Alarming antibody evasion properties of rising SARS-CoV-2 BQ and XBB subvariants.

The BQ and XBB subvariants of SARS-CoV-2 Omicron are now rapidly expanding, possibly due to altered antibody evasion properties deriving from their additional spike mutations. Here, we report that neutralization of BQ.1, BQ.1.1, XBB, and XBB.1 by sera from vaccinees and infected persons was markedly impaired, including sera from individuals boosted with a WA1/BA.5 bivalent mRNA vaccine. Titers against BQ and XBB subvariants were lower by 13- to 81-fold and 66- to 155-fold, respectively, far beyond what had been observed to date. Monoclonal antibodies capable of neutralizing the original Omicron variant were largely inactive against these new subvariants, and the responsible individual spike mutations were identified. These subvariants were found to have similar ACE2-binding affinities as their predecessors. Together, our findings indicate that BQ and XBB subvariants present serious threats to current COVID-19 vaccines, render inactive all authorized antibodies, and may have gained dominance in the population because of their advantage in evading antibodies.

Microbiota imbalance induced by dietary sugar disrupts immune-mediated protection from metabolic syndrome.

How intestinal microbes regulate metabolic syndrome is incompletely understood. We show that intestinal microbiota protects against development of obesity, metabolic syndrome, and pre-diabetic phenotypes by inducing commensal-specific Th17 cells. High-fat, high-sugar diet promoted metabolic disease by depleting Th17-inducing microbes, and recovery of commensal Th17 cells restored protection. Microbiota-induced Th17 cells afforded protection by regulating lipid absorption across intestinal epithelium in an IL-17-dependent manner. Diet-induced loss of protective Th17 cells was mediated by the presence of sugar. Eliminating sugar from high-fat diets protected mice from obesity and metabolic syndrome in a manner dependent on commensal-specific Th17 cells. Sugar and ILC3 promoted outgrowth of Faecalibaculum rodentium that displaced Th17-inducing microbiota. These results define dietary and microbiota factors posing risk for metabolic syndrome. They also define a microbiota-dependent mechanism for immuno-pathogenicity of dietary sugar and highlight an elaborate interaction between diet, microbiota, and intestinal immunity in regulation of metabolic disorders.

Intestinal microbiota-specific Th17 cells possess regulatory properties and suppress effector T cells via c-MAF and IL-10.

Commensal microbes induce cytokine-producing effector tissue-resident CD4+ T cells, but the function of these T cells in mucosal homeostasis is not well understood. Here, we report that commensal-specific intestinal Th17 cells possess an anti-inflammatory phenotype marked by expression of interleukin (IL)-10 and co-inhibitory receptors. The anti-inflammatory phenotype of gut-resident commensal-specific Th17 cells was driven by the transcription factor c-MAF. IL-10-producing commensal-specific Th17 cells were heterogeneous and derived from a TCF1+ gut-resident progenitor Th17 cell population. Th17 cells acquired IL-10 expression and anti-inflammatory phenotype in the small-intestinal lamina propria. IL-10 production by CD4+ T cells and IL-10 signaling in intestinal macrophages drove IL-10 expression by commensal-specific Th17 cells. Intestinal commensal-specific Th17 cells possessed immunoregulatory functions and curbed effector T cell activity in vitro and in vivo in an IL-10-dependent and c-MAF-dependent manner. Our results suggest that tissue-resident commensal-specific Th17 cells perform regulatory functions in mucosal homeostasis.

Antigenicity and receptor affinity of SARS-CoV-2 BA.2.86 spike.

A severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) Omicron subvariant, BA.2.86, has emerged and spread to numerous countries worldwide, raising alarm because its spike protein contains 34 additional mutations compared with its BA.2 predecessor1. We examined its antigenicity using human sera and monoclonal antibodies (mAbs). Reassuringly, BA.2.86 was no more resistant to human sera than the currently dominant XBB.1.5 and EG.5.1, indicating that the new subvariant would not have a growth advantage in this regard. Importantly, sera from people who had XBB breakthrough infection exhibited robust neutralizing activity against all viruses tested, suggesting that upcoming XBB.1.5 monovalent vaccines could confer added protection. Although BA.2.86 showed greater resistance to mAbs to subdomain 1 (SD1) and receptor-binding domain (RBD) class 2 and 3 epitopes, it was more sensitive to mAbs to class 1 and 4/1 epitopes in the 'inner face' of the RBD that is exposed only when this domain is in the 'up' position. We also identified six new spike mutations that mediate antibody resistance, including E554K that threatens SD1 mAbs in clinical development. The BA.2.86 spike also had a remarkably high receptor affinity. The ultimate trajectory of this new SARS-CoV-2 variant will soon be revealed by continuing surveillance, but its worldwide spread is worrisome.

