Objective: The purpose of this study was to explore the rate of prolonged grief disorder (PGD) and associated factors in a large sample of diverse college students. Sources of grief support and perceived helpfulness of support were also examined.Method: An online survey was administered to bereaved students at three colleges at the City University of New York. PGD measured by the Inventory of Complicated Grief was the primary outcome. Chi-squared and t-tests were used to assess the association between PGD and associated factors.Results: A total of n = 899 participants completed the Inventory of Complicated Grief (ICG) based on a significant death loss = >12 months. An estimated 13.4% (n = 120/899) met criteria for PGD. The rate of PGD was associated with race, history of anxiety or depression, trauma other than the death, insecure attachment style, kinship to the deceased, closeness to the deceased, cause of death, and sudden/unexpected death. The majority of students sought grief support from a friend or family member.Conclusion: The rate of PGD in this sample of college students is similar to that of adults and most prevalent for students of color. Identification of those most at risk is critical to referring these students to effective treatments. The death of a loved one is one of the most difficult life experiences and often triggers a grief reaction with intense and painful emotions. While acute grief usually dissipates over time and becomes integrated, a sizeable minority of people has ongoing difficulty after the loss. Severe and prolonged grief symptomatology is designated as Persistent Complex Bereavement Disorder (PCBD) in the DSM-5 . ProposeOne third of traditional age college students (average age 18–23) in the United States have lost at least one family member or friend in the past year  and nearConsequences of PGD for college students include lack of concentration and focus, lack of motivation, sleep difficulties, isolation from family and friends, suicidal ideation, poor grades, and possible dropout or dismissal, in addition to mental and physical health problems common to adults with PGD such as anxiety, depression, and chronic illness . PGD canLimited research has been conducted on the prevalence of PGD in college students and these studies have significant limitations including small sample sizes, overly restrictive inclusion criteria (limiting bereavement to the past 2 years) and a lack of diverse samples.n = 2/46) of predominantly white college students could be classified as having PGD using the 13-item Inventory of Traumatic Grief  of bereaved college students from the United Arab Emirates and the Southeast United States met criteria for PGD utilizing the Prolonged Grief Questionnaire  of graduate students in the Southeast Unites States met criteria for PGD utilizing the PG-13. All three of these studies defined bereavement as experiencing a death-loss within the past 2 years.Prevalence rates between studies are difficult to compare due to the use of different instruments and different criteria for measuring PGD. However, the rates of PGD in college students reported in prior studies are significantly lower than rates found in adults. ief ITG; . Varga ee PG-13; . Varga (n = 27/226) 12% met criteria for PGD. The highest rate of PGD was found by n = 46/326) 14% met criteria for PGD, however, time since loss was not specified.Lifetime death-losses were included in one study by The vast majority of studies on bereavement in American college students have used samples that were predominantly white. Only a handful of studies used more diverse samples and none of them reported the prevalence of PGD. However, among 1,581 bereaved students (40% Black) enrolled at a large Southern University, In addition to race , prior rFactors associated with PGD among college students, particularly students of color have been underexamined. An examination of the rate of PGD in a more diverse college student sample might yield important information about the grief experiences of this population. Based on the findings reported by Researchers in the field of complicated/prolonged grief assert that problematic thoughts, such as ruminations about why or how the person died or behaviors such as avoidance of reminders of the loss can interfere with the normal healing process, resulting in a prolonged and intense state of acute grief . CertainThe purpose of this exploratory study is 3-fold: (1) to identify the rate of PGD in a large sample of diverse college students, (2) to examine the relationship between PGD and potential risk factors, and (3) to identify sources of grief support used by these students and perceptions of the helpfulness of these supports.The sample comprised 974 students attending three colleges at a public university in a large northeastern metropolitan area during the 2019–2020 academic year. Internet survey data were collected over two semesters (at a single time point for each participant) through the college research pool for course