School-based behaviour change communication interventions could help to achieve behavioural changes in the school and enhance the enrollment of the students and teachers as health messengers to local communities. Evidence on the impacts of the school-engaged malaria preventive interventions are limited as far as the social and behaviour change communication (SBCC) is concerned. This study examined the effectiveness of the school-based SBCC approach on insecticide-treated nets (ITNs) utilization among primary school students in malaria-endemic settings of Ethiopia.Various participatory, educational, and communication interventions were implemented from 2017 to 2019 in 75 primary schools and respective villages in Jimma to promote malaria preventive practices. A quasi-experimental design was conducted with randomly selected 798 students (i.e. 399 intervention and 399 control groups). Data were collected by trained interviewers using structured questionnaires. The SPSS version 26 software was used to analyse the data. Propensity score matching analysis was performed to control for possible confounding biases. The average effects of the intervention were estimated using multivariate general linear modelling to estimate for mean differences and odds ratio based on the nature of data.The result showed that the ITNs utilization was 6.857 folds in the intervention groups compared to the counterpart; ; effect size = 39%). A mean differences (MD) of self-efficacy , knowledge , attitude , perceived malaria risk , and perceived family supports  were observed favoring the intervention. Multivariable logistic regression modelling results showed that knowledge  and perceived family supports  and self-efficacy  predicted the ITN utilization among the school children.The finding of this study suggested that the school-based SBCC approach combined with peer education activities advanced the malaria-related knowledge, attitude, self-efficacy, risk perceptions, and family supports and ultimately improved the sustained use of ITNs among school-going children. Further research should be conducted to understand the mechanism of these effects given the influences of social, health services, and school systems are considered. Anopheles species  and lowest [11 (6.4%)] exposures rate to information from peer educators' were reported among intervention and control groups, respectively. The same thing is true for teachers, community, and mass media with 135 (91.8%) and 80 (22.3%), 31 (68.9%) and 14 (31.1%), and 68 (87.8%) and 21 (23.6%) in control and intervention groups, respectively (Table .8% and cThe results showed a mean differences (MD) of self-efficacy , comprehensive knowledge , attitude , perceived malaria risk , and perceived family support  favoring the intervention group. But the finding showed a lower mean score for the perceived severity of malaria  among intervention groups. Further, comparatively, the highest and lowest impact of the intervention (effect sizes) was reflected on self-efficacy (ES = 31%), and perceived malaria (ES = 7%) reported the presence of at least one ITN in the household. Respondents who reported having at least one ITN in the household is nearly three times higher in the intervention group . There was not a considerable difference in ownership of more than one ITN per two family members between the intervention and control groups . The ITNs utilization was 6.857 folds in the intervention groups compared to the counterpart ; effect size, ES = 39%). This implies the ITNs use was 39% higher in the intervention group which was attributed to the intervention (Table Multivariable logistic regression modelling (adjusted for predicted propensity score and intervention status) were conducted to examine how psychographic dimensions interacted with each other to affect the primary outcome of the intervention (i.e. ITN utilization among primary school students). Accordingly, knowledge (MDK), self-efficacy, perceived family support, peer educators as main sources of information, and the ratio of ITN in the household were predicted the ITN utilization.This implies that students who reported having one ITN per two family members were nearly two times more likely to use ITNs . Higher scores of knowledge  and perceived family support , and self-efficacy  were associated with improved use of ITNs among students. However, important factors in the health behaviour change process such as attitude , perceived disease risk , and severity  were not predicted ITNs use among students . Those who had adequate access to the nets and thought they had greater family support were more likely to use it and vice versa  and the utilization of ITNs among target students, indicating the effectiveness of the interventions.Knowledge is considered a basic factor in the process of health behaviour change efforts. The knowledge score which was measured as knowledge was considerably higher among the intervention groups in our study. A study conducted in India to evaluate the effects of multiple behaviour change approaches  showed significant improvement in knowledge by 21% among students in the intervention schools . In MaliHaving led to a improvement in perceptions of disease risks, attitudes, and self-efficacy towards malaria preventive actions, the current interventions justified the notions of cognitive-behavioural theories . Thus, iThe possible reasons for this paradoxical result may be due to the improvement in self-efficacy and social supports on ITN utilization among the intervention groups. This finding is appears consistent with the result of a community-based study conducted to evaluate the same programme (i.e. the school-based SBCC approach), which indicated the reduction in the community’s attitude (− 3.5%) and perception of risk (− 3.3%) regarding malaria . The afoFamily participation in school health programmes was considered one of the principal components of the Health Promoting Schools policy . The theFurther, the primary outcome of interest in the current study was changes in the utilization of the ITNs in the intervention targets. In principle, the observed changes in the levels of psychographic and social supports outcomes should be translated into actions or behaviours . The resgeengoo qulqullina barnootaa” which means the circle for quality of education. The circles for the quality of education are made of members representing the community, teachers, students, and accountable to school directors have the ultimate task of improving the quality of education. More importantly, the community-based evaluation study conducted enrolling parents indicated improved acceptance of the intervention that employed students as messengers of malaria information [The possible difference might be attributed because of integrated peer education networks and community-led SBCC approaches in the current programme. Parents were actively participating in the programme through what is locally called “ormation .The multivariable logistic regression modelling identified knowledge, self-efficacy, family supports, peer educators as sources of information, and the ratio of ITNs in the household as independent predictors of ITN utilization among school students. Access to more than one ITN per two persons in the household independently predicted the use of ITN in this study. A similar finding was reported from a previous school-based study in that ownership was found as the main predictor of ITN use . EnsurinIn the previous study, the total knowledge index was associated with both ITN ownership and its use. However, the study reported varying levels of influence for different domains of knowledge . In shorThough evidence is scarce in this regard; the implication of the current study could be explained by the notion of cognitive-behavioural theories. Given that adequate knowledge, a higher tendency of perceived coping responses to certain health risks  can lead to behaviour change . HoweverPerceptions about family support independently predicted the use of ITN among school children in this study. Effective behaviour change interventions require enhancement of social support and this is the ultimate goal of SBCC strategy which influences community norms, social factors, and reinforcements to bring about sustained behaviour change . FamilieThe analysis has evaluated the effects of various sources of malaria information such as peer educators, teachers, mass media, and community  on ITN use among students. Among these various sources reported in this study; peer educators were found positively associated with the use of ITN in this study. This might be because of greater attention and opportunity for open discussions that result from perceived age similarity among the students and the length of time they spent together . The schsituational constraints related to the presence of external factors. Situational constraints are circumstances that affect (through moderation) the relationship between attitude and behaviours [strong situations and this in turn weakens the relationship between attitude and behaviours [weak situations than in strong ones.The relationship between ITN use and important factors such as attitude and perceptions of disease threat was not importantly high in this study and this contradicts the principles of many behavioural change theories . Specifihaviours . Accordihaviours . In shornon-responsive attitude or affection that is acquired or learned from the existing values or social norms. For instance, ITN may be most commonly liked in the area simply for some reason other than its actual benefits. Such a situation may also happen if attitude questions in the survey were understood more as affection rather than objectively defined benefits and this was explained in observational learning theory (the modelling effects). It was also indicated that attitudes predict behaviour to the extent that attitude and behaviour are measured at the same level of specificity [Second, students might hold a cificity . HoweverFor this study, information on a parasitological survey which might have added value to the findings was not collected. Arguably, the choice of outcome measures for health intervention is critical in that health outcomes should be considered the index of success of interventions designed to change health behaviours . HoweverFurthermore, limitation inherently to the unmatched design employed in the current study might have introduced bias that affects the result. However, an advanced statistical technique: the PSM analysis was applied as matching analysis to remove the possible confounding bias arising from the lack of matching so that the average effects of the intervention were easily estimated .On top of this, different sources of biases for this study were also considered and dealt with. For instance, the possibility of information contamination among the control groups due to human mobility and interaction is often difficult to control in communication interventions. Furthermore, performance bias and social desirability bias (SDB) due to the self-reported behaviours might have also affected the data. Presumably, the SDB may be high among the intervention group since the population under study was already aware of the aim of the intervention and could respond to the interview in a socially desirable manner; thus hiding underlying reality. The measure of SDB score was controlled during the subsequent analysis to effectively estimate the averaged effects of the intervention. The data collectors were kept blind about the intervention and the target settings to control for possible performance bias.This study suggested the school-based SBCC approach advanced the malaria-related knowledge, attitude, self-efficacy, risk perceptions, and family supports, which have ultimately improved the sustained use of ITNs among school-going children. Schools' involvement through the integrated peer education and SBCC approaches would result in effective behaviour changes in schools regarding malaria preventions and beyond, which in turn helps to accelerate malaria elimination efforts. Furthermore, research should be conducted to broadly understand the mechanism by which the SBCC approach affects students’ behaviours given the influences of social, health services, and school systems.