Antibody evasion properties of SARS-CoV-2 Omicron sublineages.

The identification of the Omicron (B.1.1.529.1 or BA.1) variant of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in Botswana in November 20211 immediately caused concern owing to the number of alterations in the spike glycoprotein that could lead to antibody evasion. We2 and others3-6 recently reported results confirming such a concern. Continuing surveillance of the evolution of Omicron has since revealed the rise in prevalence of two sublineages, BA.1 with an R346K alteration (BA.1+R346K, also known as BA.1.1) and B.1.1.529.2 (BA.2), with the latter containing 8 unique spike alterations and lacking 13 spike alterations found in BA.1. Here we extended our studies to include antigenic characterization of these new sublineages. Polyclonal sera from patients infected by wild-type SARS-CoV-2 or recipients of current mRNA vaccines showed a substantial loss in neutralizing activity against both BA.1+R346K and BA.2, with drops comparable to that already reported for BA.1 (refs. 2,3,5,6). These findings indicate that these three sublineages of Omicron are antigenically equidistant from the wild-type SARS-CoV-2 and thus similarly threaten the efficacies of current vaccines. BA.2 also exhibited marked resistance to 17 of 19 neutralizing monoclonal antibodies tested, including S309 (sotrovimab)7, which had retained appreciable activity against BA.1 and BA.1+R346K (refs. 2-4,6). This finding shows that no authorized monoclonal antibody therapy could adequately cover all sublineages of the Omicron variant, except for the recently authorized LY-CoV1404 (bebtelovimab).

Striking antibody evasion manifested by the Omicron variant of SARS-CoV-2.

The B.1.1.529/Omicron variant of SARS-CoV-2 was only recently detected in southern Africa, but its subsequent spread has been extensive, both regionally and globally1. It is expected to become dominant in the coming weeks2, probably due to enhanced transmissibility. A striking feature of this variant is the large number of spike mutations3 that pose a threat to the efficacy of current COVID-19 vaccines and antibody therapies4. This concern is amplified by the findings of our study. Here we found that B.1.1.529 is markedly resistant to neutralization by serum not only from patients who recovered from COVID-19, but also from individuals who were vaccinated with one of the four widely used COVID-19 vaccines. Even serum from individuals who were vaccinated and received a booster dose of mRNA-based vaccines exhibited substantially diminished neutralizing activity against B.1.1.529. By evaluating a panel of monoclonal antibodies against all known epitope clusters on the spike protein, we noted that the activity of 17 out of the 19 antibodies tested were either abolished or impaired, including ones that are currently authorized or approved for use in patients. Moreover, we also identified four new spike mutations (S371L, N440K, G446S and Q493R) that confer greater antibody resistance on B.1.1.529. The Omicron variant presents a serious threat to many existing COVID-19 vaccines and therapies, compelling the development of new interventions that anticipate the evolutionary trajectory of SARS-CoV-2.

Fast and efficient template-mediated synthesis of genetic variants.

Efficient methods for the generation of specific mutations enable the study of functional variations in natural populations and lead to advances in genetic engineering applications. Here, we present a new approach, mutagenesis by template-guided amplicon assembly (MEGAA), for the rapid construction of kilobase-sized DNA variants. With this method, many mutations can be generated at a time to a DNA template at more than 90% efficiency per target in a predictable manner. We devised a robust and iterative protocol for an open-source laboratory automation robot that enables desktop production and long-read sequencing validation of variants. Using this system, we demonstrated the construction of 31 natural SARS-CoV2 spike gene variants and 10 recoded Escherichia coli genome fragments, with each 4 kb region containing up to 150 mutations. Furthermore, 125 defined combinatorial adeno-associated virus-2 cap gene variants were easily built using the system, which exhibited viral packaging enhancements of up to 10-fold compared with wild type. Thus, the MEGAA platform enables generation of multi-site sequence variants quickly, cheaply, and in a scalable manner for diverse applications in biotechnology.

