The incidence of depression and anxiety disorders in spontaneous chronic urticaria patients.

Objective: A frequent condition known as chronic urticaria (CU) is characterized by the appearance of wheals, angioedema, or both. CU lowers the quality of life and may also result in psychological discomfort. The literature survey revealed few studies dealing with depression and anxiety in these patients. Hence, Hamilton scores for depression and anxiety were used in this study to evaluate the incidence of depression and anxiety in chronic urticaria patients. Methodology: To evaluate CU patients' levels of depression and anxiety, the Hamilton Rating Scale for Depression (HDRS) and the Hamilton Anxiety Rating Scale (HAMA) were applied. Moreover, in a control group of thirty healthy volunteers, thirty CU patients were included in this study. It was essential to observe the patients' urticaria activity score, medications, age, gender, comorbidities, employment status, and income. When it came to levels of depression and anxiety, a comparison was made between the case group and the healthy group. Results: In the CU patients' group, the mean age was 26.9 years. The questionnaires showed that 14 (46%) subjects in the patient group had moderate to severe signs of anxiety, and 21 (70%) had moderate to severe symptoms of depression. Besides, in the control group,7 (23.3%) had moderate to severe signs of anxiety, and 8 (26.7%) had severe depression. Conclusion: According to the study, individuals with CU exhibit depression and anxiety symptoms more frequently than the control group. Therefore, the possibility of mental comorbidities should be considered when treating individuals with CU.

Drivers of vaccine hesitancy among vulnerable populations in India: a cross-sectional multi-state study.

Objectives: India's Covid-19 vaccination campaign engaged frontline workers (FLWs) to encourage vaccination among vulnerable segments of society. The FLWs report encountering a variety of barriers to vaccination and are often unsuccessful despite multiple visits to the same person. This cross-sectional study aims to pinpoint which of these barriers drive vaccine hesitancy among these segments, to help streamline vaccine communication, including FLW training, to better safeguard the population. Methods: Trained field enumerators contacted 893 individuals from five states across India and collected self-reported assessments of fifteen vaccination barriers (identified through discussions with FLWs), current vaccination status and future vaccination intentions, and covariates (demographics/comorbidities). Factor analysis of the fifteen barriers yielded two factors, one relating to fear of vaccine adverse effects and a second focused on peripheral concerns regarding the vaccine. The covariates significantly associated with current vaccination status were combined under a latent class regime to yield three cluster types (health access, financial strength, and demographics). The primary analysis examined the effect of the two barrier factors, the covariate clusters, and comorbidity, on current vaccination status and future vaccine intentions. Results: Fear of vaccine adverse effects was the primary driver of vaccine hesitancy; peripheral concerns frequently mentioned by the FLWs had no impact. Although cluster membership and the presence of comorbidities predicted vaccine uptake, neither of them materially altered the effect of fear of vaccine adverse effects with the following exception: fear of adverse effects was not associated with vaccination status among young Muslim men. Conclusion: Subject to limitations, these results indicate that interventions to decrease vaccine hesitancy should focus primarily on fear associated with vaccines rather than spend resources trying to address peripheral concerns.

KAT6A mutations in Arboleda-Tham syndrome drive epigenetic regulation of posterior HOXC cluster.

Arboleda-Tham Syndrome (ARTHS) is a rare genetic disorder caused by heterozygous, de novo mutations in Lysine(K) acetyltransferase 6A (KAT6A). ARTHS is clinically heterogeneous and characterized by several common features, including intellectual disability, developmental and speech delay, and hypotonia, and affects multiple organ systems. KAT6A is the enzymatic core of a histone-acetylation protein complex; however, the direct histone targets and gene regulatory effects remain unknown. In this study, we use ARTHS patient (n = 8) and control (n = 14) dermal fibroblasts and perform comprehensive profiling of the epigenome and transcriptome caused by KAT6A mutations. We identified differential chromatin accessibility within the promoter or gene body of 23% (14/60) of genes that were differentially expressed between ARTHS and controls. Within fibroblasts, we show a distinct set of genes from the posterior HOXC gene cluster (HOXC10, HOXC11, HOXC-AS3, HOXC-AS2, and HOTAIR) that are overexpressed in ARTHS and are transcription factors critical for early development body segment patterning. The genomic loci harboring HOXC genes are epigenetically regulated with increased chromatin accessibility, high levels of H3K23ac, and increased gene-body DNA methylation compared to controls, all of which are consistent with transcriptomic overexpression. Finally, we used unbiased proteomic mass spectrometry and identified two new histone post-translational modifications (PTMs) that are disrupted in ARTHS: H2A and H3K56 acetylation. Our multi-omics assays have identified novel histone and gene regulatory roles of KAT6A in a large group of ARTHS patients harboring diverse pathogenic mutations. This work provides insight into the role of KAT6A on the epigenomic regulation in somatic cell types.