Forkhead Box N1 transcription factor. This transcription factor supports the development of TECs, the controllers of T cell fate, and defective TECs means no T cells.The thymus is a bi-lobed lymphoid organ localized above the heart whose primary function is to foster the development of the T cells of the adaptive immune system. T cells are a critical component of the cellular immune system, helping B cells produce antibodies, releasing cytokines to orchestrate effective immune responses, and killing infected cells and/or neoplasm/tumor cells. The recognition of an infected cell or tumor cell, and the ability to support B cell secretion of antibodies requires T cells express a cell surface receptor, termed the T cell receptor (TCR). This receptor selectively binds to peptides complexed to major histocompatibility complex antigens (MHC class I and II). MHC class I molecules are present on all nucleated cells in the body, while MHC class II is restricted to antigen presenting cells. In the thymus, thymocytes rearrange DNA loci comprising the genes encoding the TCR subunits. Using a process termed VDJ recombination, each thymocyte expresses a unique TCR with exclusive recognition specificities. However, only those thymocytes expressing TCRs that can engage self-peptides/self MHC complexes present on thymic epithelial cells (TECs) are “selected” to form the peripheral T cell repertoire. This involves processes coined positive and negative selection, with the ensuing repertoire of T cells capable of recognizing pathogen- or distinct tumor- derived peptides presented on self-MHC molecules without overt reactivity to self-peptides. The critical role of the thymus in this process is best exemplified with infants born with mutations in genes required for the formation of the thymus (22q11.2 deletion syndrome or DiGeorge). No thymus results in no T cells, leading to a severe combined immunodeficiency. A second excellent example pertains to the spontaneously arising nude mouse, which lacks a thymus and hair due to mutations in the Yet, as late as 1960, the thymus was a scientific conundrum. Dramatic variations in its size in humans and diverse species had been documented for over two millennia. Severe clinical repercussions based on these size differences ensued due to a complete lack of knowledge about its functions, which was not revealed until demonstrated in pivotal experiments performed by Jacques Miller using thymectomy and skin grafting. These experiments established the thymus as the site of T cell development. For this, Miller was awarded the Lasker Prize in Biomedicine in 2019 . Such anBhalla et al.; Giardino et al.). The contributions of key transcription factors such as T Box Transcription Factor 1 (TBX1) and FOXN1 in the patterning of the pharyngeal apparatus and the subsequent demarcation of the thymus anlage are discussed in conjunction with the differentiation and expansion of thymic epithelial cells. The formation of the two different populations of epithelial cells, cortical and medullary TECs and their selective roles in supporting and educating thymocytes to form the repertoire of T cells, which is unique to each individual, is described, revealing new subsets of these cells in the process .In the series of articles under the title “New Insights into thymic functions during stress, aging, and in disease settings” in this issue of Frontiers in Immunology, novel insights and findings by leading scientists in the field of thymus biology are presented. The articles include two reviews outlining recent discoveries into the mechanisms required for the formation and specification of the thymus within the 3rd pharyngeal pouch during embryogenesis . Such thymic hypoplasia is widespread in the population, and the different conditions leading to hypoplasia/aplasia of the thymus are presented in a combination of research and review articles in this series  For example, thymic hypoplasia is evident in pregnant women and infants born prematurely. The natural aging process causes a gradual and “permanent” hypoplasia of the thymus and this involves both loss of developing T cells and TEC subsets, with a greater impact evident in males than females, at least until middle-age . The epithelial tissue within the thymus undergoes adipogenesis and epithelial to mesenchymal transitions . Given these varied and damaging processes, improving and/or restoring thymopoiesis, from infants to the elderly, requires new therapeutic strategies. Several innovative approaches covered in this series include use of pro-T cells to improve T cell formation in the thymus  and the use of specific cytokines to restore TEC functions (Kinsella and Dudakov). Humanized mouse models and comparative mouse-human studies offer additional understanding and methodologies for improving thymus functions .As is well-recognized currently, in addition to aging, the thymus undergoes a transient and sometimes massive reduction in overall size and cellularity following infections, corticosteroid production through the hypothalamus-pituitary-adrenal axis or via injections, and chemotherapy treatments. The impact on thymocytes and stromal cell populations are presented herein . While the thymus is usually considered in the context of T cell development, there is evidence of some B cell development in normal and clinical settings. Two pronounced conditions, the autoimmune diseases, Systemic Lupus Erythematosus and Myasthenia Gravis, have substantial levels of B cell development/expansion . In the case of Myasthenia Gravis, thymectomy is actually done for patients to eliminate the autoreactive B cells in the thymus, improving their clinical outcomes. Alzheimer's Disease is another clinical condition where mouse models have established that intrathymic injections of TECs, derived from stem cells, can actually improve cognitive functions . In summary, we have come a long way from purposely destroying the thymus to focusing on its restoration and regeneration. These are nicely collated in our Frontiers in Immunology Topic.The emergence of checkpoint blockade immunotherapy to treat cancer patients has consequences as well, since the incidence of autoimmune complications with such a patient cohort increases dramatically due to the antibody treatments (All authors participated in the editorial reviews of the manuscripts. All authors participated in the writing and editing of the editorial article.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


In a previous feasibility study (APROPOS) in a single municipality of the Netherlands, we showed that a locally tailored preconception care (PCC) approach has the potential to positively affect preconceptional lifestyle behaviours. Therefore, we designed a second study (APROPOS-II) to obtain a more robust body of evidence: a larger group of respondents, more municipalities, randomization, and a more comprehensive set of  outcomes. The aim of this study is to assess the effectiveness and the implementation process of a local PCC-approach on preconceptional lifestyle behaviours, health outcomes and the reach of PCC among prospective parents and healthcare providers.This study is an effectiveness-implementation hybrid type 2 trial. This involves a stepped-wedge cluster randomized controlled trial design aiming to include over 2000 future parents from six municipalities in the Netherlands. The intervention has a dual-track approach, focusing on both the uptake of PCC among future parents and the provision of PCC by healthcare providers. The PCC-approach consists of 1) a social marketing strategy directed towards prospective parent(s) and 2) a local care pathway to improve interdisciplinary collaboration. Data will be collected before and after the introduction of the intervention through questionnaires and medical records in each of the participating municipalities. The primary outcome of this study is change in lifestyle behaviours . Secondary outcomes are pregnancy outcomes  and the uptake of PCC. Moreover, a process evaluation will be performed, providing information on the efficacy, feasibility, barriers and facilitators regarding the implementation of the intervention.The APROPOS-II study introduces a locally tailored PCC-approach in six municipalities in the Netherlands that will contribute to the body of evidence regarding the effectiveness of PCC and its implementation. If this intervention has a positive effect on lifestyle behaviour changes, leading to improved pregnancy outcomes and the future health of prospective parents and their offspring, it could subsequently be upscaled to (inter)national implementation.NL7784 .Dutch Trial register:  Despite major advances in clinical research and medical technology, the prevalence of adverse maternal and neonatal health outcomes, such as pre-eclampsia and preterm birth, have only moderately decreased over the past decade . As the One way to actively prepare for pregnancy is by using preconception care (PCC). PCC is defined as “a set of interventions that aim to identify and modify medical, behavioural and social risks to a woman’s health or pregnancy outcome through prevention and management” . DespiteA previous feasibility study performed by our group (APROPOS) in one municipality of the Netherlands showed that a locally tailored PCC-approach has the potential to positively affect preconceptional lifestyle behaviours and increases the use of PCC among prospective parents . After bTo determine the effect of the intervention on preconceptional lifestyle behaviours  and other risk factors  among women who recently conceived;to assess the effect of the intervention on the reach of prospective parents regarding PCC, the uptake of PCC and pregnancy preparation;to evaluate the implementation process in order to gain information on the feasibility, barriers and facilitators for the implementation of the local PCC-approach;to evaluate the sustainability of the intervention, with a specific focus on the diversity within and between the participating municipalities.The aim of this study is to assess the effectiveness and the implementation process of a local PCC-approach (i.e. the intervention) on preconceptional lifestyle behaviours, health outcomes and the reach of PCC among prospective parents and healthcare providers. Therefore, we have specified the following objectives:The APROPOS-II study uses a hybrid effectiveness-implementation design , 15. In The study will be conducted in six municipalities distributed throughout the Netherlands . In total, ten community midwifery practices in these municipalities participate in the study. Municipalities were selected for diversity regarding their size , population, health challenges and organization of care. All women above the age of 18 years who adequately master the Dutch, English, Polish or Turkish language are eligible to participate in this study. Men are recruited through their participating partners.Our intervention targets all men and women in their reproductive life span. In all six participating municipalities together, this is approximately 270,000 people . Figure Uptake of PCC: To improve preconceptional lifestyle behaviours and the uptake of PCC, a social marketing strategy, directed towards prospective parent(s), will be distributed in each participating municipality. This online and offline social marketing strategy aims to trigger prospective parents to actively prepare for pregnancy by directing them towards reliable PCC-information and promoting individual PCC-consults.Provision of PCC: To improve interdisciplinary collaboration