credit. Participants had to be 18 years or older and have experienced the death of a family member or close friend at any point in their life. Students indicated their consent to participate after reading a description of the study on-line. Participants were provided with a list of grief resources at the end of the survey. The university’s institutional review board approved the study.Socio-demographic characteristics were measured by a questionnaire that included age, gender identity, race/ethnicity, religiosity/spirituality, academic status, and country of origin.Loss summary questionnaire  is a 19-item self-report measure of clinically impairing grief symptoms (r = 0.80). Cronbach’s alpha in the present sample = 0.92. A diagnosis of PGD was based on three criteria: (1) bereavement for 12 months or longer, (2) ICG total score = >30, and (3) a score = >10 on the Work and Social Adjustment Scale (WSAS). While the authors of the ICG originally recommended a cut-off score of 25 to diagnose of PGD, further analysis of the instrument  is a 19-item self-report scale that measures four dimensions of social support: emotional/informational , tangible support ; affectionate support (someone to love and make you feel wanted), and positive social interaction . Each item is scored on a five-point scale . The scale has good internal consistency reliability for all subscales as well as the overall support index with alphas = 0.91–0.97  is a 12-item self-report scale that measures two dimensions of attachment style: attachment anxiety  and attachment avoidance . Each item is rated on a 7-point scale from 1 (strongly disagree) to 7 (strongly agree). The measure has adequate internal consistency {alpha = 0.78 (anxiety) and 0.84 (avoidance) and good test-retest reliability [r = 0.82 (anxiety) and r = 0.89 (avoidance)]; Grief Support was assessed with the following question: “Think about the people to whom you have turned for support regarding your grief. Please indicate whether you sought help from the following and if so, their level of helpfulness : religious leader, licensed mental health professional at your college counseling center, licensed mental health professional in the community, doctor/general practitioner, family member, friend, or other.” A qualitative question was also asked about “any other ways you have tried to cope with your grief” and “how helpful each activity was in coping with your grief.”t-tests (continuous) were used to assess the association between PGD and demographic and psychosocial variables. Descriptive statistics were stratified by PGD diagnosis. Statistical significance was set at p < =0.05 for all analyses.STATA version 16.1 was used for all statistical analyses. PGD was the primary outcome. Chi-square tests  and Demographic and psychosocial characteristics stratified by PGD are shown in n = 120/899).Loss characteristics were stratified by PGD and are shown in X2  = 12.02, p = 0.04, whereby the rate of PGD among Black students (16.7%) was twice that of white students (8.8%). The rates of PGD among Asians (15.8%) and Hispanics (10.5%) were also higher than that of whites.Having PGD was associated with race/ethnicity, X2  = 15.7, p < 0.001, history of trauma other than the death, X2  = 6.2, p = 0.01, and attachment style: anxious, t(882) = −5.97 p < 0.001 and avoidant, t(882) = −5.03, p < 0.001.Higher rates of PGD were also associated with the history of depression or anxiety, X2  = 38.8, p < 0.001, whereby rates were higher for those who had lost a significant other (60.0%), parent (29.8%), or sibling (23.5%) than those who had lost a friend (17.8%), aunt/uncle (10.2%), or grandparent (9.8%). PGD was also associated with closeness to the deceased, X2  = 30.5, p < 0.001.PGD was associated with kinship, X2  = 18.3, p < 0.001. PGD was associated with sudden/unexpected death, X2  = 9.0, p = 0.003.The rate of PGD was higher for those who were bereaved due to suicide (30.0%) than for those who were bereaved due to accident (21.6%), homicide (18.9%), or illness (11.0%), There were no statistically significant associations between PGD and age, gender, religiosity/spirituality, college attended, academic year, being born in the United States, social support, number of losses, or years since the death.Sources of grief support and perceived helpfulness are shown in Other sources of comfort/coping included exercise , meditation, visual arts, listening to and playing music, writing in a journal, writing to the deceased, watching TV/movies, reading, playing video games, looking at old pictures/videos/text messages of the deceased, using social media, housework/chores, spending time with family and friends, talking to the deceased, caring for plants/animals, God/religion, therapy, support groups, charity work (helping others), time outdoors, crying/yelling, staying busy, visiting the cemetery, and traveling. One student wrote,I know that (my loved one) loved to write poems and songs; therefore, I sometimes write about her and life in general. Sometimes