption experiments were conducted by inoculating three masses (0.5, 1, and 10 g) of each treatment (AB, GB, and RC) with E. coli O157:H7-GFP with concentration ranges of 105 -108 colony-forming units (cfu) ml-1 . Adsorption data were evaluated using Langmuir, Freundlich, and Temkin adsorption isotherm models. The Freundlich isotherm model described the observed data well for all treatments using the 10-g soil mass, with the R2 values closer to unity in all treatments. The Freundlich Kf parameter, an indicator of adsorption capacity, was higher for the AB treatment (9.93 cfu ml-1 ) compared with the GB and RC treatments (2.32 and 1.27 cfu ml-1 , respectively). The multiple pairwise comparisons test (Tukey test) of the Freundlich 1/nf parameter demonstrated a significant difference (p < .05) between the AB treatment and the RC and GB treatments. Similarly, the Kf values were significantly (p = .05) higher for the 10-g mass under the same test conditions, but no significant differences were observed in the 0.5- and 1-g masses. This study demonstrated that AB has a higher E. coli adsorption capacity and the potential for mitigating the effects of E. coli O157:H7 transport to surface or groundwater through the soil ecosystem.

An Adapted Deep Convolutional Neural Network for Automatic Measurement of Pancreatic Fat and Pancreatic Volume in Clinical Multi-Protocol Magnetic Resonance Images: A Retrospective Study with Multi-Ethnic External Validation.

Pancreatic volume and fat fraction are critical prognoses for metabolic diseases like type 2 diabetes (T2D). Magnetic Resonance Imaging (MRI) is a required non-invasive quantification method for the pancreatic fat fraction. The dramatic development of deep learning has enabled the automatic measurement of MR images. Therefore, based on MRI, we intend to develop a deep convolutional neural network (DCNN) that can accurately segment and measure pancreatic volume and fat fraction. This retrospective study involved abdominal MR images from 148 diabetic patients and 246 healthy normoglycemic participants. We randomly separated them into training and testing sets according to the proportion of 80:20. There were 2364 recognizable pancreas images labeled and pre-treated by an upgraded superpixel algorithm for a discernible pancreatic boundary. We then applied them to the novel DCNN model, mimicking the most accurate and latest manual pancreatic segmentation process. Fat phantom and erosion algorithms were employed to increase the accuracy. The results were evaluated by dice similarity coefficient (DSC). External validation datasets included 240 MR images from 10 additional patients. We assessed the pancreas and pancreatic fat volume using the DCNN and compared them with those of specialists. This DCNN employed the cutting-edge idea of manual pancreas segmentation and achieved the highest DSC (91.2%) compared with any reported models. It is the first framework to measure intra-pancreatic fat volume and fat deposition. Performance validation reflected by regression R2 value between manual operation and trained DCNN segmentation on the pancreas and pancreatic fat volume were 0.9764 and 0.9675, respectively. The performance of the novel DCNN enables accurate pancreas segmentation, pancreatic fat volume, fraction measurement, and calculation. It achieves the same segmentation level of experts. With further training, it may well surpass any expert and provide accurate measurements, which may have significant clinical relevance.

The Receptor Binding Domain of SARS-CoV-2 Lambda Variant Has a Better Chance Than the Delta Variant in Evading BNT162b2 COVID-19 mRNA Vaccine-Induced Humoral Immunity.

The SARS-CoV-2 Delta and Lambda variants had been named variants of concern (VOC) and variants of interest (VOI), respectively, by the World Health Organization (WHO). Both variants have two mutations in the spike receptor binding domain (RBD) region, with L452R and T478K mutations in the Delta variant, and L452Q and F490S mutations in the Lambda variant. We used surface plasmon resonance (SPR)-based technology to evaluate the effect of these mutations on human angiotensin-converting enzyme 2 (ACE2) and Bamlanivimab binding. The affinity for the RBD ligand, ACE2, of the Delta RBD is approximately twice as strong as that of the wild type RBD, an increase that accounts for the increased infectivity of the Delta variant. On the other hand, in spite of its amino acid changes, the Lambda RBD has similar affinity to ACE2 as the wild type RBD. The protective anti-wild type RBD antibody Bamlanivimab binds very poorly to the Delta RBD and not at all to the Lambda RBD. Nevertheless, serum antibodies from individuals immunized with the BNT162b2 vaccine were found to bind well to the Delta RBD, but less efficiently to the Lambda RBD in contrast. As a result, the blocking ability of ACE2 binding by serum antibodies was decreased more by the Lambda than the Delta RBD. Titers of sera from BNT162b2 mRNA vaccinated individuals dropped 3-fold within six months of vaccination regardless of whether the target RBD was wild type, Delta or Lambda. This may account partially for the fall off with time in the protective effect of vaccines against any variant.

Behavioural and dopaminergic signatures of resilience.

Chronic stress can have lasting adverse consequences in some individuals, yet others are resilient to the same stressor1,2. Susceptible and resilient individuals exhibit differences in the intrinsic properties of mesolimbic dopamine (DA) neurons after the stressful experience is over3-8. However, the causal links between DA, behaviour during stress and individual differences in resilience are unknown. Here we recorded behaviour in mice simultaneously with DA neuron activity in projections to the nucleus accumbens (NAc) (which signals reward9-12) and the tail striatum (TS) (which signals threat13-16) during social defeat. Supervised and unsupervised behavioural quantification revealed that during stress, resilient and susceptible mice use different behavioural strategies and have distinct activity patterns in DA terminals in the NAc (but not the TS). Neurally, resilient mice have greater activity near the aggressor, including at the onset of fighting back. Conversely, susceptible mice have greater activity at the offset of attacks and onset of fleeing. We also performed optogenetic stimulation of NAc-projecting DA neurons in open loop (randomly timed) during defeat or timed to specific behaviours using real-time behavioural classification. Both open-loop and fighting-back-timed activation promoted resilience and reorganized behaviour during defeat towards resilience-associated patterns. Together, these data provide a link between DA neural activity, resilience and resilience-associated behaviour during the experience of stress.