among healthcare providers, a local care pathway will be developed and implemented in each participating municipality, including interdisciplinary arrangements for collaboration and referral between primary, secondary and tertiary care. Moreover, a working conference will be held among local healthcare providers of multidisciplinary backgrounds and a stakeholder coalition will be formed.The intervention used in this study is a PCC-approach tailored to the needs of prospective parent(s) and healthcare providers in a local setting. This intervention has been developed based on pre-implementation research in the APROPOS feasibility study . The intwww.wokewomen.nl (in Dutch).Concerning the uptake of PCC, the social marketing strategy has been developed by a professional and experienced marketing agency. A social marketing strategy uses commercial marketing technologies to design programs that can influence the behaviour of the target audience in order to improve personal welfare and that of society . The theRegarding the provision of PCC, the working conference for the healthcare providers aims to educate them on preconceptional health and risk factors using the national Preconception Indication List (PIL), emphasizes the importance of facilitating PCC and is used to conduct a region-specific bottleneck analysis  19]. Su. Su19]. www.zwangerwijzer.nl) and the APROPOS feasibility study [Study parameters will be assessed by a questionnaire distributed among prospective parent(s) by the participating community midwifery practices in the first trimester of pregnancy. The questionnaire is based on validated questionnaires such as WHO - Quality of life questionnaire, the London Measure of Unplanned Pregnancy (LMUP), a validated preconception tool , expressed as the number of women who preconceptionally change at least one of these lifestyle behaviours from inadequate to adequate. The definitions of healthy lifestyle behaviour are based on Dutch guidelines and are provided in Table Secondary outcomes are pregnancy outcomes  and the uptake of PCC. The definitions of these pregnancy outcomes are given in Table The RE-AIM framework will be used to evaluate the intervention and the implementation strategy. This framework is a practical way of evaluating health interventions and has previously been used in studies focused on changing individual behaviours . RegardiHealthcare providers’ implementation outcomes will be measured by a 23-item validated NoMAD  questionnaire. The NoMAD questionnaire will be used to describe the health care providers’ views on how the intervention impacts their work and their expectations about whether the intervention could become a routine part of their work . The heaFinally, the feasibility of the implementation of the local care pathways will be evaluated among healthcare providers. We will measure to what degree the local care pathway was distributed amongst the healthcare providers and how healthcare providers experienced the convenience of this tool. In each municipality, focus groups with the local stakeholder coalition will be held 9 months after the start of the intervention and will be analysed using thematic analysis to identify key issues and themes. In these focus groups, information will be gathered on how the intervention complies with the heterogeneity of local settings, since differences in healthcare networks, logistics and practical issues call for tailored approaches.Based on the results from the APROPOS feasibility study, we anticipate on an odds ratio of 1.5 in lifestyle behaviour change during the preconception period (primary outcome). Based on the sample size estimation method of Hemming and Taljaard, with an alpha of 0.05, power of 80% and a low intraclass correlation Coefficient of 0.02, the sample size was calculated to be 363 participants per unit with 6 units  . ConsideP-values < 0.05 are considered statistically significant.Statistical analysis will be performed using multilevel logistic regression analysis to take into account the clustering of respondents within municipalities. Data will be analysed anonymously on two levels; the respondents’ level (before and after the intervention) and the municipal level . Baseline characteristics will be compared between the pre- and post-intervention group and compared in a baseline table. Chi-square analysis and ANCOVA will be used to study the effects of the intervention on the adverse pregnancy outcomes. Adjusted odds ratios will be calculated taking into account the potential confounders: age, educational level and parity. The APROPOS-II study introduces a locally tailored PCC-approach in six municipalities in the Netherlands, that will contribute to current knowledge regarding the implementation and the effectiveness of PCC. Until now there is little data from randomized clinical trials that prove the effectiveness of PCC on maternal and perinatal health outcomes , 42, 43.www.wokewomen.nl) with practical and evidence-based information to help them improve their preconceptional health status. In addition, we encourage these women when there is a need for extra information to visit a healthcare provider for a PCC-consult.Despite the low uptake of PCC-consults, the majority of prospective parents use the internet as their primary source of information regarding preconception health . Women aPrevious literature on social marketing strategies showed that it has the potential to improve diet, increase exercise and reduce substances-use like tobacco, and alcohol . CreatinStrengths of this stepped-wedge cluster RCT are the diversity of the population in the participating municipalities and the tailored intervention that complies with the heterogeneity of local settings. Moreover, the detailed questionnaire investigating respondents’ lifestyle behaviours, health beliefs and the extent of planning of the current pregnancy based on validated questionnaires can be considered a strength of this study. In addition, the innovative social marketing strategy, the extensive process evaluation and the involvement of the prospective father highlight different and often neglected aspects of PCC-research.A potential limitation of this study is the occurrence of selection bias, as participants who have actively prepared for their pregnancy are probably more eager to share their experience. However, in our previous feasibility study, response rates were high and the population characteristics were similar before and after introduction of the intervention . AnotherWith this study, we expect to effectively implement and evaluate a locally tailored PCC-approach. If this intervention will show to improve lifestyle behaviours, leading to improved pregnancy outcomes and future health of prospective parents and their offspring, it may be upscaled to (inter)national implementation.


The hallmark of sex chromosome evolution is the progressive suppression of recombination which leads to subsequent degeneration of the non-recombining chromosome. In birds, species belonging to the two major clades, Palaeognathae (including tinamous and flightless ratites) and Neognathae , show distinctive patterns of sex chromosome degeneration. Birds are female heterogametic, in which females have a Z and a W chromosome. In Neognathae, the highly-degenerated W chromosome seems to have followed the expected trajectory of sex chromosome evolution. In contrast, among Palaeognathae, sex chromosomes of ratite birds are largely recombining. The underlying reason for maintenance of recombination between sex chromosomes in ratites is not clear. Degeneration of the W chromosome might have halted or slowed down due to a multitude of reasons ranging from selective processes, such as a less pronounced effect of sexually antagonistic selection, to neutral processes, such as a slower rate of molecular evolution in ratites. The production of genome assemblies and gene expression data for species of Palaeognathae has made it possible, during recent years, to have a closer look at their sex chromosome evolution. Here, we critically evaluate the understanding of the maintenance of recombination in ratites in light of the current data. We conclude by highlighting certain aspects of sex chromosome evolution in ratites that require further research and can potentially increase power for the inference of the unique history of sex chromosome evolution in this lineage of birds. Sry gene (Sex-determining region Y) is located on the Y chromosome and its inheritance to the male embryo determines sex by activating the downstream male regulatory pathway [Dmrt1 (Doublesex- and mab-3-related transcription factor 1) as the sex-determining gene. Dmrt1 is located on the Z chromosome and its higher expression in males relative to females drives the formation of male gonads [Sex chromosomes are the main players in the genetic sex determination of mammals and birds. In eutherian mammals, males are the heterogametic sex, meaning that males carry two morphologically distinct sex chromosomes , and females are the homogametic sex (XX). In birds, the heterogametic sex is the female  and males are the homogametic sex (ZZ). In the mammalian XY system, the  pathway . In birde gonads . Sex chre gonads , repeat-e gonads ,6 and hee gonads . A key pe gonads ,9,10. Several theoretical studies have aimed at explaining the evolution of recombination suppression and its extension beyond the sex-determining region (reviewed in detail in ). The moIn this view of sex chromosome evolution, a progressive suppression of recombination and subsequent loss of genetic material during Y/W degeneration seems to be inevitable. However, the correlation between the age of sex chromosomes and extent of recombination suppression seems to be weak . This caDmrt1, is located on the Z chromosome in both lineages and is absent not only from the gene-poor W chromosome of Neognathae, but it has also been lost from the W chromosome in ratites [The striking difference in the state of W degeneration between Neognathae and ratites is particularly puzzling since sex chromosomes in these two lineages share a common ancestor . The sex ratites . In fact ratites ,37,39. M ratites  and avia ratites . In thisIn this section, we discuss selective processes that can influence the evolution of recombination suppression and its subsequent chromosome degeneration in the context of ratite sex chromosomes B. These The most prominent hypothesis for the evolution of suppressed recombination between the evolving sex chromosomes involves sexual antagonism . SuppresDmrt1 is located at 76.2 Mb near the chromosome end at 80.9 Mb [Dmrt1 is located at 25.9 Mb. Although the gene number and content is conserved between different avian lineages [Dmrt1 and its relative location to the chromosome centromere might have an influence on the rate of degeneration of the chromosome. For mutations to have a sexually antagonistic effect, there must be phenotypic variation between sexes that antagonistic selection can act upon. Estimating the prevalence of sexual antagonism within a species is challenging. However, if mating system can be taken as a proxy for the intensity of sexual selection, various mating systems, from monogamy to harem formation with intense male competition, exist among ratites as they do in birds with differentiated sex chromosomes ,43 Tabl. Hence,  80.9 Mb . In chiclineages , the seqIn the presence of sexual antagonism, SA mutations can only accumulate on the sex chromosome if a non-random association ) is established between the sex-determining region and the SA mutation. Without