I sing and listen to the songs she taught me. I honor her memory by wearing her skirt on her birthday. When I was a child, I watched her sew the skirt and add beautiful embroideries by hand. I feel that wearing that skirt is special and it captures her essence and creativity, in many ways that live in me. All this has helped me to accept her death and not think about the ways that I was absent. I think about the wonderful person that she was, and I look for ways that her memory lives on. Remembering her is my way of grieving.To this author’s knowledge, this study is the first to report on the prevalence of PGD in a large sample of diverse American college students. The overall rate of PGD was 13.4%, higher than what has been reported in college samples, particularly when bereavement was limited to the past 2 years (0.05–5.5%). In contrast with the aforementioned studies, the present study included lifetime death-losses, which more accurately captures the number of people with PGD. The average time since death in this sample was 6 years, suggesting that studies that have limited bereavement to the last 2 years have grossly underestimated the number of students suffering with the disorder.The rate of PGD of 13.4% found in this study is consistent with what has been found for adults. A meta-analysis of 14 studies revealed a pooled prevalence of PGD of 9.8% in people bereaved by natural causes although a variety of measurement tools and criteria for PGD were used across studies . The resp = 0.007), bereavement by homicide  and have a history of trauma other than the death . Stress theorists note that exposure to chronic stress has long-term physiological effects, which negatively impact both physical and psychological health (The rate of PGD among Black students (16.7%) was higher than any other group and nearly twice the rate of white students (8.8%). This is consistent with findings reported by l health . As a rel health , the deaAlthough history of depression or anxiety was not measured with validated instruments, the results presented here are consistent with prior research with both adolescents  and adulConsistent with prior research, attachment anxiety and attachment avoidance were both associated with PGD . Those wWhereas several studies with adults have shown that losing a spouse or a child increases the risk for prolonged grief , the preCircumstances of the death such as sudden/unexpected loss and loss due to suicide were also associated with PGD. In a literature review of bereavement and mental health after sudden and violent losses, For grief support, participants sought help primarily from a friend (80%), family member (76%), and “other helper” (54%), which included God, a boss, boyfriend/girlfriend, coach, coworker, and a pet. The least utilized supports were doctor, mental health counselor, religious leader, and professor. That 80% of the sample sought help from friends and only 60% found this support helpful suggests that colleges could better address the needs of this population by educating students about grief and how to respond to classmates who are bereaved.There are several limitations of this study. The sample was drawn from a college research pool available mainly to social science students who self-selected and may not be representative of the general population of college students. Data were collected at only one time point (after the loss), which makes it impossible to assess the directionality of the relationship between PGD and associated factors. History of depression/anxiety diagnosis and history of trauma were measured with single-item questions, which may limit the reliability of these variables. The use of self-report measures across the board may not be as reliable as clinician-reports due to distortion on the part of subjects. Finally, socio-economic status and financial consequences of the death were not included in the analysis, which could provide a more nuanced understanding of the rate of PGD in this sample. CUNY is a public university, accessible to students of all economic backgrounds. Forty percent of its students come from households that earn less than $20,000 per year and over 90% come from households considered to be low-incomeDespite these limitations, this is the first study to examine the rate of PGD in a large college student sample that includes a majority of students of color and takes into account lifetime death-losses. Results showed an overall rate of PGD that was higher than what has been reported for predominantly white college student samples. Results presented here also revealed a large disparity in rates of PGD between students of color and white students, providing further evidence of worse health outcomes for people of color in the United States.Screening of high-risk students should be conducted on college campuses and targeted treatment for PGD should be made available for those who need it. Mental health counselors at college counseling centers could be trained in evidence-based approaches