DNA-based memory devices for recording cellular events.

Measuring biological data across time and space is critical for understanding complex biological processes and for various biosurveillance applications. However, such data are often inaccessible or difficult to directly obtain. Less invasive, more robust and higher-throughput biological recording tools are needed to profile cells and their environments. DNA-based cellular recording is an emerging and powerful framework for tracking intracellular and extracellular biological events over time across living cells and populations. Here, we review and assess DNA recorders that utilize CRISPR nucleases, integrases and base-editing strategies, as well as recombinase and polymerase-based methods. Quantitative characterization, modelling and evaluation of these DNA-recording modalities can guide their design and implementation for specific application areas.

Engineering living and regenerative fungal-bacterial biocomposite structures.

Engineered living materials could have the capacity to self-repair and self-replicate, sense local and distant disturbances in their environment, and respond with functionalities for reporting, actuation or remediation. However, few engineered living materials are capable of both responsivity and use in macroscopic structures. Here we describe the development, characterization and engineering of a fungal-bacterial biocomposite grown on lignocellulosic feedstocks that can form mouldable, foldable and regenerative living structures. We have developed strategies to make human-scale biocomposite structures using mould-based and origami-inspired growth and assembly paradigms. Microbiome profiling of the biocomposite over multiple generations enabled the identification of a dominant bacterial component, Pantoea agglomerans, which was further isolated and developed into a new chassis. We introduced engineered P. agglomerans into native feedstocks to yield living blocks with new biosynthetic and sensing-reporting capabilities. Bioprospecting the native microbiota to develop engineerable chassis constitutes an important strategy to facilitate the development of living biomaterials with new properties and functionalities.

Robust direct digital-to-biological data storage in living cells.

DNA has been the predominant information storage medium for biology and holds great promise as a next-generation high-density data medium in the digital era. Currently, the vast majority of DNA-based data storage approaches rely on in vitro DNA synthesis. As such, there are limited methods to encode digital data into the chromosomes of living cells in a single step. Here, we describe a new electrogenetic framework for direct storage of digital data in living cells. Using an engineered redox-responsive CRISPR adaptation system, we encoded binary data in 3-bit units into CRISPR arrays of bacterial cells by electrical stimulation. We demonstrate multiplex data encoding into barcoded cell populations to yield meaningful information storage and capacity up to 72 bits, which can be maintained over many generations in natural open environments. This work establishes a direct digital-to-biological data storage framework and advances our capacity for information exchange between silicon- and carbon-based entities.

Metagenomic engineering of the mammalian gut microbiome in situ.

Engineering of microbial communities in open environments remains challenging. Here we describe a platform used to identify and modify genetically tractable mammalian microbiota by engineering community-wide horizontal gene transfer events in situ. With this approach, we demonstrate that diverse taxa in the mouse gut microbiome can be modified directly with a desired genetic payload. In situ microbiome engineering in living animals allows novel capabilities to be introduced into established communities in their native milieu.

Quantifying spatiotemporal variability and noise in absolute microbiota abundances using replicate sampling.

Metagenomic sequencing has enabled detailed investigation of diverse microbial communities, but understanding their spatiotemporal variability remains an important challenge. Here, we present decomposition of variance using replicate sampling (DIVERS), a method based on replicate sampling and spike-in sequencing. The method quantifies the contributions of temporal dynamics, spatial sampling variability, and technical noise to the variances and covariances of absolute bacterial abundances. We applied DIVERS to investigate a high-resolution time series of the human gut microbiome and a spatial survey of a soil bacterial community in Manhattan's Central Park. Our analysis showed that in the gut, technical noise dominated the abundance variability for nearly half of the detected taxa. DIVERS also revealed substantial spatial heterogeneity of gut microbiota, and high temporal covariances of taxa within the Bacteroidetes phylum. In the soil community, spatial variability primarily contributed to abundance fluctuations at short time scales (weeks), while temporal variability dominated at longer time scales (several months).

The alarmins high mobility group box protein 1 and S100A8/A9 display different inflammatory profiles after acute knee injury.

