bluuebunny/crossref_metadata_2025_split huggingface dataset:
Random effects modelling vs logistic regression for the inclusion of cluster level covariates in propensity scores for medical device and surgical epidemiology
Authors: Du Mike , Prats-Uribe Albert , Khalid Sara , Prieto-Alhambra Daniel , Strauss Victoria Y , Khalid Sara 
Published: February 2022

AbstractPurposeSurgeon and hospital related features such as surgeries volume can be associated with treatment choices and treatment outcomes. Accounting for these covariates with propensity score (PS) analysis can be challenging due to clustered nature of the data. Previous studies have not focused solely on the PS estimation strategy when treatment effects are estimated using random effects model(REM). We studied PS estimation for clustered data using REM compared with logistic regression.MethodsSix different PS estimation strategies were tested using simulations with variable cluster-level confounding intensity (odds ratio(OR)=1.01 to OR=2.5): i) logistic regression PS excluding cluster- level confounders; ii) logistic regression PS including cluster-level confounders; iii) same as ii) but including cross-level interactions; iv), v) and vi), similar to i), ii) and iii) respectively but using REM instead of logistic regression PS. Same analysis were tested in a randomised controlled trial emulation of partial vs total knee replacement surgery. Simulation metrics included bias and mean square error (MSE). For trial emulation, we compared observational vs trial-based treatment effect estimates.ResultsIn most simulated scenarios, logistic regression including cluster-level confounders gave more accurate estimates with the lowest bias and MSE. E.g. with 50 clusters x 200 individuals and confounding intensity OR=1.5, the relative bias= 10% and MSE= 0.003 for (i), compared to 21% and, 0.010 for (iv). In the Trial emulation, all 6 PS strategies gave similar treatment effect estimates.ConclusionsLogistic regression including patient and surgeon/hospital-level confounders appears to be the preferred strategy for PS estimation. Further investigation with more complex clustered structure is suggested.Competing interestsProf. Prieto-Alhambra’s research group has received grant support from Amgen, Chesi-Taylor, Novartis, and UCB Biopharma. His department has received advisory or consultancy fees from Amgen, Astellas, AstraZeneca, Johnson, and Johnson, and UCB Biopharma and fees for speaker services from Amgen and UCB Biopharma. Janssen, on behalf of IMI-funded EHDEN and EMIF consortiums, and Synapse Management Partners have supported training programs organised by DPA’s department and open for external participants organized by his department outside submitted work.Ethics Approval and Informed ConsentThis study was approved by the secretary of state, having considered the recommendation from the Confidentiality Advisory Group (CAG reference: 17/CAG/0174). Informed ethical approval was given on the use of pseudonymised patients data included in the study.
Intracellular flow cytometry complements RT-qPCR detection of circulating SARS-CoV-2 variants of concern
Authors: Vanhulle Emiel , Provinciael Becky , Stroobants Joren , Camps Anita , Maes Piet , Vermeire Kurt 
Published: February 2022

AbstractDespite the efficacy of current vaccines against SARS-CoV-2, the spread of the virus is still not under control, as evidenced by the ongoing circulation of the highly contagious SARS-CoV-2 Omicron variant. Basic and antiviral research on SARS-CoV-2 relies on cellular assays of virus replication in vitro. In addition, accurate detection of virus-infected cells and released virus particles is needed to study virus replication and to profile new candidate antiviral drugs. Here, by flow cytometry, we detect SARS-CoV-2 infection at single cell level and distinguish infected Vero E6 cells from uninfected bystander cells. Furthermore, based on the viral nucleocapsid expression, subpopulations of infected cells that are in an early or late phase of viral replication can be differentiated. Importantly, this flow cytometric technique complements RT-qPCR detection and can be applied to all current SARS-CoV-2 variants of concern, including the highly mutated Omicron variant.Method summaryThis study describes the characterization of SARS-CoV-2 infected cells using intracellular flow cytometric viral nucleocapsid staining that complements RT-qPCR quantification of viral RNA. The technique makes it possible to distinguish between infected cells in the early (low N) or late phase (high N) of viral replication. It can also be applied to the different variants of concern of SARS-CoV-2, including the Omicron variant.
Bile Salts and N-Acetyl-D-Glucosamine induce transcription of the vieSAB Operon in Vibrio cholerae El Tor
Authors: Shull Lauren M. , Camilli Andrew 
Published: February 2022

