Bacteria conjugate ubiquitin-like proteins to interfere with phage assembly.

Several immune pathways in humans conjugate ubiquitin-like proteins to virus and host molecules as a means of antiviral defence1-5. Here we studied an antiphage defence system in bacteria, comprising a ubiquitin-like protein, ubiquitin-conjugating enzymes E1 and E2, and a deubiquitinase. We show that during phage infection, this system specifically conjugates the ubiquitin-like protein to the phage central tail fibre, a protein at the tip of the tail that is essential for tail assembly as well as for recognition of the target host receptor. Following infection, cells encoding this defence system release a mixture of partially assembled, tailless phage particles and fully assembled phages in which the central tail fibre is obstructed by the covalently attached ubiquitin-like protein. These phages show severely impaired infectivity, explaining how the defence system protects the bacterial population from the spread of phage infection. Our findings demonstrate that conjugation of ubiquitin-like proteins is an antiviral strategy conserved across the tree of life.

The molecular mechanism of on-demand sterol biosynthesis at organelle contact sites.

Contact-sites are specialized zones of proximity between two organelles, essential for organelle communication and coordination. The formation of contacts between the Endoplasmic Reticulum (ER), and other organelles, relies on a unique membrane environment enriched in sterols. However, how these sterol-rich domains are formed and maintained had not been understood. We found that the yeast membrane protein Yet3, the homolog of human BAP31, is localized to multiple ER contact sites. We show that Yet3 interacts with all the enzymes of the post-squalene ergosterol biosynthesis pathway and recruits them to create sterol-rich domains. Increasing sterol levels at ER contacts causes its depletion from the plasma membrane leading to a compensatory reaction and altered cell metabolism. Our data shows that Yet3 provides on-demand sterols at contacts thus shaping organellar structure and function. A molecular understanding of this protein's functions gives new insights into the role of BAP31 in development and pathology.

Principles for Adapting Assessments of Executive Function across Cultural Contexts.

Direct assessments of executive functions (EFs) are increasingly used in research and clinical settings, with a central assumption that they assess "universal" underlying skills. Their use is spreading globally, raising questions about the cultural appropriateness of assessments devised in Western industrialized countries. We selectively reviewed multidisciplinary evidence and theory to identify sets of cultural preferences that may be at odds with the implicit assumptions of EF assessments. These preferences relate to motivation and compliance; cultural expectations for interpersonal engagement; contextualized vs. academic thinking; cultural notions of speed and time; the willingness to be silly, be incorrect, or do the opposite; and subject-matter familiarity. In each case, we discuss how the cultural preference may be incompatible with the assumptions of assessments, and how future research and practice can address the issue. Many of the cultural preferences discussed differ between interdependent and independent cultures and between schooled and unschooled populations. Adapting testing protocols to these cultural preferences in different contexts will be important for expanding our scientific understanding of EF from the narrow slice of the human population that has participated in the research to date.

Rim aperture of yeast autophagic membranes balances cargo inclusion with vesicle maturation.

Autophagy eliminates cytoplasmic material by engulfment in membranous vesicles targeted for lysosome degradation. Nonselective autophagy coordinates sequestration of bulk cargo with the growth of the isolation membrane (IM) in a yet-unknown manner. Here, we show that in the budding yeast Saccharomyces cerevisiae, IMs expand while maintaining a rim sufficiently wide for sequestration of large cargo but tight enough to mature in due time. An obligate complex of Atg24/Snx4 with Atg20 or Snx41 assembles locally at the rim in a spatially extended manner that specifically depends on autophagic PI(3)P. This assembly stabilizes the open rim to promote autophagic sequestration of large cargo in correlation with vesicle expansion. Moreover, constriction of the rim by the PI(3)P-dependent Atg2-Atg18 complex and clearance of PI(3)P by Ymr1 antagonize rim opening to promote autophagic maturation and consumption of small cargo. Tight regulation of membrane rim aperture by PI(3)P thus couples the mechanism and physiology of nonselective autophagy.

Variability in the age of schooling contributes to the link between literacy and numeracy in Côte d'Ivoire.