LD, it is not possible for the SA mutation to be associated with the sex it favors. Therefore, one way to prevent the accumulation of sexually antagonistic mutations and the consequent recombination suppression is selection for a higher rate of recombination in the PAR of the heterogametic sex close to the sex-determining region . This caIn the case of the establishment of LD between the sex-determining region and the SA mutation, other mechanisms besides recombination suppression might be able to guarantee the restriction of the SA allele in the sex it favors. One such mechanism is sex-biased gene expression, where a male-beneficial mutation present on the Z or W chromosome gets down-regulated in females, and vice-versa in males. One such example of this is in the emu, where it has been suggested that sex chromosomes have become masculinized due to the presence of male-biased expression of Z-linked genes . In a reIn the previous section, we discussed that under sexually antagonistic selection, the occurrence of a selected allele loosely linked to the sex-determining region will lead to selection for reduced recombination. In this section, we discuss the model by Otto that shows that, under specific conditions, there can be selection for increased recombination between a selected locus and the sex-determining region . This moIt is possible to speculate about the relevance of this hypothesis for the case of recombination maintenance in Palaeognathae. Recombination across most of the W is maintained across all species of ratites and to various degrees in tinamous, which implies that either balancing selection has acted independently in several lineages or all ratite species harbor ancestral balanced polymorphisms. Since the major lineages of Palaeognathae diverged tens of million years ago from one another ,49, lineLack of recombination between the sex chromosomes in the heterogametic sex reduces the efficacy of natural selection and eventually leads to the degeneration of the non-recombining chromosome. With a degenerated Y/W chromosome, there will be only one copy of the X/Z-chromosomal content in the heterogametic sex. Reducing number of gene copies  to half can have severe fitness consequences ,51. FitnDosage compensation does not necessarily occur uniformly for all genes. In birds, dosage compensation has been shown to act in a gene-specific manner . In chicIn this section, we discuss neutral processes that are relevant for both recombination suppression and subsequent chromosome degeneration C. DegeneRatites have long generation times and large bodies , which tThe explanation for this is the “accordion model”, where an accumulation of TEs provides substrates for non-allelic homologous recombination (NAHR)  and is, therefore, followed by an increased rate of deletions countering the genome size increase. Due to this effect, birds with more TE accumulation end up with smaller genomes . In linedS (synonymous substitutions per synonymous site) between 0.0464 and 0.1898 reside within an about 10 Mb region of the Z chromosome . ConverTaken together, the slow molecular evolution of ratites and the low degree of heterochromatinization of their W chromosomes might simply reflect that fewer recombination-reducing mutations occurred or became fixed than in other birds. Whether recombination suppression mainly spreads via discrete steps  or gradual changes  remains to be determined because reference-quality W chromosome assemblies are scarce in birds .Sex chromosome evolution has been a topic of intense research for several decades. The most common hypothesis to explain recombination suppression  has not provided a conclusive explanation for the observed patterns of recombination suppression and chromosomal degeneration in sex chromosomes . This isTo assess the relevance of the sexual antagonism hypothesis in ratite sex chromosome evolution, we must first aim at quantifying levels of sexual conflict in these birds. This of course is another difficult task to achieve. One approach is to compare the mating system and levels of sexual dimorphism in ratites with tinamous and other groups of Neognathae that show various extents of sex chromosome degeneration. Estimates of genetic diversity across the sex chromosomes can also inform us about the presence of sexually antagonistic selection. Theoretical work has shown that levels of genetic diversity are expected to be higher in the PAR region compared to autosomal loci, because the PAR is in linkage to the sex-determining region in the heterogametic sex  . The comIt remains unclear how much of recombination suppression occurred in discrete vs. gradual steps. To this end, obtaining high-quality assemblies of Z and W chromosomes of ratites would be instrumental for detecting inversions. Studying the chromatin state of sex chromosomes, particularly the region close to the sex-determining region, using chromatin immunoprecipitation (ChIP) with massively parallel sequencing, will likely provide a better understanding of the impact of repressive histone states of DNA sequences on sex chromosome evolution. Taking all neutral and selective processes into account will reveal whether there is some lineage-specific mechanism that slowed down the rate of sex chromosome recombination suppression and W degeneration in ratites.


This population growth poses several challenges to the global food system, which will need to produce more healthy food using fewer natural resources, reducing the environmental impact, conserving biodiversity and flexibly adjusting to changing societal expectations. Meeting this demand requires environmentally sustainable improvements to farmed animal health and welfare, and of efficiency and diversification  [The Food and Agriculture Organisation of the United Nations (FAO) reports that by the year 2050 the global human population is likely to reach 9.7 billion, rising to 11.2 billion by 2100  project . The firIn the past 20 years, genomic selection has substantially increased genetic gain in some farmed animal species through the use of large training populations . Howeverhttps://gtexportal.org/home/) has achieved this very effectively across human tissues, enabling expression QTL (eQTL) studies linking gene expression to genetic variation [GTEx). Large farmed animal cohorts in controlled and well-characterised environments with extensive pedigree information and molecular phenotypes would allow researchers, in partnership with industry, to (1) build better predictive models of genotype-to-phenotype, (2) better understand genotype-by-environment interactions and (3) prioritise functional variants for inclusion in breeding programmes [The first phase of FAANG is using a specific set of transcriptomic and epigenomic assays to define functional regions of the genome in tissues . Due to ariation  and provogrammes . Hundredogrammes . A projeogrammes .Beyond its use in genomic prediction, the functional data produced by FAANG will provide new perspectives for informed management decisions. Epigenetic and expression information for individual animals could be combined with microbiome data and high-throughput phenotypes from new management technologies  . These dThrough large-scale sequencing efforts by the farmed animal genomics community data are now accumulating that characterise the sequence diversity of farmed animals including locally adapted breeds/populations. As a consequence, future genetic management is likely to include the use of pangenomes that will capture all available population-level genomic information for a given farmed animal species. Using graph-based frameworks, we can more accurately genotype and annotate the genomic diversity present in any given individual . In thisSingleCell). The organisational processes, standardisation and data sharing infrastructure established by the community for the first stages of FAANG [SingleCell to progress quickly and efficiently. The FAANGSingleCell project should build on existing functional tissue maps for other species [GTEx project described above, single-cell atlases will provide a powerful layer of resolution including cell-specific molecular phenotypes, enabling the fine-scale dissection of complex traits of interest.The use of bulk tissue samples in the FAANG studies performed to date captures regulatory elements and expression signals averaged across all represented cell types but fails to reveal the cell-specific basis of the molecular phenotypes of interest. In order to more accurately link genotype to phenotype, data at the level of individual cell types are required. Single-cell sequencing technologies enable the deconvolution of the transcriptional and regulatory complexity in tissues made up of multiple cell types. New technologies to detect gene expression as well as chromatin accessibility, structure and interactions within single cells provide more comprehensive data to predict function and interaction partners for regulatory elements. As a consequence, one of the main priorities for FAANG within the next 5 to 10 years is to create single-cell atlases for the key tissues of farmed animal species  is the focal point for this activity. Technological development, coordination and standardisation by the DCC will continue to be crucial for the shift towards population scale studies, single-cell datasets, cell atlases and pangenomes, across a growing number of species. This will require new reproducible analysis pipelines and infrastructure, metadata validation services, data portal features such as a centralised atlas browser and online training resources. Single-cell atlases and in vitro systems for farmed animal species will be accompanied by high quality metadata, archiving and visualisations across species, organ systems, tissues and cell types. As FAANG datasets continue to increase in complexity, there is a growing need for new methods of data visualisation and integration to be made available. These future developments, and the distributed data and analysis infrastructure, will be crucial for the successful application of functional data to farmed animal breeding programmes.As a scientific community, FAANG continues to develop a coordinated analysis and data collection infrastructure crucial for its success . The FAAhttps://faang.org/proj.php) was a major success for the first stage of FAANG and its leadership. Current funding for FAANG supports the research community to improve the functional annotation of key farmed animal species and to facilitate more refined genomics-enabled animal breeding/genetic improvement. The research priorities outlined here are already strategically aligned to the objectives of the European Green Deal  and current USDA National Institute for Food and Agriculture programmes . International cooperation will be essential to secure funding for their achievement. Given the scale and cost of the research involved, it will likely be necessary to initially prioritise the development of in vitro systems and the enhancement of data infrastructure to provide a solid foundation for FAANGSingleCell and FAANGGTEx.The research priorities we have outlined for FAANG for the coming decade are depicted in Fig. The timely achievement of all of the research priorities we outline here for the next stages of FAANG will together increase the capacity of the farmed animal production industry to face the challenges of the future, empowering genomic selection, enhancing adaptation to changing environments, conserving biodiversity and bridging the gaps between cellular and whole animal scale knowledge.