AbstractThe vieSAB operon regulates motility, biofilm formation and cholera toxin production in the classical biotype of V. cholerae. Classical biotype mutants lacking the cyclic-di-GMP phosphodiesterase and response regulator vieA or its cognate sensor kinase vieS fail to effectively colonize the infant mouse small intestine, are less motile than the wild-type strain, and are hyper-biofilm formers. However, deletion of vieA or vieS in the currently-circulating El Tor biotype does not have a demonstrable effect on these phenotypes. To begin to understand the role of vieSAB in the El Tor biotype, we studied its regulation and the effect of vieA expression on bacterial physiology, separating the functional roles of VieA’s enzymatic and DNA-binding activities. We identify bile salts and N-acetylglucosamine, which can be found at high concentration in the small intestine, as stimulating expression of vieSAB during in vitro growth. We show that, in contrast to many response regulators, VieA does not regulate its own expression. Together, these data indicate that V. cholerae upregulates vieSAB in response to small-intestinal signals, allowing VieA to alter the physiology of the cell through its transcriptional regulation and cyclic-di-GMP signaling activities in order to adapt to the human host.
Exploring different virulent proteins of human respiratory syncytial virus for designing a novel epitope-based polyvalent vaccine: Immunoinformatics and molecular dynamics approaches
Authors: Moin Abu Tayab , Ullah Md. Asad , Patil Rajesh B. , Faruqui Nairita Ahsan , Sarkar Bishajit , Araf Yusha , Das Sowmen , Kapil Uddin Khaza Md. , Hossain Md Shakhawat , Miah Md. Faruque , Moni Mohammad Ali , Chowdhury Dil Umme Salma , Islam Saiful 
Published: February 2022

AbstractHuman Respiratory Syncytial Virus (RSV) is one of the most prominent causes of lower respiratory tract infections (LRTI), contributory to infecting people from all age groups - a majority of which comprises infants and children. The implicated severe RSV infections lead to numerous deaths of multitudes of the overall population, predominantly the children, every year. Consequently, despite several distinctive efforts to develop a vaccine against the RSV as a potential countermeasure, there is no approved or licensed vaccine available yet, to control the RSV infection effectively. Therefore, through the utilization of immunoinformatics tools, a computational approach was taken in this study, to design and construct a multi-epitope polyvalent vaccine against the RSV-A and RSV-B strains of the virus. Potential predictions of the T-cell and B-cell epitopes were followed by extensive tests of antigenicity, allergenicity, toxicity, conservancy, homology to human proteome, transmembrane topology, and cytokine-inducing ability. The most promising epitopes (i.e. 13 CTL epitopes, 9 HTL epitopes, and 10 LBL epitopes) exhibiting full conservancy were then selected for designing the peptide fusion with appropriate linkers, having hBD-3 as the adjuvant. The peptide vaccine was modeled, refined, and validated to further improve the structural attributes. Following this, molecular docking analysis with specific TLRs was carried out which revealed excellent interactions and global binding energies. Additionally, molecular dynamics (MD) simulation was conducted which ensured the stability of the interactions between vaccine and TLR. Furthermore, mechanistic approaches to imitate and predict the potential immune response generated by the administration of vaccines were determined through immune simulations. Owing to an overall evaluation, in silico cloning was carried out in efforts to generate recombinant pETite plasmid vectors for subsequent mass production of the vaccine peptide, incorporated within E.coli. However, more in vitro and in vivo experiments can further validate its efficacy against RSV infections.
Imaging minimal bacteria at the nanoscale: a reliable and versatile process to perform Single Molecule Localization Microscopy in mycoplasmas
Authors: Rideau Fabien , Villa Audrey , Belzanne Pauline , Verdier Emeline , Hosy Eric , Arfi Yonathan 
Published: February 2022