Green synthesis of silver nanoparticles using Ocimum sanctum Linn. and its antibacterial activity against multidrug resistant Acinetobacter baumannii.

The biosynthesis of nanoparticles using the green route is an effective strategy in nanotechnology that provides a cost-effective and environmentally friendly alternative to physical and chemical methods. This study aims to prepare an aqueous extract of Ocimum sanctum (O. sanctum)-based silver nanoparticles (AgNPs) through the green route and test their antibacterial activity. The biosynthesized silver nanoparticles were characterised by colour change, UV spectrometric analysis, FTIR, and particle shape and size morphology by SEM and TEM images. The nanoparticles are almost spherical to oval or rod-shaped with smooth surfaces and have a mean particle size in the range of 55 nm with a zeta potential of -2.7 mV. The antibacterial activities of AgNPs evaluated against clinically isolated multidrug-resistant Acinetobacter baumannii (A. baumannii) showed that the AgNPs from O. sanctum are effective in inhibiting A. baumannii growth with a zone of inhibition of 15 mm in the agar well diffusion method and MIC and MBC of 32 µg/mL and 64 µg/mL, respectively. The SEM images of A. baumannii treated with AgNPs revealed damage and rupture in bacterial cells. The time-killing assay by spectrophotometry revealed the time- and dose-dependent killing action of AgNPs against A. baumannii, and the assay at various concentrations and time intervals indicated a statistically significant result in comparison with the positive control colistin at 2 µg/mL (P < 0.05). The cytotoxicity test using the MTT assay protocol showed that prepared nanoparticles of O. sanctum are less toxic against human cell A549. This study opens up a ray of hope to explore the further research in this area and to improve the antimicrobial activities against multidrug resistant bacteria.

KAT6A mutations in Arboleda-Tham syndrome drive epigenetic regulation of posterior HOXC cluster.

Arboleda-Tham Syndrome (ARTHS) is a rare genetic disorder caused by heterozygous, de novo truncating mutations in Lysine(K) acetyltransferase 6A (KAT6A). ARTHS is clinically heterogeneous and characterized by several common features including intellectual disability, developmental and speech delay, hypotonia and affects multiple organ systems. KAT6A is highly expressed in early development and plays a key role in cell-type specific differentiation. KAT6A is the enzymatic core of a histone-acetylation protein complex, however the direct histone targets and gene regulatory effects remain unknown. In this study, we use ARTHS patient (n=8) and control (n=14) dermal fibroblasts and perform comprehensive profiling of the epigenome and transcriptome caused by KAT6A mutations. We identified differential chromatin accessibility within the promoter or gene body of 23%(14/60) of genes that were differentially expressed between ARTHS and controls. Within fibroblasts, we show a distinct set of genes from the posterior HOXC gene cluster (HOXC10, HOXC11, HOXC-AS3, HOXC-AS2, HOTAIR) that are overexpressed in ARTHS and are transcription factors critical for early development body segment patterning. The genomic loci harboring HOXC genes are epigenetically regulated with increased chromatin accessibility, high levels of H3K23ac, and increased gene-body DNA methylation compared to controls, all of which are consistent with transcriptomic overexpression. Finally, we used unbiased proteomic mass spectrometry and identified two new histone post-translational modifications (PTMs) that are disrupted in ARTHS: H2A and H3K56 acetylation. Our multi-omics assays have identified novel histone and gene regulatory roles of KAT6A in a large group of ARTHS patients harboring diverse pathogenic mutations. This work provides insight into the role of KAT6A on the epigenomic regulation in somatic cell types.

Multiomics of Bohring-Opitz syndrome truncating ASXL1 mutations identify canonical and noncanonical Wnt signaling dysregulation.