There is humoral-neural coupling between the systemic and brain RASs, which is important for controlling sympathetic tone and the release of endocrine factors that collectively determine blood pressure (BP). Similar to the interactions between the systemic and brain RASs is the communication between the peripheral and brain immune systems. Systemic inflammation activates the brain’s immune response. Importantly, the RAS and inflammatory factors act synergistically in brain regions involved in the regulation of BP. This review presents evidence of how such interactions between the brain RAS and central immune mechanisms contribute to the pathogenesis of hypertension. Emphasis focuses on the role of these interactions to induce neuroplastic changes in a central neural network resulting in hypertensive response sensitization (HTRS). Neuroplasticity and HTRS can be induced by challenges (stressors) presented earlier in life such as a low-dose of angiotensin II or high fat diet (HFD) feeding in adults. Similarly, the offspring of mothers with gestational hypertension or of mothers ingesting a HFD during pregnancy are reprogrammed and manifest HTRS when exposed to new stressors as adults. Consideration of the actions and interactions of the brain RAS and inflammatory mediators in the context of the induction and expression of HTRS will provide insights into the etiology of high BP that may lead to new strategies for the prevention and treatment of hypertension.Mounting evidence indicates that the renin-angiotensin (RAS) and immune systems interact with one another in the central nervous system (CNS) and that they are importantly involved in the pathogenesis of hypertension. Components comprising the classic RAS were first identified in the periphery, and subsequently, similar factors were found to be generated  Hypertension is a major risk factor for the development of cardiovascular diseases that are responsible for high morbidity and mortality. Approximately one-third of adults in the United States have hypertension, and only half of hypertensive patients show a satisfactory response to treatment . This maCNS mechanisms driving sympathetic nervous system (SNS) overactivity play an important role in the development and progression of many forms of experimental hypertension . The braAbout ten years ago, we discovered that the CNS can be reprogrammed by mild non-pressor challenges that induce a neurological state that is expressed as a sensitized hypertensive response when a challenge (stressor) is presented once again. Thicausal mechanisms in prenatals hypertensive response sensitization (HTRS) is the result of the induction of CNS neuroplasticity in the brain neural pathways controlling the tone of the SNS . Our finBesides a prohypertensive axis of the RAS, an antihypertensive axis of the RAS has also been identified and studied extensively regarding its protective role in the development of hypertension . This reThe hypothalamus is a key brain region contributing to the regulation of the cardiovascular system, fluid balance, and energy homeostasis. Maintenance of these functions requires the integration of information from neural and humoral signals initially derived from the systemic (aka the circulating or classic) RAS and the systemic immune system. The SFO and organum vasculosum (OVLT) lie along the lamina terminalis (LT) and the area postrema (AP), located in the caudal medulla, function as sensory circumventricular organs (SCVOs) . By virtThe RAS plays a fundamental role in the regulation of BP and hydromineral homeostasis. ANG II generated by angiotensin converting enzyme (ACE) 1 acts on ANG type 1 receptors (AT1-R) to produce its major physiological and pathophysiological effects, including direct acting vasoconstriction, baroreflex modulation, sympathetic activation, aldosterone release, triggering oxidative stress, and inflammation . ComponeThe metabolic cascade of the systemic RAS (aka as the circulating or systemic RAS), involves renal renin release to act on renin substrate to generate ANG I and then via ACE1 action, ANG II. Following the elucidation of the components of the classic RAS, comparable elements were identified in many body organs including the brain. The so-called brain RAS is associated with neurons, astrocytes and microglia functions in a paracrine, autocrine, and even intracrine signaling role . Many CNRecent studies demonstrate that either AT2-R or ACE2/ANG-(1-7)/Mas-R are present either within or adjacent to cardiovascular nuclei in the brainstem and hypothalamus that express the AT1-R . FunctioRecent studies demonstrate that ACE2 is expressed on GABAergic neurons that in turn synapse on excitatory neurons within the hypothalamus . ADAM17,2, which in turn serves as a humoral mediator to communicate with surrounding tissue in the mode of a volume transmitter  or PIC administration induces enhanced expression of PICs in the brain. Because PICs are large lipophobic polypeptides, they are unlikely to pass the blood-brain barrier. Therefore, as in the case of classic brain RAS communication, the mode of body-brain signaling is worthy of consideration. Current evidence indicates that there are five putative means of informing the brain of systemic inflammation and elevated peripheral PICs. First, sensory circumventricular organs, most notably the OVLT have been implicated in the mediation of fever. There is evidence to indicate that the pyrogenic effect of interleukin (IL)-6 is produced by action on the OVLT to release prostaglandin Ensmitter . Second,nsmitter . Third, activity . Fourth,activity . Fifth, activity .The brain has its own innate immune system, which involves microglia as the resident immune cell. Recent studies indicate that microglia and astrocytes in the brainstem and hypothalamus participate in orchestrating many cardiovascular and metabolic functions . Glial cIncreased local production of ANG II is one of the main mechanisms responsible for the development and progression of hypertension. Apart from the control of SNS activity and extracellular volume regulation, ANG II also acts as a stimulator of PICs and promotes activation of the brain innate immune system . ConversToll-like receptors are pathogen receptors, which are a major component of the innate immune system . A growiNF-κB is a key regulator of PIC expression and the inflammatory response observed in hypertension. In ANG II-induced hypertension, peripheral ANG II infusion increases PICs, ROS, and prohypertensive RAS components (ACE1 and AT1-R) and decreases the antihypertensive components (ACE2 and Mas-R) within the PVN. Bilateral PVN NF-κB blockade decreases the prohypertensive RAS axis and increases the protective antihypertensive RAS axis through an ROS-mediated mechanism. The result is an attenuation of the ANG II-induced hypertensive response . FurtherEndoplasmic reticulum (ER) stress and autophagy are also implicated in the development of hypertension. In ANG II-induced hypertensive mice there is upregulation of many members of the ER stress pathway in the SFO. Central inhibition of ER stress prevented ANG II-induced hypertension . LikewisRecent studies demonstrate that blood-borne PICs induce a pressor response and sympathetic activation through acting on the SFO to increase RAS activity and inflammation. Blockade of AT1-R or ACE or knockdown of AT1-R in the SFO attenuates those PIC-elicited responses . Tumor nCollectively, the studies just described indicate that crosstalk between the RAS and TLR4/NF-κB/TNF-α as well as ER stress and autophagy within the brain tips the balance of the RAS in favor of the prohypertensive axis while decreasing the antihypertensive axis thereby enhancing cardiovascular inflammation. This process results in triggering positive feedback from periphery to brain to drive a progressive increase in sympathetic tone and the pathogenesis of hypertension.per se or consuming a high fat diet (HFD) produces a chronic low-grade inflammatory state characterized by activation of microglia, astrocytes, and increased expression of PICs in the CNS. The mediators of the brain innate immune system, including TLR4, IκB kinase-β/NF-κB (IKKβ/NF-κB), ER stress and autophagy, are involved in obesity-related inflammatory signaling  pathway . The CNSThere are many examples of changes in behavioral and physiological responses resulting from prior experience. The impact of exposure to certain stimuli or physiological states can last a lifetime. When a response entails an increase in magnitude because of an antecedent condition, it is considered as an example of sensitization . SensitiWe found that peripheral or central pretreatment with non-pressor doses of ANG II, aldosterone, or TNF-α sensitized the hypertensive response to a slow-pressor dose of ANG II. At the end of DEL the mild challenges used for IND upregulated mRNA expression of several components of the RAS and PICs and of microglial marker in the brain, whereas central inhibition of AT1-R, mineralocorticoid receptors, and inflammation reversed the changes in gene expression and also prevented the sensitization of ANG II-elicited hypertension seen during EXP. Importantly, the results showed that either central blockade of the RAS or central blockade of inflammation during IND attenuated both the upregulated expression of RAS components and PICs regardless of the mild challenges used for IND ,b. Such In studies of dietary sensitization, 3 weeks of HFD feeding increased white adipose tissue mass, plasma leptin levels and mRNA expression of leptin and leptin receptor in both the LT and PVN . CentralDespite the prohypertensive axis of the RAS playing a predominant role in the sensitization of hypertension, the antihypertensive axis of the RAS has a protective effect against the sensitization process. This is based on its counter-regulatory effects on hypertension. In our studies, low doses of ANG II or aldosterone infusion during IND not only enhances ANG II-induced increases in the expression of renin, AT1-R, and ACE mRNA, but also upregulates mRNA expression of ACE2 and AT2-R in forebrain nuclei , b. FurtTaken together, our studies provide insight into the actions of the RAS and of PICs in the hypothalamus to mediate cardiovascular sensitizing effects of mild physiological or metabolic challenges, even though these factors do not induce any apparent immediate abnormalities in the cardiovascular function. The demonstrations of sensitization also indicate that challenges to homeostasis can act to reprogram the neural network controlling BP to generate an enhanced pressor response when a hypertensive stimulus is either sustained or encountered at a point later in life  Figure .There is a strong association between maternal health during pregnancy and cardiovascular disease in adult offspring. Many kinds of prenatal insult such as hypertension, obesity or HFD, protein restriction, uteroplacental perfusion insufficiency or glucocorticoid treatment during pregnancy have been demonstrated to have consequences on health of the offspring. Such prenatal challenges alter responses to environmental stressors and predispose human and experimental animals to cardiovascular and metabolic disorders .Increased RSNA and overactivity of the RAS and PICs have been implicated as causal mechanisms in prenatal programming of hypertension . Either The prevalence of obesity in women of childbearing age and during pregnancy has steadily increased over the past two decades . Prior eMaternal separation during early life, so called early life stress, sensitizes rats to ANG II-induced hypertension in adult life. Early life stress offspring displayed renal dysfunction, elevated renal and systemic sympathetic activity, enhanced ANG II-mediated vasoconstriction and vascular inflammation and reduced endothelial nitric oxide buffering capacity , 2013a,bOur studies investigating the effects of gestational maternal challenges on the offspring when they reach adulthood have focused on the role of reprogramming central mechanisms involving the RAS and PICs on SNA. This was done by inducing sensitization of the offspring during the prenatal period by subjecting pregnant dams to ANG II-elicited hypertension or to HFD feeding . In the Sex-specific effects are widely observed in perinatal developmental programming studies and female offspring are protected as compared to male offspring . We alsoTaken together, these experiments demonstrate that reprogramming of the mechanisms controlling BP during a sensitive prenatal period produces long-lasting phenotypic changes in the CNS and increased responsiveness to a prohypertensive challenge .In this review, we have described the roles of the RAS and inflammation in the CNS in the development of hypertension. These two systems, either independently or synergistically, activate brain nuclei involved in BP regulation to increase SNS drive that in turn, generates renin-angiotensin-elicited or obesity-related hypertension. We further highlighted the interactions of the RAS and inflammation in the HTRS.The predisposition of the offspring of mothers exposed to various insults during pregnancy to display HTRS appears to be very similar to sensitization of this response produced by prior challenges that were found in adult animals. The permanence of HTRS is associated with CNS reprogramming instantiated as changes in expression of RAS components and of PICs. The studies on the new models of sensitization provide many possibilities to elucidate central mechanisms underlying the long-term maintenance of sensitization and the etiology of hypertension.Because maternal hypertension and HFD/obesity are common and rapidly increasing, it is likely that future generations will be at increased risk for cardiovascular diseases including hypertension. Because only half of hypertensive patients show a satisfactory response to conventional treatment , it is cBX conceived, drafted, revised, and approved the manuscript. YZ revised and approved the manuscript. AJ conceived, edited, revised, and approved the manuscript. All authors contributed to the article and approved the submitted version.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