AbstractMycoplasmas are the smallest free-living organisms. These bacteria are important models for both fundamental and Synthetic Biology, owing to their highly reduced genomes. They are also relevant in the medical and veterinary fields, as they are pathogenic of both humans and most livestock species. Mycoplasma cells have minute sizes, often in the 300-800 nanometers range. As these dimensions are close to the diffraction limit of visible light, fluorescence imaging in mycoplasmas is often poorly informative. Recently developed “Super-Resolution Imaging” techniques can break this diffraction limit, improving the imaging resolution by an order of magnitude and offering a new nanoscale vision of the organization of these bacteria. These techniques have however not been applied to mycoplasmas before. Here, we describe an efficient and reliable protocol to perform Single-Molecule Localization Microscopy (SMLM) imaging in mycoplasmas. We provide a polyvalent transposon-based system to express the photo-convertible fluorescent protein mEos3.2, enabling Photo-Activated Localization Microscopy (PALM) in most Mycoplasma species. We also describe the application of direct STochastic Optical Reconstruction Microscopy (dSTORM). We showcase the potential of these techniques by studying the subcellular localization of two proteins of interest. Our work highlights the benefits of state-of-the-art microscopy techniques for mycoplasmology and provides an incentive to further the development SMLM strategies to study these organisms in the future.ImportanceMycoplasmas are important models in biology, as well as highly problematic pathogens in the medical and veterinary fields. The very small size of these bacteria, well below the micron, limits the usefulness of traditional fluorescence imaging methods as their resolution limit is similar to the dimensions of the cells. Here, to bypass this issue, we established a set of state-of-the-art “Super-Resolution Microscopy” techniques in a wide range of Mycoplasma species. We describe two strategies: PALM, based on the expression of a specific photo-convertible fluorescent protein; and dSTORM, based on fluorophore-coupled antibody labeling. With these methods, we successfully performed single-molecule imaging of proteins of interest at the surface of the cells and in the cytoplasm, at lateral resolutions well below 50 nanometers. Our work paves the way toward a better understanding of mycoplasma’s biology through imaging of subcellular structures at the nanometer scale.
CACNB4 overexpression decreases dendritic spine density in sex-specific manner
Authors: Parker Emily M. , Kindja Nathan L. , DeGiosio Rebecca A. , Salisbury Ryan B. , Krivinko Josh M. , Cheetham Claire E. J. , MacDonald Matthew L. , Sweet Robert A. 
Published: February 2022

ABSTRACTThe canonical voltage-gated calcium channel (VGCC) subunit complex is comprised of the α1 subunit, the ion permeable channel, plus three auxiliary subunits: β, α2δ and γ. β is the most extensively studied auxiliary subunit and is necessary for proper forward trafficking of the α1 subunit to the plasma membrane. α1 subunits mediate voltagedependent movement of calcium ions into the cytoplasm of neurons, including at dendritic sites, where increased intracellular calcium initiates signaling cascades that shape structural and functional plasticity of dendritic spines. Genetic studies strongly implicate calcium signaling dysfunction in the etiology of neurodevelopmental disorders including schizophrenia. Dendritic spine density (DSD) is significantly decreased in schizophrenia in primary auditory cortex where DSD is driven by loss of small spines, and small spine loss is associated with increased peptide levels of ALFDFLK found in the VGCC β subunit β4. OverexpessingCACNB4to increase β4 levels selectively reduced small spine density in cortical neuron cultures. The studies described herein set out to validate thisin vitroobservation in an intact mammalian system within a neurodevelopmental context. We overexpressedCACNB4in neurodevelopment and assessed DSD and morphology in cerebral cortex of male and female mice at an adult timpoint. We then characterized β protein levels and β4 protein-protein interactions in male and female mouse cortex. Overexpression selectively reduced small dendritic spine density but this effect was present only in female mice and did not appear to result from estrous stage. Instead, the sex-dependent effect on DSD corresponded to sex differences in the β4 interactome of male versus female mice: the VGCC β subunit β1b was significantly enriched in the β4 interactome of brain tissue of male mice, and thus may have served to mitigate VGCC overexpression-mediated spine loss in male mice.
Sex and age differences in cognitive bias and neural activation in response to cognitive bias
Authors: Hodges Travis E. , Lee Grace Y. , Noh Sophia H. , Galea Liisa A.M. 
Published: February 2022