such as Complicated Grief Treatment , which cAdditionally, several important demographic and psychosocial factors should be viewed as potential risk factors when screening for PGD including race/ethnicity, history of depression/anxiety and trauma, insecure attachment style, kinship, closeness to the deceased, sudden/unexpected death, and death due to suicide. College counselors and peer educators should be alert for these factors in order to identify and refer bereaved students who may be most at risk for PGD.The raw data supporting the conclusions of this article will be made available by the author, without undue reservation.The studies involving human participants were reviewed and approved by the University Integrated Institutional Review Board of The City University of New York. The participants provided their written informed consent to participate in this study.KG conducted all aspects of the project including research design, data collection, data analysis, preparation of the manuscript, and revisions.The author declares that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


Objectives: We aimed to develop a scale and evaluate this scale's validity and reliability to measure factors affecting people's knowledge and attitudes toward the pandemic breaking transmission chain.Methods: This exploratory mixed-method study was carried out in two phases: (1) item generation using literature reviews and interviews and, (2) item reduction by psychometric assessments of the developed scale. The face, content, construct , convergent, and discriminant validity of the scale were assessed in the Iranian population (n = 500) from March to June 2020. The Composite Reliability (CR) and the internal consistency correlation coefficient were estimated.Results: The Knowledge and Attitude Scale Toward COVID-19 Pandemic Breaking Transmission Chain (KA-C) among the Iranian population included 18 items. Two factors with a whole variance of 66.05% were identified by exploratory factor analysis. Factors were labeled as “health literacy” and “home health empowerment.” The confirmatory factor analysis showed the goodness of fit. The CR of the scale for first and second factors were 0.965 and 0.833 receptively. The scale's internal consistency correlation coefficient was acceptable .Conclusion: The KA-C scale can be exerted to screen the people's knowledge and attitude about the COVID-19 pandemic breaking the transmission chain as a valid and reliable scale for further policymaking, health care providers, and for a multi-dimensional psychosocial assessment of the pandemic period. The world has been facing a new challenge in the past several last months. The new coronavirus, SARS-CoV-2, first emerged in Wuhan, Hubei province China, and has since then turned into a global pandemic . As a fiCOVID-19 can be transmitted through human-to-human contact and indirect contact with contaminated objects. COVID-19 can be transmitted through body fluid droplets from the mouth or nose, spreading when a person with COVID-19 coughs, sneezes, and talks. Droplets typically cannot transverse more than six feet . COVID-19 remains intact and contagious in droplets and can be suspended in the air for up to three hour. Additionally, contaminated droplets can settle on plastic, stainless steel, copper, and cardboard objects. A person can become infected if they touch the surface contaminated with COVID-19 objects and then contact mucous membranes such as the eyes, nose, or mouth. Therefore, health experts advise frequent hand-washing with soap and water . It seemPeople's awareness of the strategies mentioned is also crucial to breaking the transmission and contamination chain. During public health emergencies, official sources  and unofficial sources  play a major role in people's awareness , 9. TherIn Iran, after the first case of COVID-19 was registered on February 19, 2020, attempts have been developed to shut down religious congregations, schools, and colleges, as well as non-essential businesses. Efforts have been directed toward continuing education courses online. All cinemas, theaters, sporting activities, and gymnasiums were closed. Car and real estate businesses were also closed and hotels and accommodation centers received nearly zero guests. It was eventually declared that no place of business was safe from COVID-19 . During This exploratory mixed-method study was conducted in two phases, including item generation and item reduction. Literature reviews and interviews were conducted in the item generation phase, and psychometric assessment of the scale was conducted in the item reduction phase .In the literature review, the identification of terms attributed to knowledge and attitude was reviewed using WHO, CDC, and Johns Hopkins COVID-19 Hub websites from December 2019 until May 2020. The terms that have been searched in different databases  were “COVID-19,” “awareness,” “health protective behaviors,” “animal hygiene” and “social distancing.” The inclusion criteria were defined as