However, it remains largely unclear whether the abstract pattern of spatial arrangement of radicals can be rapidly extracted, and if so, whether this extraction takes place at the visual cortex, the very first processing center. As the visual cortex is documented to actively participate in orthographic processing, it may also play a role in the possible extraction of abstract orthographic patterns of Chinese characters. Hence, we hypothesize that abstract orthographic patterns of Chinese characters are covertly extracted at the visual cortex during reading. In this study, we investigated whether the visual cortex could rapidly extract abstract structural patterns of Chinese characters, using the event-related potential (ERP) technique. We adopted an active oddball paradigm with two types of deviant stimuli different only in one feature, structural or tonal, from standard stimuli; in each of the two sessions, subjects focused conscious attention on one feature and neglected the other. We observed that the ERPs recorded at occipital electrodes responded differentially to standard and structural deviant stimuli in both sessions, especially within the time range of the occipital P200 component. Then, we extracted three source waves arising from different levels of the visual cortex. Early response differences (from 88 to 456 ms after stimulus onset) were observed between the source waves, probably arising from left primary/secondary and bilateral associative visual cortices, in response to standard and deviant stimuli that violated abstract structural patterns, whether subjects focused their attention on the character structure or not. This suggests rapid extraction of abstract structural patterns of Chinese characters in the visual cortex, no matter the abstract structural pattern was explicit or implicit to subjects. Note that the source waves arising from right primary/secondary visual cortices in response to standard and structural deviant stimuli did not differ at all, indicating that this extraction of the abstract structural pattern of Chinese characters was left-lateralized. Besides, no difference was observed between source waves originating from any level of the visual cortex to standard and deviant stimuli that violated abstract tonal patterns, until 768 ms when a late effect related to conscious detection of targets occurred at higher levels of the visual cortex. Note that at late stages (later than 698 ms after stimulus onset), responses arising from bilateral associative visual cortices to standard and target stimuli differed for both sessions, no matter the structural or tonal feature was attended to. Our findings support the primitive intelligence of visual cortex to rapidly extract abstract orthographic patterns of Chinese characters that might be engaged in further lexical processing. Our findings also suggest that this rapid extraction can take place implicitly during reading.Orthographic processing is crucial in reading. For the Chinese language, sub-lexical processing has already taken place at radical level. Previous literature reported early position-specific radical representations and later position-general radical representations, implying a possible separating process of abstract position information irrespective of radicals  As a critical carrier of communications among humans, the written language is considerably variable. It is marvelous for humans to be able to perceive written scripts and extract relevant features fast ,2. DurinIt is a key step to combine units of written scripts into a whole during the lexical processes before word representations are formed . For exaRadical position information seems to be encoded and represented during orthographic processing. Although behavioral studies provide evidence supporting the emergence of both position-specific and position-general radical representations in Chinese character recognition ,12, the per se and representations of abstract radical positions. In alphabetic languages, the separation of letter identity and within-word letter position is crucial for subsequent semantic and phonological processing; the letter identity assists semantic extraction and the position information is necessary for representations of within-word-position-coded letter identities that will be utilized in phonological processing [per se and abstract radical positions may be more complex than in alphabetic languages. The role of the abstracted position information in orthographic processing of Chinese characters, to our surprise, was investigated by few studies. Representing the way that radicals of Chinese characters are combined, the abstract position information irrespective of radicals per se indeed affects the recognition of Chinese characters during reading. For instance, characters with left-right structure tend to be perceived faster than those with up-down structure [per se and representations of abstract radical positions, the impact of the spatial arrangement of radicals on the recognition of characters, and the fact that the literate brain of Chinese speakers is continuously learning the statistics of the linguistic surroundings lead us to the hypothesis that the abstract pattern of spatial arrangement of radicals in Chinese characters are extracted during reading, probably in an implicit way.The emergence order of position-specific and position-general radical representations inevitably suggests a possible abstracting process that isolates representations of radicals ocessing . Since tocessing ,8, the structure . In addiper se is extracted and represented during Chinese character recognition. As a part of orthographic processing, previous literature on the time course of orthographic processing for both alphabetic [per se and their positions, if existing, is likely to be involved as well in the extraction of the abstract spatial pattern of radical configuration. Previous literature also observed activations of a group of brain regions including fusiform gyrus and middle occipital gyrus  specifically during orthographic processing [To our knowledge, it remains largely unclear whether the abstract pattern of spatial arrangement of radicals irrespective of the radicals phabetic  and logophabetic ,21 makesphabetic ,16, sincocessing ,23, whicIn this study, we attempted to explore whether the abstract pattern of spatial arrangement of radicals could be implicitly extracted during reading. We decided to use the ERP technique in order to track the time course of this covert procedure during orthographic possessing of Chinese characters. We were most interested in an ERP component named the visual mismatch negativity (vMMN), a response originating from the visual cortex and reflecting its predictive error of the statistics of the visual surroundings . From anImportantly, the vMMN can also be elicited by violations of the regularity of lexical information. For instance, Wei et al. reported significant vMMN effects for surprising alternations between non-characters and real characters/pseudo-characters with legally-positioned radicals, among native Chinese speakers but not non-native speakers, suggesting that the brain of native Chinese speakers is capable of extracting lexical features with an early-triggered automatic process . Their fHere, we adopted an active dual-deviant oddball paradigm, simulating a scene of reading. To be more specific, the visual stimulus stream used in this experiment was composed of three types of Chinese characters: those with left-right structure and falling tone, those with left-right structure and rising tone, and those with up-down structure and falling tone (probability: 8:1:1). In one of the two sessions in the experiment, subjects were instructed to respond to the target characters with a deviant structure (up-down) from the standard one (left-right). By doing so, subjects explicitly identified the violations of the standard abstract pattern of spatial arrangement of radicals in this session. We expected to observe differences within in the time range of the vMMN between ERPs in response to standard stimuli and target ones, which represented the active extraction of the abstract structural pattern. In the other session with exactly the same stimulus presentation, subjects were instructed to respond to the target characters with a deviant tone (rising) from the standard one . Then, we examined whether there were also differences within in the time range of the vMMN between ERPs in response to standard stimuli and non-target ones with a different structure that were targets in the structure identification session. We expected to observe similar difference effects on ERPs as in the structure identification session; if so, this meant that the differences in ERPs in response to changes in the abstract structure pattern of Chinese characters could be extracted implicitly during reading.This paradigm was also used to replicate the findings that abstract phonological patterns of Chinese characters are implicitly extracted in the visual cortex ,32. In tSixteen subjects  participated in the experiment. All subjects were native Chinese speakers with normal or corrected-to-normal vision. None of the subjects had history of neurological or psychiatric disorders. All subjects were paid for their time and gave written informed consent to the experimental protocol approved by the local Ethics Committee of Tsinghua University.We used an active oddball paradigm with two types of deviant stimuli, simulating a scene of reading . In a sthttp://corpus.zhonghuayuwen.org). The final images of stacking-up stimulus characters for the three stimulus types were considerably similar, suggesting minimal physical heterogenicity of visual stimuli (The stream contained 1200 characters: 960 standard stimuli (LR4), 120 structural deviant stimuli (UD4), and 120 tonal deviant stimuli (LR2). Owing to the limitation of the number of real characters with left-right structure and falling tone, the stimulus pool contained 524 standard characters and 436 of them were presented twice. The characters were presented in a pseudo-random order, any two neighboring deviant stimuli were separated by at least two standard stimuli, and the standard characters that were delivered twice were not presented consecutively. All the stimuli were in white color, displayed at the center of a gray background. Subjects’ eyes were at the same height as the center of the screen and the eye-to-screen distance was ~1 m. The width and height of the Chinese characters were about 10 cm; they were all in Imitation Song typeface. The occurrence frequency and the stroke number of the characters were matched across stimulus types ; the occ stimuli . The dur stimuli . The delThe experiment included two sessions where exactly the same stream of characters was presented. In one session, subjects were instructed to respond to one type of deviant stimuli by pressing a button and neglect the other type; in the other session, they were instructed to respond to the other type of deviant stimuli. The session order was counterbalanced across subjects. As shown in During data acquisition, subjects were comfortably seated in an armchair. They were instructed to press a button in response to each target stimulus as quickly as possible. While they were doing the tasks, continuous electroencephalography (EEG) data were being recorded at a sampling rate of 500 Hz with a BrainAmp DC amplifier (Brain Products GmbH) and the software Recorder , using Ag/AgCl electrodes, simultaneously from 61 channels  according to the international 10/10 system, with the reference electrode placed at FCz and the ground electrode placed at AFz. Vertical and horizontal electro-oculograms were also recorded for elimination of ocular artifacts. Electrode impedance was maintained below 10 kΩ.A hit was recorded when the subject pressed the response button between 200 to 1400 ms after the onset of a target stimulus. The hit rate is the ratio of the number of hits to the number of target stimuli. The reaction time is the median time across trials between stimulus onset and the first hit. The indices were calculated in Matlab  and compared between sessions with paired t-tests in SPSS ; a priori significance level was set at 0.05.The pre-processing of event-related potentials (ERPs) included re-referencing, filtering, artifact elimination, bad-block rejection, segmenting, and averaging. First, the continuous EEG data were re-referenced to the average of all channels, and band-pass filtered between 1 and 70 Hz. Next, ocular and myogenic artifacts were largely eliminated with a data cleaning toolbox implanted in EEGLAB  on the bp values were false discovery rate (FDR) corrected in order to control the overall Type I error rate. We also compared the ERPs in response to the