Literacy and numeracy are correlated throughout development, however, our understanding of this relation is limited. We explored the predictors of literacy and numeracy covariance (i.e., shared fluency between literacy and numeracy) in children (N = 1167, girls = 563) in rural Côte d'Ivoire, with specific focus on how developmental timing of instruction may relate to covariance. Many Ivorian children experience late enrollment and grade repetition, leading to variation in age-for-grade; participants were between grades 1 to 6, but their ages ranged from 5 to 15 (M = 9.19, SD = 2.07). Phonological awareness, numerical magnitude, ordinality, working memory, and inhibitory control were cognitive predictors of covariance. Age-for-grade was negatively related to covariance suggesting that covariance is related to timing of instruction.

Nudging parents and teachers to improve learning and reduce child labor in Cote d'Ivoire.

Whether SMS-based nudge interventions can increase parent engagement and improve child learning outcomes across diverse contexts such as rural West Africa is unknown. We conducted a school-randomized trial to test the impacts of an audio or text-message intervention (two messages per week for one school year) to parents and teachers of second and fourth grade students (N = 100 schools, 2246 students) in Cote d'Ivoire. Schools were randomly assigned to have messages sent to (i) parents only, (ii) teachers only, (iii) parents and teachers together, or (iv) control. There were statistically non-significant impacts of the parents-only treatment on learning, although with typical effect sizes (d = 0.08, p = 0.158), and marginally statistically significant increases in child labor (d = 0.11, p < 0.10). We find no impacts of the other treatment conditions. Subgroup analyses based on pre-registered subgroups show significantly larger improvements in learning for children with below-median baseline learning levels for the parents-only arm and negative impacts on learning for girls for the teachers-only arm, suggesting different conclusions regarding impacts on equity for vulnerable children.

Unraveling the multifaceted resilience of arsenic resistant bacterium Deinococcus indicus.

Arsenic (As) is a toxic heavy metal widely found in the environment that severely undermines the integrity of water resources. Bioremediation of toxic compounds is an appellative sustainable technology with a balanced cost-effective setup. To pave the way for the potential use of Deinococcus indicus, an arsenic resistant bacterium, as a platform for arsenic bioremediation, an extensive characterization of its resistance to cellular insults is paramount. A comparative analysis of D. indicus cells grown in two rich nutrient media conditions (M53 and TGY) revealed distinct resistance patterns when cells are subjected to stress via UV-C and methyl viologen (MV). Cells grown in M53 demonstrated higher resistance to both UV-C and MV. Moreover, cells grow to higher density upon exposure to 25 mM As(V) in M53 in comparison with TGY. This analysis is pivotal for the culture of microbial species in batch culture bioreactors for bioremediation purposes. We also demonstrate for the first time the presence of polyphosphate granules in D. indicus which are also found in a few Deinococcus species. To extend our analysis, we also characterized DiArsC2 (arsenate reductase) involved in arsenic detoxification and structurally determined different states, revealing the structural evidence for a catalytic cysteine triple redox system. These results contribute for our understanding into the D. indicus resistance mechanism against stress conditions.

Visualization of Organelles In Situ by Cryo-STEM Tomography.

Cryogenic electron microscopy (cryo-EM) relies on the imaging of biological or organic specimens embedded in their native aqueous medium; water is solidified into a glass (i.e., vitrified) without crystallization. The cryo-EM method is widely used to determine the structure of biological macromolecules recently at a near-atomic resolution. The approach has been extended to the study of organelles and cells using tomography, but the conventional mode of wide-field transmission EM imaging suffers a severe limitation in the specimen thickness. This has led to a practice of milling thin lamellae using a focused ion beam; the high resolution is obtained by subtomogram averaging from the reconstructions, but three-dimensional relations outside the remaining layer are lost. The thickness limitation can be circumvented by scanned probe imaging, similar to the scanning EM or the confocal laser scanning microscope. While scanning transmission electron microscopy (STEM) in materials science provides atomic resolution in single images, the sensitivity of cryogenic biological specimens to electron irradiation requires special considerations. This protocol presents a setup for cryo-tomography using STEM. The basic topical configuration of the microscope is described for both two- and three-condenser systems, while automation is provided by the non-commercial SerialEM software. Enhancements for batch acquisition and correlative alignment to previously-acquired fluorescence maps are also described. As an example, we show the reconstruction of a mitochondrion, pointing out the inner and outer membrane and calcium phosphate granules, as well as surrounding microtubules, actin filaments, and ribosomes. Cryo-STEM tomography excels in revealing the theater of organelles in the cytoplasm and, in some cases, even the nuclear periphery of adherent cells in culture.