ASXL1 (additional sex combs-like 1) plays key roles in epigenetic regulation of early developmental gene expression. De novo protein-truncating mutations in ASXL1 cause Bohring-Opitz syndrome (BOS; OMIM #605039), a rare neurodevelopmental condition characterized by severe intellectual disabilities, distinctive facial features, hypertrichosis, increased risk of Wilms tumor, and variable congenital anomalies, including heart defects and severe skeletal defects giving rise to a typical BOS posture. These BOS-causing ASXL1 variants are also high-prevalence somatic driver mutations in acute myeloid leukemia. We used primary cells from individuals with BOS (n = 18) and controls (n = 49) to dissect gene regulatory changes caused by ASXL1 mutations using comprehensive multiomics assays for chromatin accessibility (ATAC-seq), DNA methylation, histone methylation binding, and transcriptome in peripheral blood and skin fibroblasts. Our data show that regardless of cell type, ASXL1 mutations drive strong cross-tissue effects that disrupt multiple layers of the epigenome. The data showed a broad activation of canonical Wnt signaling at the transcriptional and protein levels and upregulation of VANGL2, which encodes a planar cell polarity pathway protein that acts through noncanonical Wnt signaling to direct tissue patterning and cell migration. This multiomics approach identifies the core impact of ASXL1 mutations and therapeutic targets for BOS and myeloid leukemias.

Going Virtual: Adapting an Institutional Annual Bereavement Event During the COVID-19 Pandemic.

Our institution's annual bereaved family event was modified to a virtual format during the COVID-19 pandemic. While necessary to comply with physical distancing directives, the transition also provided greater accessibility for families. Virtual events were feasible and appreciated by attendees. Future hybrid bereavement events should be considered to allow families more flexibility and accessibility.

Anemia in Indian Men: An Emerging Public Health Challenge.

Approximately one-quarter of the world's population is suffering from anemia, out of which 12.7% of men suffer from anemia around the globe. In India, anemia is a moderate public health problem (ranging from 20 to 39.9%). Prevalence of anemia is 25% in men and 31% in adolescent boys, in India. In recent years, there has been an alarming rise in anemia prevalence among adolescent boys (15-19 years) in 23 states, and among men (15-49 years) in 17 states. Anemia in the states of Jammu and Kashmir and Ladakh is a severe public health problem. Despite being a vital issue, indicators for anemia in men are not given due importance in national health statistics reports. This article highlights the issue to prioritize men's health in the context of anemia at state and national levels.

Recent updates on nano-phyto-formulations based therapeutic intervention for cancer treatment.

Cancer is a leading cause of death globally, with limited treatment options and several limitations. Chemotherapeutic agents often result in toxicity which long-term conventional treatment. Phytochemicals are natural constituents that are more effective in treating various diseases with less toxicity than the chemotherapeutic agents providing alternative therapeutic approaches to minimize the resistance. These phytoconstituents act in several ways and deliver optimum effectiveness against cancer. Nevertheless, the effectiveness of phyto-formulations in the management of cancers may be constrained due to challenges related to inadequate solubility, bioavailability, and stability. Nanotechnology presents a promising avenue for transforming current cancer treatment methods through the incorporation of phytochemicals into nanosystems, which possess a range of advantageous characteristics such as biocompatibility, targeted and sustained release capabilities, and enhanced protective effects. This holds significant potential for future advancements in cancer management. Herein, this review aims to provide intensive literature on diverse nanocarriers, highlighting their applications as cargos for phytocompounds in cancer. Moreover, it offers an overview of the current advancements in the respective field, emphasizing the characteristics that contribute to favourable outcomes in both in vitro and in vivo settings. Lastly, clinical development and regulatory concerns are also discussed to check on the transformation of the concept as a promising strategy for combination therapy of phytochemicals and chemotherapeutics that could lead to cancer management in the future.

Caregivers' Preferences of COVID-19 Vaccination for Children: A Cross-sectional Study From Rural South India.