standard and the target deviant stimuli, using the same analyzing approaches.We compared the ERPs in response to the standard and deviant stimuli with paired t-tests. The comparison was first conducted in sensor space; the ERPs recorded at occipital regions  were averaged and further analyzed. A sliding window with duration of 40 ms moved with 2-ms step and the mean amplitude was compared between responses to the standard and the unattended deviant stimuli for each time point with paired t-tests. The In order to compare the ERPs in response to different types of stimuli in source space, we extracted six components from the averaged ERPs using spatial filters that were obtained via the independent component analysis (ICA) . The numIn order to localize the neural sources for each spatial filter, we applied dipole fitting to the components derived from the ICA-based decomposition with DIPFIT 3.3 toolbox implanted in EEGLAB. We used a boundary element model derived from the MNI  averaged brain as the head model. The location, orientation, and strength of a couple of mirrored rotating dipoles  were obtained through a scanning procedure aimed at minimizing the deviation between the estimated topography and the measured one . The locAfter the source waves were derived, we compared the ERPs in response to the standard and deviant stimuli in source space with paired t-tests. The comparing approach was the same as that for ERPs in sensor space. The ERP pre-processing, the ICA-based decomposition of ERPs, and the dipole fitting were conducted with the EEGLAB toolbox in Matlab, the eLORETA was conducted in the LORETA-KEY software package , and senp < 0.001). The mean hit rate was 94% for TaskS and 86% for TaskT (p < 0.001). This suggests that the tone identification task was more difficult than the structure identification task, ensuring the extraction of abstract structural pattern in TaskT to be implicit.The mean reaction time was 684 ms for TaskS and 839 ms for TaskT . A paireor TaskT . A paireWe observed significant differences between occipital ERPs in response to standard (LR4) and target deviant stimuli (UD4) in TaskS. As shown in We observed significant differences between occipital ERPs in response to standard (LR4) and target deviant stimuli (LR2) in TaskT. As shown in We extracted six components that explained 95.18% of the whole data. Based on visual inspection, their topographies suggest that three components arose from the occipital lobe  and target deviant stimuli (UD4) in TaskS. The time range when the differences occurred was 248–332 ms (target deviant > standard) and 334–456 ms (target deviant < standard) for IC5 , and 120In contrast, there was no significant difference between source waves arising from the visual cortex elicited by standard (LR4) and target deviant stimuli (LR2) in TaskT, except the significant differences between IC1 elicited by standard (LR4) and target deviant stimuli (LR2) in the time range of 768–960 ms (target deviant > standard), which was quite late. There was no significant difference between source waves arising from the visual cortex elicited by standard (LR4) and non-target deviant stimuli (LR2) in TaskS . These fWhen attentional resources were consciously and actively devoted to detect the target stimuli, there were significant differences between source waves representing higher-level processes (IC3 and IC2) elicited by standard and target deviant stimuli in both TaskS and TaskT. For IC3, whose sources were in the frontal lobe, the time range when the differences occurred was 360–520 ms (target deviant > standard) in TaskS and 558–756 ms (target deviant > standard) in TaskT . For IC2Interestingly, we also observed significant differences between IC1 elicited by standard and target deviant stimuli at late stages of both TaskS and TaskT. The time range when the differences occurred was 484–540 ms (target deviant < standard), and 698–776 ms (target deviant > standard) in TaskS, and 768–960 ms (target deviant > standard) in TaskT . This suIn this study, we investigated the possible extraction of abstract structural patterns of Chinese characters in the visual cortex, by examining whether the ERPs, especially the source waves arising from the visual cortex, changed in case of violations of explicit and implicit abstract structural patterns. Early response differences were observed between 88 and 456 ms after stimulus onset at different levels of the visual cortex, probably the left primary/secondary visual cortices and the bilateral associative visual cortices, whether the abstract structural pattern was explicit or implicit to subjects. On the other hand, we did not observe early response difference in the visual cortex for abstract tonal patterns. Moreover, response differences later than 698 ms after stimulus onset were also observed in the bilateral associative visual cortices for both explicit abstract structural and tonal patterns. These findings suggest that the visual cortex could rapidly extract abstract structural patterns of Chinese characters, whether or not conscious attention was focused on the target feature. Besides, the extraction of abstract tonal patterns was still feasible in the visual cortex at a relatively late stage if attentional resources were sufficiently devoted.The differences between ERPs elicited by explicit and implicit changes in the abstract structural pattern were observed both at the sensor level and at the source level. In the sensor space, we selected occipital electrodes mainly because the responses of the visual cortex were of our primary interest. We observed ERP differences, most significantly in the time range of the P200 component, for both explicit and implicit violations of the abstract structural pattern, corresponding to the vMMN. However, there were still several other time ranges in which the ERP differences were also significant. Owing to the volume conduction of neurophysiological signals, it was not easy to convincingly attribute the ERPs recorded at these occipital electrodes to the brain region under them. That is to say, although we are confident to assume that the occipital ERPs mainly represents the neural activity of the visual cortex, especially at the latencies when typical visual ERP waves occurs, chances are that the scalp responses are dominated by cortices other than the visual cortex within some time periods. When ERPs in response to standard and deviant stimuli differed significantly, we relied on the topography of mean amplitude to examine whether the effects reflected certain processes that actually took place in the visual cortex or simply were products of volume conduction of processes that took place elsewhere. For example, the ERPs at occipital electrodes in response to LR4 and UD4 in TaskS differed around 638–756 ms, but it was almost unlikely that this difference effect reflected neural activities in the visual cortex, as the topographies of mean amplitude within this time range showed concentrated power around Pz, suggesting volume conduction of the P300 component. Therefore, it is necessary to conduct subsequent source-level analyses to clarify what brain regions the ERP difference effects could be attributed to and when these effects actually began and ended at their respective neural sources.Using ICA-based space filters, we extracted three source waves arising from the visual cortex. Although the source reconstruction results based on dipole fitting and eLORETA were not exactly the same, they both supported that IC4 and IC5 arose from relatively lower levels of the visual cortex , whereas IC1 arose from higher levels of the visual system . We found evidence supporting that abstract structural patterns could be rapidly and implicitly extracted in the visual cortex, as a part of the orthographic processing of Chinese characters. We observed significant differences between IC1 in response to the standard (LR4) and the target deviant stimuli (UD4) in TaskS (before 374 ms). Moreover, significant differences between IC1 in response to the standard (LR4) and the non-target deviant stimuli (UD4) were also observed in TaskT (before 356 ms). Similar response differences were also observed for IC5 (before 456 ms), but not for IC4. These findings suggest that both explicit and implicit abstract structural patterns of Chinese characters can be extracted rapidly in the visual cortex, and that this rapid extraction is lateralized to the left hemisphere.The sensory cortex is considered to have primitive or sensory intelligence that extracts abstract stimulus patterns from ongoing sequence . The audper se. Note that one advantage of the current study over previous ones is that we have provided evidence documenting where in the brain the ERP effects originated and pinpointed the neural sources underlying the ERP effects to the visual cortex. Despite the relatively poor spatial resolution of the ERP technique, the current study uses multiple source localization methods and obtains resembling results on the generating sites of the ERP difference effects, providing converging evidence in favor of the vital role of the visual cortex in the extraction of abstract structural patterns of Chinese characters during orthographic processing.This primitive intelligence might be relevant to the processing underlying the emergence of position-specific and position-general radical representations. Previous literature documents that early radical representations are position-specific, but position-general radical representations also appears later , implyinWe did not observe evidence supporting the detection of violations of implicit abstract patterns of lexical tone in the visual system at low levels. This seems inconsistent with previous studies documenting a rapid, automatic detection of violations of abstract patterns of Chinese character phonology ,32. The There are two limitations of this study that may affect the generalization of our findings. First, our use of an active oddball paradigm, together with the relatively long presentation time of the stimulus characters, renders it difficult for us to examine the role of attention in the extraction of the abstract structural pattern of Chinese characters. Although the dual-deviant oddball paradigm ensures that the ERP differences we found are not task-specific or target-specific, it is impossible for us to examine whether the rapid and implicit extraction of abstract structural patterns of Chinese characters was automatic and pre-attentive. After all, subjects paid conscious attention to all the characters in the character stream but focused on different features in different sessions. However, as an exploratory investigation, this study provides solid evidence supporting the implicit extraction of abstract structural patterns of Chinese characters. Whether this extraction is automatic and pre-attentive would be investigated in future studies using a passive oddball paradigm, e.g. a common passive paradigm used in vMMN studies where characters are presented peripherally and subjects pay attention to an irrelevant visual task at the center ,32.Second, it is not clear whether our findings may be generalized to structures of Chinese characters other than left-right structure or up-down structure. However, even if the results of this study failed to be generalized to other types of Chinese character structure and Chinese speakers could only differentiate characters with left-right structure from characters with up-down structure at the level of visual cortex, it would still be appropriate for us to conclude that the extraction of the abstract structural pattern of Chinese characters has already started in the visual cortex. The core question here is what caused the ERP differences we observed in this study. They cannot be induced due to the differences in familiarity of the characters, as we controlled the occurrence frequency of the stimulus characters to ensure they were matched across stimulus types. We cannot rule out the possibility that they are induced by the differences in familiarity of the abstract structural patterns. This could be examined in future studies where the structure familiarity difference between standard and deviant stimuli is manipulated so that it could be examined whether the ERP differences change with structure familiarity correspondingly. However, since it is likely that the visual system of Chinese speakers learns to extract the abstract structural pattern of Chinese characters during their long-term linguistic experience, it is possible that the differences in occurrence frequency of various structures determine, at least partially, the learning at the visual cortex and consequently the exact extraction strategy heavily depends on the familiarity.In conclusion, this study documents that the visual cortex is able to extract abstract patterns of spatial arrangement of radicals (i.e. structure) of Chinese characters rapidly, no matter the abstract pattern is explicit or implicit to readers. The left primary/secondary visual cortices and the bilateral associative visual cortices are probably involved in this left-lateralized extraction. Nevertheless, we have not found evidence in favor of a similar rapid extraction in the visual cortex of abstract patterns of the lexical tone of Chinese characters. These findings suggest that the primitive intelligence of the visual cortex may be restricted to extract abstract patterns of limited features of Chinese characters.