Correlating cryo-super resolution radial fluctuations and dual-axis cryo-scanning transmission electron tomography to bridge the light-electron resolution gap.

Visualization of organelles and their interactions with other features in the native cell remains a challenge in modern biology. We have introduced cryo-scanning transmission electron tomography (CSTET), which can access 3D volumes on the scale of 1 micron with a resolution of nanometers, making it ideal for this task. Here we introduce two relevant advances: (a) we demonstrate the utility of multi-color super-resolution radial fluctuation light microscopy under cryogenic conditions (cryo-SRRF), and (b) we extend the use of deconvolution processing for dual-axis CSTET data. We show that cryo-SRRF nanoscopy is able to reach resolutions in the range of 100 nm, using commonly available fluorophores and a conventional widefield microscope for cryo-correlative light-electron microscopy. Such resolution aids in precisely identifying regions of interest before tomographic acquisition and enhances precision in localizing features of interest within the 3D reconstruction. Dual-axis CSTET tilt series data and application of entropy regularized deconvolution during post-processing results in close-to-isotropic resolution in the reconstruction without averaging. The integration of cryo-SRRF with deconvolved dual-axis CSTET provides a versatile workflow for studying unique objects in a cell.

Sociodemographic predictors of depression and anxiety symptomatology among parents in rural Côte d'Ivoire.

BACKGROUND: In Côte d'Ivoire, cocoa farming is a widespread practice in rural households, an occupation with increased risks of depression and anxiety exacerbated by economic instability. We used the Goldberg-18 Depression and Anxiety diagnostic tool to identify predictors of depressive and anxiety symptomatology among a sample of parents in rural cocoa farming communities. METHODS: In a cross-sectional survey, the Goldberg-18 was administered to Ivorian parents (N = 2471). Confirmatory factor analysis (CFA) was conducted to confirm the factor structure of the assessment tool, and Ordinary Least Squares (OLS) regression with clustered standard errors was used to identify sociodemographic predictors of symptomatology. RESULTS: CFA showed adequate fit statistics for a two-factor model measuring depressive and anxiety symptoms. Among respondents, 87 % screened positive for requiring further referral for clinical diagnosis. Sociodemographic predictors of depressive and anxiety symptoms were similar for males and females. For the total sample, higher monthly income, more years of education, and belonging to the Mandinka ethnic group predicted fewer depressive and anxiety symptoms. In contrast, higher depressive and anxiety symptomatology were associated with age. Single marital status predicted increased anxiety but not depressive symptoms for the full sample model and the female only sample, but not the male sample. LIMITATIONS: This is a cross-sectional study. CONCLUSIONS: The Goldberg-18 measures distinct domains of depressive and anxiety symptoms in a rural Ivorian sample. Age and single marital status are predictors of increased symptoms. Higher monthly income, higher education, and certain ethnic affiliations are protective factors.

Teacher depressive symptoms and children's school readiness in Ghana.

This study investigated associations between kindergarten teachers' (N = 208) depressive symptoms and students' (Ghanaian nationals, N = 1490, Mage  = 5.8) school-readiness skills (early literacy, early numeracy, social-emotional skills, and executive function) across 208 schools in Ghana over one school year. Teachers' depressive symptoms in the fall negatively predicted students' overall school-readiness skills in the spring, controlling for school-readiness skills in the fall. These results were primarily driven by social-emotional skills (r = .1-.3). There was evidence of heterogeneity by students' fall skill levels; teacher depressive symptoms predicted more negative spring overall school readiness for children who had higher fall school-readiness skills. Findings underscore the importance of teachers' mental health in early childhood education globally, with implications for policy and practice.

Elemental compositions of sea urchin larval cell vesicles evaluated by cryo-STEM-EDS and cryo-SEM-EDS.