AbstractCognitive symptoms of depression, including negative cognitive bias, are more severe in women than in men. Current treatments to reduce negative cognitive bias are not effective and sex differences in the neural activity underlying cognitive bias may play a role. Here we examined sex and age differences in cognitive bias and functional connectivity in a novel paradigm. Male and female rats underwent an 18-day cognitive bias procedure, in which they learned to discriminate between two contexts (shock paired context A, no-shock paired context B), during either adolescence (postnatal day (PD 40)), young adulthood (PD 100), or middle-age (PD 210). Cognitive bias was measured as freezing behaviour in response to an ambiguous context (context C), with freezing levels akin to the shock paired context coded as negative bias. All animals learned to discriminate between the two contexts, regardless of sex or age. However, adults (young adults, middle-aged) displayed a greater negative cognitive bias compared to adolescents, and middle-aged males had a greater negative cognitive bias than middle-aged females. Females had greater neural activation of the nucleus accumbens, amygdala, and hippocampal regions to the ambiguous context compared to males, and young rats (adolescent, young adults) had greater neural activation in these regions compared to middle-aged rats. Functional connectivity between regions involved in cognitive bias differed by age and sex, and only adult males had negative correlations between the frontal regions and hippocampal regions. These findings highlight the importance of examining age and sex when investigating the underpinnings of negative cognitive bias and lay the groundwork for determining what age- and sex-specific regions to target in future cognitive bias studies.HighlightsMiddle-aged males had a greater negative cognitive bias than middle-aged femalesAdult rats displayed a greater negative cognitive bias compared to adolescentsGreater neural activity in females than males in limbic and reward regionsGreater role of the frontal cortex activation in the cognitive bias of adultsFunctional connectivity in response to cognitive bias differed by age and sex
The new archaeal order Lutiacidiplasmatales reveals convergent evolution in Thermoplasmatota
Authors: Sheridan Paul O. , Meng Yiyu , Williams Tom A. , Gubry-Rangin Cécile 
Published: February 2022

AbstractThe archaeal Terrestrial Miscellaneous Euryarchaeota Group (TMEG) has been identified in various environments, and the single genome investigated thus far suggests that these organisms are anaerobic sulfite reducers. We assembled 35 new TMEG genomes that, based on genome analysis, appear to possess aerobic and facultative anaerobic lifestyles and may oxidise rather than reduce sulfite. We propose naming this order (representing 16 genera) “Lutiacidiplasmatales” due to their occurrence in various acidic environments and placement within the phylum Thermoplasmatota. A phylum-level analysis revealed that Thermoplasmatota evolution had been punctuated by several periods of high levels of novel gene family acquisition. Several essential metabolisms, such as aerobic respiration and acid tolerance, were likely acquired independently by divergent lineages through convergent evolution rather than inherited from a common ancestor. Ultimately, this study describes the terrestrially prevalent Lutiacidiciplasmatales and indicates convergent evolution as an important evolutionary driving force in archaeal lineages with complex histories.
Breasi-CRISPR: an efficient genome editing method to interrogate protein localization and protein-protein interactions in the embryonic mouse cortex
Authors: Meyerink Brandon L. , Pratiksha KC , Tiwari Neeraj K. , Kittock Claire M. , Klein Abigail , Evans Claire , Pilaz Louis-Jan 
Published: February 2022

AbstractIn developing tissues, knowing the localization and interactors of proteins of interest is key to understanding their function. This can be challenging when the researched protein lacks reliable antibodies. Here, we combine Easi-CRISPR with in utero electroporation to tag endogenous proteins within embryonic mouse brains. This method is called Breasi-CRISPR (Brain Easi-CRISPR), and enables knock-in of both short and long epitope tag sequences in genes of interest with high efficiency. Using Breasi-CRISPR, we were able to visualize epitope tagged proteins known to have either high or low expression levels, such as ACTB, LMNB1, EMD, FMRP, NOTCH1, and RPL22. Detection was possible by immunohistochemistry as soon as one day after electroporation at embryonic day 13 (E13). Two and five days after electroporation, we observed efficient gene editing in up to 50% of electroporated cells. Moreover, tagged proteins could be detected by immunoblotting in lysates from individual cortices two days after electroporation. Next, we demonstrated that Breasi-CRISPR enables the tagging of proteins with fluorophores in an efficient manner, allowing the visualization of endogenous proteins via live-imaging in organotypic brain slices two days after electroporation. Finally, we used Breasi-CRISPR to perform co-IP mass-spectrometry analyses of tagged autism-related protein FMRP to discover its interactome in the embryonic cortex. Together, these data show Breasi-CRISPR is a powerful tool with diverse applications that will propel the understanding of protein function in neurodevelopment.
A dendritic mechanism for balancing synaptic flexibility and stability
Authors: Yaeger Courtney E. , Vardalaki Dimitra , Zhang Qinrong , Pham Trang L.D. , Brown Norma J. , Ji Na , Harnett Mark T. 
Published: February 2022