any type of article and document related to COVID-19 prevention which were published between December 2019 and May 2020. Fifteen studies (13 English and two in Persian) were selected. After reviewing abstracts and full texts and excluding duplicate articles, seven articles were included in the study. Articles were mostly about the factors influencing the transmission and prevention of the virus such as correct and incorrect behaviors, knowledge, attitude, and beliefs related to the spread of the virus in the general population. Two independent researchers (MA and MM) performed article screening. Through this process, if there was a disagreement, a third reviewer (SK) made the final decision. According to the steps proposed by Graneheim and Lundman , the conTwenty semi-structured face-to-face interviews were organized to achieve a better overview of the concept of attitude and knowledge. Using purposeful sampling, the participants  were selected from the general population. On average, individual interviews took around 30 min. The interview questions were focused on the following sections:We want to know and assess what factors are involved in improving the contributor's (you) collaboration to obey instructions published from health organizations?Also, to assess people's tendency to consume formal or informal sources of information, we asked how patients get their information about COVID-19 or how they prepare themselves for protective behaviors?The interviews were recorded and transcribed. The texts of the interviews were analyzed with a directed content analysis method using MAXQDA software Ver.10.The components of people's awareness about breaking the transmission chain extracted in the first and second phases of item generation have emerged.Based on extracted codes, categories, and subcategories, an item pool with 23 items was prepared. Items were reviewed by researchers several times, and then some were removed and modified. Finally, an initial format of the knowledge and attitude toward COVID-19 (KA-C) scale with 23 items was prepared. The response options were based on a five-point Likert scale: “I completely agree, I agree, I don't agree nor disagree, I disagree, and I completely disagree.”Quantitative and qualitative methods were used to determine the face validity of KA-C . For quaN = 2), psychology (N = 2), epidemiology (N = 2), and disaster management (N = 4) were requested to apply their analysis of the scale in terms of grammar, phrasing, item allocation, and scaling to assess the qualitative content validity. Their recommendations were considered in the scale. The quantitative content validity was examined by calculating the content validity ratio (CVR) and content validity index (CVI). The same 10 experts then specified each item's necessity for the CVR evaluation. They scored each item as 1 = unnecessary, 2 = somewhat necessary, or 3 = necessary. Based on the Lawshe formula (1975), when 10 experts' opinions are considered, the minimum acceptable CVR is 0.64 (Ten experts with expertise and experience in psychometric assessment ( is 0.64 . For eva is 0.64 . Based ohttps://forms.gle/4qngsnDXFnkAcJdj7. (in the Persian language). Google Forms is an online questionnaire creation software. Considering 10 samples for each item  were used for data gathering. The 21-item scale was prepared using Google Forms at ach item , 210 indIn this study, the EFA was evaluated through maximum likelihood and Promax rotation. Bartlett's sphericity test and the Kaiser-Meyer-Olkin (KMO) index were used to determine the sample's adequacy for performing the factor analysis. A KMO of higher than 0.9 indicated an adequate sample . The fac2), Degree of Freedom (DF), Parsimonious Comparative Fit Index (PCFI) >0.5, Parsimonious Normed Fit Index (PNFI) >0.5, Minimum Discrepancy Function divided by Degrees of Freedom (CMIN/DF) , Comparative Fit Index (CFI) > 0.90, Incremental Fit Index (IFI) > 0.90, Tucker–Lewis index > 0.90, and Root Mean Square Error of Approximation (RMSEA) <0.08  <0.08 . To evalBased on the Fornell  criteriaThe internal consistency of the current scale was calculated by Cronbach's alpha, Average Interitem Correlation (AIC), and McDonald's omega . The accThe protocol of this study was approved by the Ethics Committee of the University of Medical Sciences of Mazandaran (IR.MAZUMS.REC.1399.596). All individuals voluntarily participated in this study. The names of the contributors were not mentioned in the report, and codes were used in the interview texts instead of their names. The declarations of the participants were not shared with other members of the research team to protect the confidentiality of the information. Furthermore, in the initial set of data gathering, participants were ensured that all answers will remain confidential and that involvement in the study is voluntary.A total of 500 individuals participated in this study. The majority of them (65.2%) were female. In the age group, most of the participants were 21–40 years old. 55.6% of the