BACKGROUND:Hepatitis B virus (HBV) infections are a critical public health issue, especially in lower resource setting where the potential for chronic infections is increased. In Nepal an HBV vaccination program has been in place since the early 2000s, though knowledge and awareness of HBV has been recorded as being low within community settings. This study explored inequalities which may influence the overall healthcare system within Nepal regarding HBV, specifically vaccination programs, knowledge, awareness, and stigma associated with HBV. METHODS:In-depth interviews (n = 21) were conducted with healthcare professionals from different healthcare districts in Nepal. These interviews were then transcribed before thematic analysis was performed. RESULTS:Several potential sources of inequality of service provision were identified. Awareness of HBV was reasonable among the participants; however, level of knowledge was variable. Participants felt while the vaccination program had been somewhat successful, there were still several areas in the Nepalese health system which needed to be strengthened to further mitigate risk of HBV infections. This included the need to strengthen and standardise vaccination processes within Nepal. CONCLUSION:Based on our findings, efforts to provide appropriate resources and equipment to community health centres needed to be reinforced. An increase is also needed in education and awareness programs around health issues, specifically those involving infectious diseases such as Hepatitis B at the community level. There also should be continued commitment to enhancing the healthcare services provided in rural and other underserved areas of Nepal.

Globally, recent increases in skilled birth attendant (SBA) coverage do not appear to be translating to proportionate reductions in maternal and neonatal mortality. One hypothesized driver is a quality gap in the childbirth care provided by health professionals. This research was conducted as part of a multi-country study to assess the quality of pre-service education of health professionals providing childbirth care and to enhance the measurement related to the coverage of SBAs and SHP. This work introduces a new metric of SBA quality, uses that metric to identify tracer indicators of quality that could be integrated into routine monitoring without the use of complex statistical methods, and identifies priorities for improving quality of care in Nepal, Senegal and Zambia. We gathered primary and secondary data, aligned to a comprehensive conceptual framework about quality of maternal and newborn care, in three “Exemplar” countries (Nepal, Senegal and Zambia). We applied latent variable analysis principles to measure quality, and constructed estimates of effective coverage of SBA. The five highest weighted indicators most predictive of quality of care were presence of a national compulsory continued development system, presence of a national licensing, relicensing and registration system, presence of a national education regulation system, proportion of health facilities with electricity available at all times, and proportion of health facilities with amoxicillin suspension available. Validity tests showed that effective SBA coverage had a stronger relationship with early neonatal mortality rates than crude SBA coverage at the subnational level. Our analysis suggests that effective SBA coverage is substantially lower than crude SBA coverage, ranging from 18% to 30% in Senegal, 25% to 43% in Nepal, and 12% to 35% in Zambia. Lower effective coverage levels demonstrated here reveal that mothers and newborns are receiving far less of the essential care than official statistics may portray. This work presents a novel approach to measure one of the most challenging concepts in global health, and the highest weighted indicators in this analysis may be useful for routine monitoring in LMICs and at the global level. These results also add to the evidence supporting a basic policy package to improve the quality of maternal and newborn care, including a national compulsory continued education system, national licensing, relicensing and registration systems and national education regulation systems. Finally, programs in these countries may benefit from using the subnational estimates to geographically tailor interventions and to improve health outcomes.