Autophagy is a catabolic process that takes place under both normal and adverse conditions and is important for the degradation of various organelles and proteins that are no longer needed. Thus, it can be viewed as both a constitutive recycling machinery and an adaptation mechanism. Increase in the activity of autophagy can be caused by multiple biotic and abiotic stress factors. Though intensive research in the past decade has elucidated many molecular details of plant autophagy, the mechanisms of induction and regulation of the process remain understudied. Here, we discuss the role of ATG8 proteins in autophagic signaling and regulation with an emphasis on the significance of ATG8 diversification for adapting autophagy to the changing needs of plants. Plants are unable to escape from unfavorable environmental conditions or damaging interactions with other organisms. Hence, in order to survive, they need to adapt to the changes in their surroundings as fast as possible. Due to these characteristics, it is important for plants to efficiently acquire essential elements, produce and reuse metabolites, and optimize energy consumption by recycling of cellular components.Saccharomyces cerevisiae identified the main players in autophagy and revealed the detailed molecular mechanisms of autophagic degradation  and phytohormones can influence the activity of autophagy and ATG proteins in plants in a TOR-dependent manner. Amino acid availability and abundance of their degradation products affect metabolic and developmental processes  includinArabidopsis thaliana although the mechanisms are unclear 1, promoting its own selective degradation ] or Ubiquitin-interacting motif  . UDS-intith ATG8 . Autopharadation . It is uin vivo and in planta data are acquired, in the next section we use existing resources such as available plant genomes together with transcriptomics and proteomics data to explore ATG8 specificities in the plant kingdom.Given the diversity of ATG8s in higher plants, it is difficult to conclude on the importance of each iso-form based on single knockout mutants due to functional redundancy. Different ATG8s appear to be induced by varying stress factors or to alleviate them although some overlap has been discovered . The abiATG8 proteins have shown a tendency toward increase in number of iso-forms and diversification in multicellular organisms. Yeast and algae have a single homolog while in the genomes of higher plants, multiple homologs can be found . The ninZea mays), none of the four ATG8s belong to the ATG8h-i group, while one of the four rice (Oryza sativa) homologs is cleavage-free. Additionally, while Medicago (Medicago truncatula) and soybean (Glycine max) have two ATG8h-i iso-forms each, only one in Medicago is cleavage-free. The rapeseed (Brassica napus) has six h-i homologs, all ending with a glycine (G) and as the plant is a recent alopolyploid    and memb-i group . Whether-i group  and in r-i group . HoweverATG8 genes, namely ATG8a, b, e, and h  likely through ROS signaling or direct binding to their promoters  by yeast one-hybrid screen able to bind the promoters of four e, and h  and thusbidopsis , most libidopsis  (Figure bidopsis . ET is aromoters . It is hATG8 genes seem to be expressed in every analyzed tissue (Apart from the evidence for differential transcriptional regulation of the ATG8 homologs, differences in protein levels have recently been documented by large scale quantitative transcriptome and proteome analysis of 30 different tissues . While ad tissue , only Atd tissue . Notablyd tissue  (Mergnerd tissue . The AtAThe presence of multiple regulation targets providing grounds for activity modulation of autophagy could be considered as an indicator that such intricate regulatory system is in fact necessary. It is without a doubt, that understanding the molecules and signals affecting the autophagy machinery would contribute to the elucidation of autophagy induction and regulation.Arabidopsis thaliana, identifying potential upstream and downstream components of the TOR network (One of the most upstream targets of autophagy modulation, the TOR kinase, has recently featured in a phosphoproteomic screen combined with targeted proteomics analysis of interacting proteins, in  network  that couIncrease in number of iso-forms and diversification of ATG8 in plants along the course of evolution might reflect the need to accommodate the requirements of the increased complexity of flowering species. It is possible that iso-form specific or clade-specific ATG8 functions exist or that certain homologs are more abundant in autophagosomes under given conditions, in specific tissues and organs or during certain developmental stages. Additionally, exploring the specificity of the ATG8 interactions and the significance of the cleavage free iso-forms, also by obtaining more functional data could further clarify the diversity of the ATG8 homologs in plants and contribute to the understanding of autophagy induction and regulation. In this line of thought, clarifying the molecular mechanisms regulating ATG4 and ATG8 would provide more insight into the process of fine-tuning autophagy.SBW provided the initial draft while both SBW and EI were involved in the further editing of the manuscript and the preparation of the figures. Both authors conceptualized the idea.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


Adopting effective drowning prevention measures are dependent on collecting data on drowning rate, since drowning prevention requires the planning and policy-making on known risk factors. When interventions are carried out, data are still required to monitor and evaluate the evaluation of the strategy. Therefore, recording drowning cases accurately can be effective in its preventing. The purpose of this study was to provide the challenges of implementing a drowning registry.Various sessions were held with attendance of the experts from the related provincial organizations such as Red Crescent, EMS, Lifeguard, and Governor General Office, Ports and Shipping, among others, as well as the health experts from medical universities. The existing challenges were recognized using qualitative research and semi-structured interview method. All related challenges were mentioned and prepared through qualitative interviews with twenty experts. Finally, they were analyzed utilizing MAXQDA software.The most important problems in implementing the drowning registration system included:1- Lack of enough time to collect data on the registration system when providing emergency services on the beach2- Lack of proper map and GPS system in identifying drowning locations 3- Lack of a single responsible organization for dealing with the beaches’ issues4- Lack of correspondence in organization involved in drowning in northern coastal provinces5- Lack of sufficient information about important variables related to the registration system6- The absence of the rescue forces in some areas other than the sea protected areas and the lack of access to clinical data on drowning in these zones at night hours7- Need for more cooperation by the Forensic Medicine Organizations of provinces to provide data The drowning registration system has many challenges that can mostly be solved by providing appropriate funding and right registering of drowning and purchasing appropriate equipment.Drowning registration, Challenges, Coastal provinces


Cryptococcosis is an important opportunistic infection and a leading cause of meningitis in patients with HIV infection. The antifungal pharmacological treatment is limited to amphotericin B, fluconazole and 5- flucytosine. In addition to the limited pharmacological options, the high toxicity, increased resistance rate and difficulty of the currently available antifungal molecules to cross the blood–brain barrier hamper the treatment. Thus, the search for new alternatives for the treatment of cryptococcal meningitis is extremely necessary. In this review, we describe the therapeutic strategies currently available, discuss new molecules with antifungal potential in different phases of clinical trials and in advanced pre-clinical phase, and examine drug nanocarriers to improve delivery to the central nervous system. Cryptococcus infection and development of cryptococcosis in humans generally occurs due to inhalation of yeasts or spores of Cryptococcus neoformans and Cryptococcus gattii present in the environment ) based micellar drug delivery system loaded with AMB (Angiopep-PEG-PE/AMB) was more effectively transported across the BBB than the deoxycholate AMB and the micelle without angiopep-2 in in vitro and in vivo assays. This superiority was attributed to angiopep-2 being a ligand of low-density lipoprotein receptor-related protein (LRP) present in the brain, which has been reported to mediate the transport of ligands across endothelial cells of the BBB . When AnITC and its derivative compound POS, both triazole agents, are acceptable alternatives for treatment of the pulmonary form of cryptococcosis, but not for cryptococcal meningitis due to its difficulty of crossing the BBB . ĆURIĆ aSearching for new alternatives for treatment of fungal infections has been pivotal. Here, we highlighted the remarkable deficit of antifungal options in the treatment of cryptococcosis recommended by the current guidelines. Studies have been carried out with the intention of finding new therapeutic alternatives as described in this work; some of these molecules and the use of nanocarriers to improve drug delivery into the CNS may become future therapies for cryptococcal meningitis.