Objective: This study aimed to document caregivers' perceptions and preferences regarding coronavirus disease-19 (COVID-19) vaccination among children. Methods: This cross-sectional study analyzed 272 caregivers with 347 children (aged 1-18 years) attending a subdistrict rural hospital in February-March 2022. Results: Vaccine acceptance was high (93.4%). Although fear of side effects was the most common reason not to vaccinate, a higher proportion of caregivers willing to vaccinate children had consulted healthcare personnel to clarify queries related to side effects. Familiar vaccination sites, where children had previously received routine immunization (RI), such as government hospitals, and Anganwadis (community-based childcare centers) where vaccines were available free of cost on all working days, were the most preferred for COVID-19 vaccination, followed by schools. Only 5.5% of the caregivers preferred private hospitals. Vaccination at home was desired for chronically ill and out-of-school children. RI as per age was associated with the willingness to vaccinate. In addition to protection from COVID-19, other benefits identified by willing parents were being able to attend schools, recreation, and travel. Conclusions: Out-of-school children, children left or missed out in RI, and children with chronic illness can be at risk of being left out for COVID-19 vaccination and can be included by expanding vaccination services house-to-house as in adults. Media engagement and communication must be interactive to address issues, such as fear of side effects, and promote additional benefits of vaccination.

A Participatory Design Approach to Develop Visualization of Wearable Actigraphy Data for Health Care Professionals: Case Study in Qatar.

BACKGROUND: Several tools have been developed for health care professionals to monitor the physical activity of their patients, but most of these tools have been considering only the needs of users in North American and European countries and applicable for only specific analytic tasks. To our knowledge, no research study has utilized the participatory design (PD) approach in the Middle East region to develop such tools, involving all the stakeholders in the product development phases, and no clear use cases have been derived from such studies that could serve future development in the field. OBJECTIVE: This study aims to develop an interactive visualization tool (ActiVis) to support local health care professionals in monitoring the physical activity of their patients measured through wearable sensors, with the overall objective of improving the health of the Qatari population. METHODS: We used PD and user-centered design methodologies to develop ActiVis, including persona development, brainwriting, and heuristic walkthrough as part of user evaluation workshops; and use cases, heuristic walkthrough, interface walkthrough, and survey as part of expert evaluation sessions. RESULTS: We derived and validated 6 data analysis use cases targeted at specific health care professionals from a collaborative design workshop and an expert user study. These use cases led to improving the design of the ActiVis tool to support the monitoring of patients' physical activity by nurses and family doctors. The ActiVis research prototype (RP) compared favorably with the Fitbit Dashboard, showing the importance of design tools specific to end users' needs rather than relying on repurposing existing tools designed for other types of users. The use cases we derived happen to be culturally agnostic, despite our assumption that the local Muslim and Arabic culture could impact the design of such visualization tools. At last, taking a step back, we reflect on running collaborative design sessions in a multicultural environment and oil-based economy. CONCLUSIONS: Beyond the development of the ActiVis tool, this study can serve other visualization and human-computer interaction designers in the region to prepare their design projects and encourage health care professionals to engage with designers and engineers to improve the tools they use for supporting their daily routine. The development of the ActiVis tool for nurses, and other visualization tools specific to family doctors and clinician researchers, is still ongoing and we plan to integrate them into an operational platform for health care professionals in Qatar in the near future.

Trends in college student mental health and help-seeking by race/ethnicity: Findings from the national healthy minds study, 2013-2021.

BACKGROUND: A considerable gap in knowledge exists around mental health trends in diverse racial and ethnic adolescent and young adult populations. The purpose of this study is to examine annual trends for mental health and help-seeking by race/ethnicity in a national sample of college students. METHODS: Survey data come from >350,000 students at 373 campuses that participated in the Healthy Minds Study between 2013 and 2021. Analyses are descriptive in nature focusing on year-by-year prevalence and help-seeking rates for each racial/ethnic group. RESULTS: In 2020-2021, >60% of students met criteria for one or more mental health problems, a nearly 50% increase from 2013. Mental health worsened among all groups over the study period. American Indian/Alaskan Native students experienced the largest increases in depression, anxiety, suicidal ideation, and meeting criteria for one or more mental health problem. Students of color had the lowest rates of mental health service utilization. The highest annual rate of past-year treatment for Asian, Black, and Latinx students was at or below the lowest rate for White students. Although Arab American students experienced a 22% increase in prevalence, there was an 18% decrease in treatment. LIMITATIONS: Response rates raise the potential of nonresponse bias. Sample weights adjust along known characteristics, but there may be differences on unobserved characteristics. CONCLUSIONS: Findings have important implications for campus mental health programming and underscore the urgency of reducing mental health inequalities in college student populations through the identification and implementation of best practices both in clinical settings and through system-level change.