Objectives Community health workers are essential to front-line health outreach throughout low-income and middle-income countries, including programming for early childhood immunisation. Understanding how community health workers are engaged for successful early childhood vaccination among countries who showed success in immunisation coverage would support evidence-based policy guidance across contexts.Design We employed a multiple case study design using qualitative research methods.Setting We conducted research in Nepal, Senegal and Zambia.Participants We conducted 207 interviews and 71 focus group discussions with 678 participants at the national, regional, district, health facility and community levels of the health systems of Nepal, Senegal and Zambia, from October 2019 to April 2021. We used thematic analysis to investigate contributing factors of community health worker programming that supported early childhood immunisation within each country and across contexts.Results Implementation of vaccination programming relied principally on the (1) organisation, (2) motivation and (3) trust of community health workers. Organisation was accomplished by expanding cadres of community health workers to carry out their roles and responsibilities related to vaccination. Motivation was supported by intrinsic and extrinsic incentives. Trust was expressed by communities due to community health worker respect and value placed on their work.Conclusion Improvements in immunisation coverage was facilitated by community health worker organisation, motivation and trust. With the continued projection of health worker shortages, especially in low-income countries, community health workers bridged the equity gap in access to vaccination services by enabling wider reach to underserved populations. Although improvements in vaccination programming were seen in all three countries—including government commitment to addressing human resource deficits, training and remuneration; workload, inconsistency in compensation, training duration and scope, and supervision remain major challenges to immunisation programming. Health decision-makers should consider organisation, motivation and trust of community health workers to improve the implementation of immunisation programming.

Respiratory burst oxidase homolog (RBOH) proteins in plants generate reactive oxygen species (ROS) in the apoplast to regulate developmental processes and stress responses. Herein, a total of 40 TaRBOH genes are identified in the genome of Triticum aestivum by a genome-wide search using the latest database. Phylogenetic analysis separated the RBOH proteins into five clusters and close clustering suggested an evolutionary relationship among them. The presence of duplication events (DEs) and the nature of selection (purifying) during evolutionary analysis revealed their role in the expansion of the TaRBOH gene family. The interaction analyses revealed their extended roles and coordinated functioning with various stress-related signaling pathways, including ABA- and Ca2+-signaling. Expression profiling in different tissue developmental stages and under stress conditions disclosed their involvement in growth, development and stress responses. In addition, the presence of assorted groups of cis-regulatory elements, interaction with the diverse nature of transcription factors and miRNA related to plant development, hormones and various stresses further suggested their association with developmental and stress-responsive pathways. This study provides inclusive information related to the functioning of TaRBOH genes in bread wheat and would provide a valuable reference for their functional characterization for crop improvement.

Vascular plant one-zinc-finger (VOZ) transcription factors play indispensable roles in orchestrating growth and stress responses in plants. These plant-specific regulators are integral to important biological processes such as flowering. Despite being a vital regulator of numerous biological processes, significant information about their role is not available. Herein, we identified and characterized six TaVOZ genes in T. aestivum by genome-wide investigation, which are phylogenetically clustered into two groups. They consist of conserved gene and protein architecture with two important domains (A and B). The occurrence of assorted cis-regulatory elements related to growth and stress response and their interaction with various transcription factors revealed the possible involvement of TaVOZ genes in numerous biological processes. Tissue-specific differential expression of a few TaVOZ genes divulged their role during plant developmental processes. Additionally, the modulated expression of TaVOZ2-3B gene during heat, drought, salinity and fungal infestations unveiled their involvement in stress response. The interaction analyses identified other vital components in the intricate network of TaVOZ-mediated stress signaling. This study provides inclusive knowledge and significance of TaVOZ genes for future crop improvement programs.

Pre-mRNA processing factor 4 kinases (PRP4Ks) are crucial for eukaryotic organisms. These proteins are dual specificity kinases that phosphorylate the proteins involved in spliceosomal assembly during pre-mRNA processing. A repertoire of six TaPRP4K genes with conserved architecture is found in the allohexaploid genome of Triticum aestivum. These genes are evenly distributed on chromosomes 4 and 5. The phylogenetic analysis reveals the PRP4Ks’ distinct groups of various monocots and dicots. The DMFA, EGY/DH-box and a few other motifs of the protein kinase domain are found conserved in all the TaPRP4K proteins. Additionally, an atypical protein kinase domain, ABC1 is also present, which increases the structural complexity of each TaPRP4Ks. The promoter sequence analysis reveals the occurrence of light, growth, hormone and stress-related cis-regulatory elements, which shows interaction with a diverse group of transcription factors (TFs). TaPRP4K genes are also presumed to be regulated by miRNAs associated with plant growth and stress response. In addition, TaPRP4Ks display interaction with numerous other proteins such as CLO, BRR2a, SMU1, STA1, etc., which are involved in the regulation of the splicing mechanism. The expression study reveals significant expression of certain TaPRP4K genes in above-ground tissues, which indicates their roles in vegetative and reproductive tissue development. Further, TaPRP4K2 group genes show upregulated expression during salt stress and fungus (Blumeria graminis f. sp. tritici) infestation, which suggests their role in both abiotic and biotic stress response. Our study provides numerous important features and probable functioning of TaPRP4K genes, which would be useful in the detailed functional characterization of these genes in future studies.