OBJECTIVE: To compare the concentrations of high mobility group box 1 protein (HMGB1) and S100A8/A9 in synovial fluid between patients with knee injuries and osteoarthritis (OA), and knee healthy subjects. To investigate associations of alarmin levels with different joint injuries and with biomarkers of inflammation, Wnt signaling, complement system, bone and cartilage degradation. METHODS: HMGB1 and S100A8/A9 were measured in synovial fluid by immunoassays in patients with knee injuries, with OA and from knee healthy subjects, and were related to time from injury and with biomarkers obtained from previous studies. Hierarchical cluster and enrichment analyses of biomarkers associated to HMGB1 and S100A8/A9 were performed. RESULTS: The synovial fluid HMGB1 and S100A8/A9 concentrations were increased early after knee injury; S100A8/A9 levels were negatively associated to time after injury and was lower in the old compared to recent injury group, while HMGB1 was not associated to time after injury. The S100A8/A9 levels were also increased in OA. The initial inflammatory response was similar between the alarmins, and HMGB1 and S100A8/A9 shared 9 out of 20 enriched pathways. The alarmins displayed distinct response profiles, HMGB1 being associated to cartilage biomarkers while S100A8/A9 was associated to proinflammatory cytokines. CONCLUSIONS: HMGB1 and S100A8/A9 are increased as an immediate response to knee trauma. While they share many features in inflammatory and immunoregulatory mechanisms, S100A8/A9 and HMGB1 are associated to different downstream responses, which may have impact on the OA progression after acute knee injuries.

Scalable and cost-effective ribonuclease-based rRNA depletion for transcriptomics.

Bacterial RNA sequencing (RNA-seq) is a powerful approach for quantitatively delineating the global transcriptional profiles of microbes in order to gain deeper understanding of their physiology and function. Cost-effective bacterial RNA-seq requires efficient physical removal of ribosomal RNA (rRNA), which otherwise dominates transcriptomic reads. However, current methods to effectively deplete rRNA of diverse non-model bacterial species are lacking. Here, we describe a probe and ribonuclease based strategy for bacterial rRNA removal. We implemented the method using either chemically synthesized oligonucleotides or amplicon-based single-stranded DNA probes and validated the technique on three novel gut microbiota isolates from three distinct phyla. We further showed that different probe sets can be used on closely related species. We provide a detailed methods protocol, probe sets for >5000 common microbes from RefSeq, and an online tool to generate custom probe libraries. This approach lays the groundwork for large-scale and cost-effective bacterial transcriptomics studies.

Food-related parenting practices and styles in households with sibling children: A scoping review.

Interventions that promote responsive feeding in early childhood have been shown to reduce obesity risks. However, interventions mostly target parent-child dyads without considering the complexities of implementing responsive feeding across multiple children within a family unit. This scoping review aims to assess the extent and nature of current literature examining feeding in the context of siblings. Six electronic databases were searched (APA PsycINFO, CINAHL, Embase, Medline, ProQuest Dissertations & Theses Global, and Scopus) for articles published up until November 25, 2021. Studies were included if they compared the use of parent feeding practices and/or styles for two or more siblings aged ≤18 years. Data were extracted from relevant studies and analysed using basic descriptive statistics. A total of 18 studies from North America (n = 12) and Europe (n = 6) were included, with the majority targeting children between 6 and 18 years of age (n = 12). All studies were cross-sectional, with most designed to test differences in parent-reported feeding practices for siblings, primarily restriction and/or pressure to eat, in relation to differences in their characteristics (n = 12). The studies provide some evidence that parents may modify certain feeding practices or styles for siblings in response to differences in their characteristics, such as weight status and eating behaviours. Future research should examine processes that underlie feeding decisions in the context of siblings, including the contexts and consequences of differential feeding, with particular focus on early childhood when feeding interventions may be most effective.

Metagenomic mining of regulatory elements enables programmable species-selective gene expression.

Robust and predictably performing synthetic circuits rely on the use of well-characterized regulatory parts across different genetic backgrounds and environmental contexts. Here we report the large-scale metagenomic mining of thousands of natural 5' regulatory sequences from diverse bacteria, and their multiplexed gene expression characterization in industrially relevant microbes. We identified sequences with broad and host-specific expression properties that are robust in various growth conditions. We also observed substantial differences between species in terms of their capacity to utilize exogenous regulatory sequences. Finally, we demonstrate programmable species-selective gene expression that produces distinct and diverse output patterns in different microbes. Together, these findings provide a rich resource of characterized natural regulatory sequences and a framework that can be used to engineer synthetic gene circuits with unique and tunable cross-species functionality and properties, and also suggest the prospect of ultimately engineering complex behaviors at the community level.