AbstractBiological and artificial neural networks learn by modifying synaptic weights, but it is unclear how these systems can retain previous knowledge and also acquire new information. Here we show that cortical pyramidal neurons can solve this plasticity-versus-stability dilemma by differentially regulating synaptic plasticity at distinct dendritic compartments. Oblique dendrites of adult mouse layer 5b cortical pyramidal neurons selectively received monosynaptic thalamic input, integrated linearly, and— surprisingly—lacked synaptic potentiation. In contrast, basal dendrites, which do not receive thalamic input, exhibited conventional NMDA receptor-mediated supralinear integration and synaptic potentiation. Congruently, spiny synapses on oblique branches showed low structural plasticity in vivo. A selective decline in NMDAR activity and expression at synapses on oblique dendrites was controlled by an experience-dependent critical period. Our results demonstrate a new biological mechanism for how single neurons can safeguard a set of inputs from ongoing plasticity by altering synaptic properties at distinct dendritic domains.
Microbiomes of the bloom-forming alga,Phaeocystis globosa, are stable, consistently recruited communities with symbiotic and opportunistic modes
Authors: Mars Brisbin Margaret , Mitarai Satoshi , Saito Mak A. , Alexander Harriet 
Published: February 2022

AbstractPhaeocystisis a cosmopolitan, bloom-forming phytoplankton genus that contributes significantly to global carbon and sulfur cycles. During blooms,Phaeocystisspecies produce large carbon-rich colonies, thus creating a unique interface for bacterial interactions. While bacteria are known to interact with phytoplankton—e.g., they promote growth by producing phytohormones and vitamins—such interactions have not been shown forPhaeocystis. Therefore, we investigated the composition and function ofP. globosamicrobiomes. Specifically, we tested whether microbiome compositions are consistent across individual colonies from fourP. globosastrains, whether similar microbiomes are re-recruited after antibiotic treatment, and how microbiomes affectP. globosagrowth under limiting conditions. Results illuminated a core colonialP. globosamicrobiome—including bacteria from the orders Alteromonadales, Burkholderiales, and Rhizobiales—that was re-recruited after microbiome disruption. Consistent microbiome composition and recruitment is indicative thatP. globosamicrobiomes are stable-state systems undergoing deterministic community assembly and suggests there are specific, beneficial interactions betweenPhaeocystisand bacteria. Growth experiments with axenic and nonaxenic cultures demonstrated that microbiomes allowed continued growth when B-vitamins were withheld, but that microbiomes accelerated culture collapse when nitrogen was withheld. In sum, this study reveals interactions betweenPhaeocystiscolonies and microbiome bacteria that could influence large-scale phytoplankton bloom dynamics and biogeochemical cycles.
Identifying inputs to visual projection neurons in Drosophila lobula by analyzing connectomic data
Authors: Tanaka Ryosuke , Clark Damon A. 
Published: February 2022

AbstractElectron microscopy-based connectomes provide important insights into how visual circuitry of fruit fly Drosophila computes various visual features, guiding and complementing behavioral and physiological studies. However, connectomic analyses of lobula, a putative center of object-like feature detection, remains underdeveloped, largely because of incomplete data on the inputs to the brain region. Here, we attempted to map the columnar inputs into the Drosophila lobula neuropil by performing connectivity- and morphology-based clustering on a densely reconstructed connectome dataset. While the dataset mostly lacked visual neuropils other than lobula, which would normally help identify inputs to lobula, our clustering analysis successfully extracted clusters of cells with homogeneous connectivity and morphology, likely representing genuine cell types. We were able to draw a correspondence between the resulting clusters and previously identified cell types, revealing previously undocumented connectivity between lobula input and output neurons. While future, more complete connectomic reconstructions are necessary to verify the results presented here, they can serve as a useful basis for formulating hypotheses on mechanisms of visual feature detection in lobula.