During millions of years of co-evolution, both plants and herbivores developed multiple strategies to counter each other. Plant defense responses against herbivores are a highly complex process and have at least two layers along with an outer structural defense barrier. Protein kinases are the central molecule in host defense response and are involved in multiple steps from perception to distal downstream signaling. In the current article, we explore all roles of these protein kinases of plants against herbivores and discuss their future prospective and bottleneck to achieve herbivore resistance.

Introduction The fundamental components of a vaccine delivery system are well-documented, but robust evidence is needed on how the related processes and implementation strategies - including the facilitators and barriers-contribute to improvements in childhood vaccination coverage. The purpose of this study was to identify critical facilitators and barriers to the implementation of common interventions across three countries that have dramatically increased coverage of early childhood vaccination over the past 20 years, and to qualify common or divergent themes in their success. Methods We conducted 277 key informant interviews and focus group discussions with public health leaders at the regional, district, and local levels and community members in Nepal, Senegal, and Zambia to identify intervention activities and the facilitators and barriers to implementation. We used thematic analysis grounded in the Consolidated Framework for Implementation Research (CFIR) to identify immunization program key facilitators and barriers. Results We found that the common facilitators to program implementation across the countries were the CFIR inner setting constructs of 1) networks and communications, 2) goals and feedback, 3) relative priority, and 4) readiness for implementation; and outer setting constructs of 4) cosmopolitanism and 5) external policies and mandates. The common barriers were incentives and rewards, available resources, access to knowledge and information, and patients needs and resources. Critical to the success of these national immunization programs were prioritization and codification of health as a human right, clear chain of command and shared ownership of immunization, communication of program goals and feedback, offering of incentives at multiple levels, training of staff central to vaccination education, the provision of resources to support the program, key partnerships and guidance on implementation and adoption of vaccination policies. Conclusion Adequate organizational commitment, resources, communication, training, and partnerships were the most critical facilitators for these countries to improve childhood vaccination.

ABSTRACT Background Hepatitis B virus (HBV) remains a public health issue in many developing nations, including Nepal. In Nepal the vaccination program was implemented in 2002-3 and aimed to reduce national prevalence of HBV. This study investigated socio-demographic, behavioural, and health service factors associated with HBV infection in low (Pokhara) and high (Dolpa) prevalence populations. Methods A serosurvey of 400 participants from within each population was conducted (final N = 799). The study combined a blood-draw to ascertain HBV status and a questionnaire that included demographic questions and items on knowledge of HBV, behavioural, and social factors associated with the risk of HBV infection. The blood draws were used to confirm HBV status as well as identify any previous infections from which participants may have recovered. Results There were 8/399 (2.01%, 95% CI 0.87%, 3.91%) current HBV infections among participants from Dolpa, compared with 2/400 (0.5%, 95% CI0.06%, 1.79%) among participants from Pokhara. The average age of each of the cohorts was relatively high, indicating an unclear association between vaccination programs and the low prevalence observed in this study. There was evidence from both sites there had been previous infections within the community as many of the participants indicated some level of protection from HBV either through vaccination or past infection (Dolpa 58/399, Pokhara 21/400) and recent cases who had recovered (Dolpa 4/399, Pokhara 3/400). Due to the relatively low prevalence of active cases of HBV, no meaningful associations between demographic, behavioural, and healthcare factors could be calculated. In both samples low level of knowledge of HBV and stigma towards people with HBV was evident. Conclusions This study found a low prevalence of HBV infection in both low and putatively high prevalence populations. However, knowledge of how HBV can be spread was quite low in each of the groups, indicating participants are managing to avoid infections while not necessarily being cautious around behaviours representing the greatest infection risk. There was also evidence of stigma being associated with having an active HBV infection, which may reduce the willingness of individuals to seek diagnosis or treatment.