Novel variants in KAT6B spectrum of disorders expand our knowledge of clinical manifestations and molecular mechanisms.

The phenotypic variability associated with pathogenic variants in Lysine Acetyltransferase 6B (KAT6B, a.k.a. MORF, MYST4) results in several interrelated syndromes including Say-Barber-Biesecker-Young-Simpson Syndrome and Genitopatellar Syndrome. Here we present 20 new cases representing 10 novel KAT6B variants. These patients exhibit a range of clinical phenotypes including intellectual disability, mobility and language difficulties, craniofacial dysmorphology, and skeletal anomalies. Given the range of features previously described for KAT6B-related syndromes, we have identified additional phenotypes including concern for keratoconus, sensitivity to light or noise, recurring infections, and fractures in greater numbers than previously reported. We surveyed clinicians to qualitatively assess the ways families engage with genetic counselors upon diagnosis. We found that 56% (10/18) of individuals receive diagnoses before the age of 2 years (median age = 1.96 years), making it challenging to address future complications with limited accessible information and vast phenotypic severity. We used CRISPR to introduce truncating variants into the KAT6B gene in model cell lines and performed chromatin accessibility and transcriptome sequencing to identify key dysregulated pathways. This study expands the clinical spectrum and addresses the challenges to management and genetic counseling for patients with KAT6B-related disorders.

Plasmodium's journey through the Anopheles mosquito: A comprehensive review.

The malaria parasite has an extraordinary ability to evade the immune system due to which the development of a malaria vaccine is a challenging task. Extensive research on malarial infection in the human host particularly during the liver stage has resulted in the discovery of potential candidate vaccines including RTS,S/AS01 and R21. However, complete elimination of malaria would require a holistic multi-component approach. In line with this, under the World Health Organization's PATH Malaria Vaccine Initiative (MVI), the research focus has shifted towards the sexual stages of malaria in the mosquito host. Last two decades of scientific research obtained seminal information regarding the sexual/mosquito stages of the malaria. This updated and comprehensive review would provide the basis for consolidated understanding of cellular, biochemical, molecular and immunological aspects of parasite transmission right from the sexual stage commitment in the human host to the sporozoite delivery back into subsequent vertebrate host by the female Anopheles mosquito.

Maternal vitamin B12 deficiency in rats alters DNA methylation in metabolically important genes in their offspring.

Vitamin B12 deficiency is a critical problem worldwide and peri-conceptional deficiency of this vitamin is associated with the risk of complex cardio-metabolic diseases. Nutritional perturbations during these stages of development may lead to changes in the fetal epigenome. Using Wistar rat model system, we have earlier shown that low maternal B12 levels are associated with low birth weight, adiposity, insulin resistance, and increased triglyceride levels in the offspring, which might predispose them to the risk of cardio-metabolic diseases in adulthood. In this study, we have investigated the effects of maternal B12 deficiency on genome-wide DNA methylation profile of the offspring and the effect of rehabilitation of mothers with B12 at conception. We have performed methylated DNA immunoprecipitation sequencing of liver from pups in four groups of Wistar rats: Control (C), B12-restricted (B12R), B12-rehabilitated at conception (B12RC), and B12-rehabilitated at parturition (B12RP). We have analyzed differentially methylated signatures between the three groups as compared to controls. We have identified a total of 214 hypermethylated and 142 hypomethylated regions in the 10 kb upstream region of transcription start site in pups of B12-deficient mothers, which are enriched in genes involved in fatty acid metabolism and mitochondrial transport/metabolism. B12 rehabilitation at conception and parturition is responsible for reversal of methylation status of many of these regions to control levels suggesting a causal association with metabolic phenotypes. Thus, maternal B12 restriction alters DNA methylation of genes involved in important metabolic processes and influences the offspring phenotype, which is reversed by B12 rehabilitation of mothers at conception.

Benchmark on a large cohort for sleep-wake classification with machine learning techniques.