During spicule formation in sea urchin larvae, calcium ions translocate within the primary mesenchymal cells (PMCs) from endocytosed seawater vacuoles to various organelles and vesicles where they accumulate, and subsequently precipitate. During this process, calcium ions are concentrated by more than three orders of magnitude, while other abundant ions (Na, Mg) must be removed. To obtain information about the overall ion composition in the vesicles, we used quantitative cryo-SEM-EDS and cryo-STEM-EDS analyzes. For cryo-STEM-EDS, thin (500 nm) frozen hydrated lamellae of PMCs were fabricated using cryo-focused ion beam-SEM. The lamellae were then loaded into a cryo-TEM, imaged and the ion composition of electron dense bodies was measured. Analyzes performed on 18 Ca-rich particles/particle clusters from 6 cells contained Ca, Na, Mg, S and P in different ratios. Surprisingly, all the Ca-rich particles contained P in amounts up to almost 1:1 of Ca. These cryo-STEM-EDS results were qualitatively confirmed by cryo-SEM-EDS analyzes of 310 vesicles, performed on high pressure frozen and cryo-planed samples. We discuss the advantages and limitations of the two techniques, and their potential applicability, especially to study ion transport pathways and ion trafficking in cells involved in mineralization. STATEMENT OF SIGNIFICANCE: The 'inorganic side of life', encompassing ion trafficking and ion storage in soft tissues of organisms, is a generally overlooked problem. Addressing such a problem becomes possible through the application of innovative techniques, performed in cryogenic conditions, which preserve the tissues in quasi-physiological state. We developed here a set of analytical tools, cryo-SEM-EDS, and cryo-STEM-EDS, which allow reconstructing the ion composition inside vesicles in sea urchin larval cells, on their way to deposit mineral in the skeletons. The techniques are complex, and we evaluate here the advantages and disadvantages of each technique. The methodologies that we are developing here can be applied to other cells and other pathways as well, eventually leading to quantitative elemental analyzes of tissues under cryogenic conditions.

Lively Minds: improving health and development through play-a randomised controlled trial evaluation of a comprehensive ECCE programme at scale in Ghana.

INTRODUCTION: Many children in developing countries grow up in environments that lack stimulation, leading to deficiencies in early years of development. Several efficacy trials of early childhood care and education (ECCE) programmes have demonstrated potential to improve child development; evidence on whether these effects can be sustained once programmes are scaled is much more mixed. This study evaluates whether an ECCE programme shown to be effective in an efficacy trial maintains effectiveness when taken to scale by the Government of Ghana (GoG). The findings will provide critical evidence to the GoG on effectiveness of a programme it is investing in, as well as a blueprint for design and scale-up of ECCE programmes in other developing countries, which are expanding their investment in ECCE programmes. METHODS AND ANALYSIS: This study is a cluster randomised controlled trial, in which the order that districts receive the programme is randomised. A minimum sample of 3240 children and 360 schools will be recruited across 72 district school cohort pairs. The primary outcomes are (1) child cognitive and socioemotional development measured using the International Development and Early Learning Assessment tool, the Strengths and Difficulties Questionnaire, and tasks from the Harvard Laboratory for Development Studies; (2) child health (measured using height/weight for age, height-for-weight Z scores). Secondary outcomes include (1) maternal mental health, (using Kessler-10 and Warwick Edinburgh Mental Wellbeing Scale) and knowledge of ECCE practices; (2) teacher knowledge, motivation and teaching quality (measured with classroom observation); (3) parental investment (using the Family Care Index and Home Observation Measurement of the Environment and the Child-Parent Relationship Scale); (4) water, sanitation, and hygiene (WASH) practices; (5) acute malnutrition (using mid-upper arm circumference). We will estimate unadjusted and adjusted intent-to-treat effects. ETHICS AND DISSEMINATION: Study protocols have been approved by ethics boards at the University College London (21361/001), Yale University (2000031549) and Ghanaian Health Service Ethics Review Committee (028/09/21). Results will be made available to participating communities, funders, the wider public and other researchers through peer-reviewed journals, conference presentations, social and print media and various community/stakeholder engagement activities. TRIAL REGISTRATION NUMBER: ISRCTN15360698, AEARCTR-0008500.

SARS-CoV-2 Production, Purification Methods and UV Inactivation for Proteomics and Structural Studies.