BACKGROUND:Hepatitis B virus (HBV) is a major source of disease burden worldwide, with an estimated 296 million individuals living with infections worldwide. Although vaccine programs exist to control infections, certain sub-populations around the world continue to have very high prevalence of HBV infection. METHODS:A systematic search of studies of HBV published after 2010 was conducted for India, Pakistan, Bangladesh, Nepal, Sri Lanka and Bhutan. Each paper was independently screened for risk of bias and inclusion. Data were extracted from included studies before being analysed to estimate pooled prevalence, and to conduct sub-group analyses. Random-effects models were used for estimating summary prevalence due to a high level of heterogeneity between studies, and funnel plots were combined with Egger's test to assess publication bias. Meta-regression was conducted to investigate sources of between-study heterogeneity. RESULTS:The pooled prevalence of HBV across all studies was 3% (95% CI 0.02, 0.05). For countries with multiple studies, the pooled prevalence in India was 3% (95% CI 0.02, 0.04), in Pakistan 6% (95% CI 0.03, 0.09), in Bangladesh 5% (95% CI of 0.02, 0.12), and in Nepal 1% (95% CI 0.00, 0.08). There was some evidence of publication bias, and a high level of heterogeneity across studies. Risk of bias analysis found most studies to be of fair or moderate quality. CONCLUSIONS:The prevalence of HBV among countries in the sub-continent was higher than the global average, but was not as high as some other regions. Countries with greater numbers of displaced persons had higher prevalence of HBV, with a wide range of prevalence between subpopulations likely reflecting differential uptake, and implementation, of vaccination programs.

Phtheirospermum japonicum shows induced expression of PjPME and PjPMEI genes during haustoria development in rice and Arabidopsis with increased PME activity, which leads to the modulated cell wall during parasitism. Moreover, how PME and PMEI proteins interact and balance during haustoria development remains elusive.

Volatile organic compounds emanating from plant surfaces serve as a sustainable natural solution to combat biotic stresses in plants. Leaf methanol is the simplest and second major volatile organic compound after isoprene emitted through the leaf surface. Methanol has been neglected as a by-product of other secondary metabolites for a long time, but recent studies have suggested its importance in development and stress responses. In our previous findings, we had revealed that transgenic plants over-expressing PME, enhanced methanol production providing resistance against a broad spectrum of insects. In the current study, we extended our previous work to provide new insights by performing differential transcriptomics of high-methanol-producing insect-resistant transgenic lines. We found that 2262 genes were differentially expressed in the transgenics plants, including transcription factors, cell wall modulating, phytohormones signaling and development-related genes. Our results demonstrated that the expression levels of transcription factors associated with development and biotic stress were altered in the transgenic lines. In addition, phytohormones ABA and gibberellin signalling genes were upregulated, whereas ethylene and auxin signalling genes were downregulated. Moreover, biochemical characteristics of cell walls in both transgenic tobacco plants were comparable to the control plants.

Objectives Vaccination averts an estimated 2–3 million deaths annually. Although vaccine coverage improvements across Africa and South Asia have remained below global targets, several countries have outperformed their peers with significant increases in coverage. The objective of this study was to examine these countries’ vaccination programmes and to identify and describe critical success factors that may have supported these improvements.Design Multiple case study design using qualitative research methods.Setting Three countries with high routine immunisation rates: Nepal, Senegal, and Zambia.Participants We conducted 207 key informant interviews and 71 focus group discussions with a total of 678 participants. Participants were recruited from all levels, including government officials, health facility staff, frontline workers, community health workers, and parents. Participants were recruited from both urban and rural districts in Nepal, Senegal, and Zambia.Results Our data revealed that the critical success factors for vaccination programmes relied on the cultural, historical, and statutory context in which the interventions were delivered. In Nepal, Senegal, and Zambia, high immunisation coverage was driven by (1) strong governance structures and healthy policy environments; (2) adjacent successes in health system strengthening; (3) government-led community engagement initiatives, and (4) adaptation considering contextual factors at all levels of the health system.Conclusions Throughout this project, our analysis returned to the importance of defining and understanding the context, governance, financing, and health systems within a country, rather than focusing on any one intervention. This paper augments findings from existing literature by highlighting how contextual factors impact implementation decisions that have led to improvements in childhood vaccine delivery. Findings from this research may help identify transferable lessons and support actionable recommendations to improve national immunisation coverage in other settings.