Accurately measuring sleep and its quality with polysomnography (PSG) is an expensive task. Actigraphy, an alternative, has been proven cheap and relatively accurate. However, the largest experiments conducted to date, have had only hundreds of participants. In this work, we processed the data of the recently published Multi-Ethnic Study of Atherosclerosis (MESA) Sleep study to have both PSG and actigraphy data synchronized. We propose the adoption of this publicly available large dataset, which is at least one order of magnitude larger than any other dataset, to systematically compare existing methods for the detection of sleep-wake stages, thus fostering the creation of new algorithms. We also implemented and compared state-of-the-art methods to score sleep-wake stages, which range from the widely used traditional algorithms to recent machine learning approaches. We identified among the traditional algorithms, two approaches that perform better than the algorithm implemented by the actigraphy device used in the MESA Sleep experiments. The performance, in regards to accuracy and F 1 score of the machine learning algorithms, was also superior to the device's native algorithm and comparable to human annotation. Future research in developing new sleep-wake scoring algorithms, in particular, machine learning approaches, will be highly facilitated by the cohort used here. We exemplify this potential by showing that two particular deep-learning architectures, CNN and LSTM, among the many recently created, can achieve accuracy scores significantly higher than other methods for the same tasks.

Implementing 360° Quantified Self for childhood obesity: feasibility study and experiences from a weight loss camp in Qatar.

BACKGROUND: The explosion of consumer electronics and social media are facilitating the rise of the Quantified Self (QS) movement where millions of users are tracking various aspects of their daily life using social media, mobile technology, and wearable devices. Data from mobile phones, wearables and social media can facilitate a better understanding of the health behaviors of individuals. At the same time, there is an unprecedented increase in childhood obesity rates worldwide. This is a cause for grave concern due to its potential long-term health consequences (e.g., diabetes or cardiovascular diseases). Childhood obesity is highly prevalent in Qatar and the Gulf Region. In this study we examine the feasibility of capturing quantified-self data from social media, wearables and mobiles within a weight lost camp for overweight children in Qatar. METHODS: Over 50 children (9-12 years old) and parents used a wide range of technologies, including wearable sensors (actigraphy), mobile and social media (WhatsApp and Instagram) to collect data related to physical activity and food, that was then integrated with physiological data to gain insights about their health habits. In this paper, we report about the acquired data and visualization techniques following the 360° Quantified Self (360QS) methodology (Haddadi et al., ICHI 587-92, 2015). RESULTS: 360QS allows for capturing insights on the behavioral patterns of children and serves as a mechanism to reinforce education of their mothers via social media. We also identified human factors, such as gender and cultural acceptability aspects that can affect the implementation of this technology beyond a feasibility study. Furthermore, technical challenges regarding the visualization and integration of heterogeneous and sparse data sets are described in the paper. CONCLUSIONS: We proved the feasibility of using 360QS in childhood obesity through this pilot study. However, in order to fully implement the 360QS technology careful planning and integration in the health professionals' workflow is needed. TRIAL REGISTRATION: The trial where this study took place is registered at ClinicalTrials.gov on 14 November 2016 ( NCT02972164 ).

Comparative whole-genome analysis of clinical isolates reveals characteristic architecture of Mycobacterium tuberculosis pangenome.

The tubercle complex consists of closely related mycobacterium species which appear to be variants of a single species. Comparative genome analysis of different strains could provide useful clues and insights into the genetic diversity of the species. We integrated genome assemblies of 96 strains from Mycobacterium tuberculosis complex (MTBC), which included 8 Indian clinical isolates sequenced and assembled in this study, to understand its pangenome architecture. We predicted genes for all the 96 strains and clustered their respective CDSs into homologous gene clusters (HGCs) to reveal a hard-core, soft-core and accessory genome component of MTBC. The hard-core (HGCs shared amongst 100% of the strains) was comprised of 2,066 gene clusters whereas the soft-core (HGCs shared amongst at least 95% of the strains) comprised of 3,374 gene clusters. The change in the core and accessory genome components when observed as a function of their size revealed that MTBC has an open pangenome. We identified 74 HGCs that were absent from reference strains H37Rv and H37Ra but were present in most of clinical isolates. We report PCR validation on 9 candidate genes depicting 7 genes completely absent from H37Rv and H37Ra whereas 2 genes shared partial homology with them accounting to probable insertion and deletion events. The pangenome approach is a promising tool for studying strain specific genetic differences occurring within species. We also suggest that since selecting appropriate target genes for typing purposes requires the expected target gene be present in all isolates being typed, therefore estimating the core-component of the species becomes a subject of prime importance.