Programmable CRISPR-Cas transcriptional activation in bacteria.

Programmable gene activation enables fine-tuned regulation of endogenous and synthetic gene circuits to control cellular behavior. While CRISPR-Cas-mediated gene activation has been extensively developed for eukaryotic systems, similar strategies have been difficult to implement in bacteria. Here, we present a generalizable platform for screening and selection of functional bacterial CRISPR-Cas transcription activators. Using this platform, we identified a novel CRISPR activator, dCas9-AsiA, that could activate gene expression by more than 200-fold across genomic and plasmid targets with diverse promoters after directed evolution. The evolved dCas9-AsiA can simultaneously mediate activation and repression of bacterial regulons in E. coli. We further identified hundreds of promoters with varying basal expression that could be induced by dCas9-AsiA, which provides a rich resource of genetic parts for inducible gene activation. Finally, we show that dCas9-AsiA can be ported to other bacteria of clinical and bioindustrial relevance, thus enabling bacterial CRISPRa in more application areas. This work expands the toolbox for programmable gene regulation in bacteria and provides a useful resource for future engineering of other bacterial CRISPR-based gene regulators.

Light Dark Matter Search with a High-Resolution Athermal Phonon Detector Operated above Ground.

We present limits on spin-independent dark matter-nucleon interactions using a 10.6 g Si athermal phonon detector with a baseline energy resolution of σ_{E}=3.86±0.04(stat)_{-0.00}^{+0.19}(syst) eV. This exclusion analysis sets the most stringent dark matter-nucleon scattering cross-section limits achieved by a cryogenic detector for dark matter particle masses from 93 to 140 MeV/c^{2}, with a raw exposure of 9.9 g d acquired at an above-ground facility. This work illustrates the scientific potential of detectors with athermal phonon sensors with eV-scale energy resolution for future dark matter searches.

Constraints on Lightly Ionizing Particles from CDMSlite.

The Cryogenic Dark Matter Search low ionization threshold experiment (CDMSlite) achieved efficient detection of very small recoil energies in its germanium target, resulting in sensitivity to lightly ionizing particles (LIPs) in a previously unexplored region of charge, mass, and velocity parameter space. We report first direct-detection limits calculated using the optimum interval method on the vertical intensity of cosmogenically produced LIPs with an electric charge smaller than e/(3×10^{5}), as well as the strongest limits for charge ≤e/160, with a minimum vertical intensity of 1.36×10^{-7} cm^{-2} s^{-1} sr^{-1} at charge e/160. These results apply over a wide range of LIP masses (5 MeV/c^{2} to 100 TeV/c^{2}) and cover a wide range of ßγ values (0.1-10^{6}), thus excluding nonrelativistic LIPs with ßγ as small as 0.1 for the first time.

Salinity-dependent expression of calcium-sensing receptors in Atlantic salmon (Salmo salar) tissues.

Multiple reports suggest that calcium-sensing receptors (CaSRs) are involved in calcium homeostasis, osmoregulation, and/or salinity sensing in fish (Loretz 2008, Herberger and Loretz 2013). We have isolated three unique full-length CaSR cDNAs from Atlantic salmon (Salmo salar) kidney that share many features with other reported CaSRs. Using anti-CaSR antibodies and PCR primers specific for individual salmon CaSR transcripts we show expression in osmoregulatory, neuroendocrine and sensory tissues. Furthermore, CaSRs are expressed in different patterns in salmon tissues where mRNA and protein expression are modified by freshwater or seawater acclimation. For example, in seawater, CaSR mRNA and protein expression is increased significantly in kidney as compared to freshwater. Electrophysiological recordings of olfactory responses produced upon exposure of salmon olfactory epithelium to CaSR agonists suggest a role for CaSRs in chemoreception in this species consistent with other freshwater, anadromous, and marine species where similar olfactory responses to divalent and polyvalent cations have been reported. These data provide further support for a role of CaSR proteins in osmoregulatory and sensory functions in Atlantic salmon, an anadromous species that experiences a broad range of environmental salinities in its life history.