Introduction: Childhood vaccination is an effective intervention for lowering the burden of infectious disease. Vaccine coverage has increased globally, but vaccine hesitancy and refusal threatens these gains. The 5C psychological antecedents of vaccination (“5C”) provides a validated measure of “vaccine hesitancy or confidence” to assess individual thoughts and behaviors behind vaccination. We investigated population-level factors that contributed to high and sustained vaccination coverage in Zambia, Nepal, and Senegal, and alignment with the 5Cs. Methods: Data was collected in the larger Exemplars in Vaccine Delivery study, from focus group discussions (FDGs) and key informant interviews (KIIs) at the national, regional, district, health facility, and community levels of health systems in Zambia, Nepal, and Senegal. We assessed the demand environment, as relayed by participants, and identified interventions reported as successful for demand generation, then retroactively aligned the interventions with the 5C constructs. Results: Demand was positively correlated with high confidence and collective responsibility. Psychological constraints sometimes impacted demand. Physical constraints created barriers in some communities, particularly difficult to access (i.e., mountainous). Occasionally, physical constraints did not affect vaccination behavior - parents believed the benefits of vaccination worth pursuing. Factors negatively correlated with demand and intent, complacency and calculation, had limited impact. Critical interventions were: targeted and tailored health education activities (media partnerships, school outreach); community engagement; community ownership; and community involvement (community health workers, leaders, religious figures). Conclusion: We found similar interventions used to generate demand, with strategies aligned with the 5C constructs. Categorizing interventions by drivers of demand may help strategic planning and the division of resources; decision makers may choose to implement our suggested interventions. Assessing the 5Cs allows decision-makers to operationalize demand generation into concrete interventions and policies, and determine the individual impact of these constructs on the population and focus efforts on interventions tailored to a specific need.

Plant-pest interactions involve multifaceted processes encompassing a complex crosstalk of pathways, molecules, and regulators aimed at overcoming defenses developed by each interacting organism. Among plant defensive compounds against phytophagous arthropods, cyanide-derived products are toxic molecules that directly target pest physiology. Here, we identified the Arabidopsis (Arabidopsis thaliana) gene encoding hydroxynitrile lyase (AtHNL, At5g10300) as one gene induced in response to spider mite (Tetranychus urticae) infestation. AtHNL catalyzes the reversible interconversion between cyanohydrins and derived carbonyl compounds with free cyanide. AtHNL loss- and gain-of-function Arabidopsis plants showed that specific activity of AtHNL using mandelonitrile as substrate was higher in the overexpressing lines than in wild-type (WT) and mutant lines. Concomitantly, mandelonitrile accumulated at higher levels in mutant lines than in WT plants and was significantly reduced in the AtHNL overexpressing lines. After mite infestation, mandelonitrile content increased in WT and overexpressing plants but not in mutant lines, while hydrogen cyanide (HCN) accumulated in the three infested Arabidopsis genotypes. Feeding bioassays demonstrated that the AtHNL gene participated in Arabidopsis defense against T. urticae. The reduced leaf damage detected in the AtHNL overexpressing lines reflected the mite's reduced ability to feed on leaves, which consequently restricted mite fecundity. In turn, mites upregulated TuCAS1 encoding β-cyanoalanine synthase to avoid the respiratory damage produced by HCN. This detoxification effect was functionally demonstrated by reduced mite fecundity observed when dsRNA-TuCAS-treated mites fed on WT plants and hnl1 mutant lines. These findings add more players in the Arabidopsis-T. urticae interplay to overcome mutual defenses.