Severe acute respiratory syndrome coronavirus-2 is the causative agent of COVID-19. During the pandemic of 2019-2022, at least 500 million have been infected and over 6.3 million people have died from COVID-19. The virus is pleomorphic, and due to its pathogenicity is often handled in very restrictive biosafety containments laboratories. We developed two effective and rapid purification methods followed by UV inactivation that allow easy downstream handling of the virus. We monitored the purification through titering, sequencing, mass spectrometry and electron cryogenic microscopy. Although pelleting through a sucrose cushion, followed by gentle resuspension overnight gave the best particle recovery, infectivity decreased, and the purity was significantly worse than if using the size exclusion resin Capto Core. Capto Core can be used in batch mode, and was seven times faster than the pelleting method, obviating the need for ultracentrifugation in the containment laboratory, but resulting in a dilute virus. UV inactivation was readily optimized to allow handling of the inactivated samples under standard operating conditions. When containment laboratory space is limited, we recommend the use of Capto Core for purification and UV for inactivation as a simple, rapid workflow prior, for instance, to electron cryogenic microscopy or cell activation experiments.

Executive functions, motivation, and children's academic development in Côte d'Ivoire.

The role of executive function skills and motivation in supporting children's academic achievement is well-documented, but the vast majority of evidence is from high-income countries. Classrooms in sub-Saharan Africa tend to be large, teacher-driven, and lecture-focused, which may provide extra challenges for children to stay engaged in the learning process. Based in self-regulated learning theory, we tested the contributions of executive functions and motivation for children's literacy and numeracy skills over 1 school year. Our preregistered study of 2,500 primary school students in Côte d'Ivoire used lagged models with a robust set of demographic covariates. Executive functions were directly assessed, and children reported on their levels of motivation for schooling. Findings indicated strong longitudinal continuity of both literacy and numeracy skills over the academic year. Further, we found unique associations of executive functions and intrinsic motivation with changes in children's literacy skills and a unique association of executive functions, but not intrinsic motivation, with changes in children's numeracy skills over the school year. Overall, these results provide evidence for the importance of nonacademic skills for children's learning in a rural, sub-Saharan African context. Implications include improving access to preprimary education, the quality of primary school experiences, and teacher training and supports to increase children's early academic skills, self-regulation skills, and motivation for learning. (PsycInfo Database Record (c) 2022 APA, all rights reserved).

Electron microscopy of cellular ultrastructure in three dimensions.

Electron microscopy in three dimensions (3D) of cells and tissues can be essential for understanding the ultrastructural aspects of biological processes. The quest for 3D information reveals challenges at many stages of the workflow, from sample preparation, to imaging, data analysis and segmentation. Here, we outline several available methods, including volume SEM imaging, cryo-TEM and cryo-STEM tomography, each one occupying a different domain in the basic tradeoff between field-of-view and resolution. We discuss the considerations for choosing a suitable method depending on research needs and highlight recent developments that are essential for making 3D volume imaging of cells and tissues a standard tool for cellular and structural biologists.

Medium-term protective effects of quality early childhood education during the COVID-19 pandemic in Ghana.

The COVID-19 pandemic led to extended school closures globally. Access to remote learning opportunities during this time was vastly unequal within and across countries. Higher-quality early childhood education (ECE) can improve later academic outcomes, but longer-term effects during crises are unknown. This study provides the first experimental evidence of how previously attending a higher-quality ECE program affected child engagement in remote learning and academic scores during pandemic-related school closures in Ghana. Children (N = 1668; 50.1% male; Mage  = 10.1 years; all Ghanaian nationals) who attended higher-quality ECE at age 4 or 5 years had greater engagement in remote learning (d = .14) in October 2020, but not better language and literacy and math scores. Previous exposure to higher-quality ECE may support educational engagement during crises.

The roles of intracellular and extracellular calcium in Bacillus subtilis biofilms.

In nature, bacteria reside in biofilms- multicellular differentiated communities held together by an extracellular matrix. This work identified a novel subpopulation-mineral-forming cells-that is essential for biofilm formation in Bacillus subtilis biofilms. This subpopulation contains an intracellular calcium-accumulating niche, in which the formation of a calcium carbonate mineral is initiated. As the biofilm colony develops, this mineral grows in a controlled manner, forming a functional macrostructure that serves the entire community. Consistently, biofilm development is prevented by the inhibition of calcium uptake. Our results provide a clear demonstration of the orchestrated production of calcite exoskeleton, critical to morphogenesis in simple prokaryotes.