Glucosinolates are antiherbivory chemical defense compounds in Arabidopsis (Arabidopsis thaliana). Specialist herbivores that feed on brassicaceous plants have evolved various mechanisms aimed at preventing the formation of toxic isothiocyanates. In contrast, generalist herbivores typically detoxify isothiocyanates through glutathione conjugation upon exposure. Here, we examined the response of an extreme generalist herbivore, the two-spotted spider mite Tetranychus urticae (Koch), to indole glucosinolates. Tetranychus urticae is a composite generalist whose individual populations have a restricted host range but have an ability to rapidly adapt to initially unfavorable plant hosts. Through comparative transcriptomic analysis of mite populations that have differential susceptibilities to Arabidopsis defenses, we identified β-cyanoalanine synthase of T. urticae (TuCAS), which encodes an enzyme with dual cysteine and β-cyanoalanine synthase activities. We combined Arabidopsis genetics, chemical complementation and mite reverse genetics to show that TuCAS is required for mite adaptation to Arabidopsis through its β-cyanoalanine synthase activity. Consistent with the β-cyanoalanine synthase role in detoxification of hydrogen cyanide (HCN), we discovered that upon mite herbivory, Arabidopsis plants release HCN. We further demonstrated that indole glucosinolates are sufficient for cyanide formation. Overall, our study uncovered Arabidopsis defenses that rely on indole glucosinolate-dependent cyanide for protection against mite herbivory. In response, Arabidopsis-adapted mites utilize the β-cyanoalanine synthase activity of TuCAS to counter cyanide toxicity, highlighting the mite's ability to activate resistant traits that enable this extreme polyphagous herbivore to exploit cyanogenic host plants.

Environmental RNAi has been developed as a tool for reverse genetics studies and is an emerging pest control strategy. The ability of environmental RNAi to efficiently down-regulate the expression of endogenous gene targets assumes efficient uptake of dsRNA and its processing. In addition, its efficiency can be augmented by the systemic spread of RNAi signals. Environmental RNAi is now a well-established tool for the manipulation of gene expression in the chelicerate acari, including the two-spotted spider mite, Tetranychus urticae . Here, we focused on eight single and ubiquitously-expressed genes encoding proteins with essential cellular functions. Application of dsRNAs that specifically target these genes led to whole mite body phenotypes—dark or spotless. These phenotypes were associated with a significant reduction of target gene expression, ranging from 20 to 50%, when assessed at the whole mite level. Histological analysis of mites treated with orally-delivered dsRNAs was used to investigate the spatial range of the effectiveness of environmental RNAi. Although macroscopic changes led to two groups of body phenotypes, silencing of target genes was associated with the distinct cellular phenotypes. We show that regardless of the target gene tested, cells that displayed histological changes were those that are in direct contact with the dsRNA-containing gut lumen, suggesting that the greatest efficiency of the orally-delivered dsRNAs is localized to gut tissues in T. urticae .

Plant secondary metabolites are well-recognized medicinally important compounds. Gossypol is an important plant secondary metabolite with several medicinal properties. Carbon nanotubes (CNTs) are allotropes of carbon with diverse applicability in chemical, physical, and biological sciences due to their high surface area. The current study demonstrates the enhancement of gossypol production in cotton cell suspension culture in culture media supplemented with water-soluble carbon nanotubes. The fresh and dry weights of cotton cell suspension culture grown in MS media with 20 µg/mL CNTs were, respectively, 1.9 and 2.13 fold higher than in control MS media after one month. The net enhancement of gossypol production in MS media supplemented with 20 µg/mL CNTs was 2.47 fold higher than the control. Confocal and SEM imaging showed the presence CNTs on the cell surface, which mediated the formation of extra channels that resulted in high biomass production in cotton cell suspension culture. The gossypol produced by this cell suspension culture showed antiproliferative activity against the prostate cancer cell line. Thus, this study demonstrated a new method for enhanced gossypol production, which can prove beneficial for the production of other plant-based biological active compounds.