participants were educated in the field of experimental science, with only a few (39.2%) being educated in the field of medical and paramedical sciences. The remaining participants (60.8%) were educated in other fields see .P < 0.001).After performing face and content validity, the 21-item scale entered the construct validity step. In MLEFA, the KMO test value was 0.960, and the Bartlett's test value was 8,513.860 (Two factors were extracted (18 items) and named as “health literacy” (15 items) and “home health empowerment” (three items). These two factors explained 66.05% of the variance see .P < 0.001) and CMIN/DF = 2.893. Then, other model fit indices were calculated, and the good fit of the final model validated those values  pandemic breaking the transmission chain among the Iranian population between March to June 2020. The scale was developed using a comprehensive review of the literature and interviews, expert panel reviews to prospect content and face validity, EFA, and Cronbach's alpha coefficients to descry internal consistency and reliability. With 18 items, the KA-C focused on the profound impact of knowledge and attitude in COVID-19 prevention.People's knowledge and attitudes are important in public health, especially in pandemic disease control strategies. Knowledge (cognitive behavior) impresses on attitude , and attitude impresses on health behavior. So, when an outbreak happens, controlling disease spread is linked to disease-related literacy and people's attitudes . Thus, svia health education because COVID-19 has been declared a pandemic. Therefore, an improved understanding will affect attitudes and practices toward COVID-19 , and the second one is “home health empowerment” (three items). The “health literacy” can reflect the knowledge of participants, which can directly affect their attitudes. The first step in guiding future efforts in the educational or policymaking process, which has been shown to impact future actions, is to determine knowledge of precautionary actions for contracting the disease . Proper COVID-19 . The “hoCOVID-19 .Since this disease is new and, despite the extensive research done in the study of different ways of its transmission, we see that observers still sometimes suffer from this disease. Therefore, the scale items are prepared to examine how people are informed by news related to this disease, how this disease could be transmitted, and how they can prevent themselves and others from becoming infected with this disease. The items were designed so that factors such as age and gender do not significantly impact how to answer. Still, factors such as cultural and economic features in the answers may be effective .In this study, based on CVI and CVR, two items were omitted from the scale.From the results, the two factors' variance is 66.05%, which is acceptable, considering that this value is above 50%. It is also mentioned that if a factor above 60% achieves the sum of the two factors, it is considered an acceptable value –41. The vice versa. So it is crucial to having protective attitudes that keep both humans and pets distanced, to prevent becoming infected and to prevent further transmissions of the virus  hygiene in the COVID-19 era is vital because recent studies have indicated that the virus could be transmitted from pets to humans and he virus .Hence, based on EFA, the final version of KA-C is considered a two-factor scale with construct validity. In this analysis, the second-order CFA was performed to explore a more detailed model of structural equations. This approach aims to obtain a more important data collection method, assuming that one or more higher-order factors are responsible for the latent variables in the common variance and that the proposed scale has two orders . A high Furthermore, the most common indicators of goodness-of-fit were evaluated. Based on the results, goodness-of-fit for all indicators was appropriate. Some research has indicated that by first-order factor analysis, the intended construct must first be developed. The conceptual construct's excellent fit is then calculated using the second-order factor analysis to test the structural equation model .Cronbach's alpha correlation coefficient, omega McDonald and AIC of factors was an acceptable internal consistency. Internal consistency is usually a measure by which it tests the correlation of different items in a test . It also examines whether several items that measure the same overall structure produce identical scores .SARS-CoV-2 is a new and extremely infectious virus, and there is currently no precise treatment for COVID-19. It must be considered that if no appropriate action is taken, and if medications, vaccines, and patient monitoring programs are not widely enforced or effective, preventive behaviors are likely to continue until 2022 , 48. CurAltogether, the primary intervention in disaster management is based on proper information about “one health” and attitudes to this information, to best control this pandemic. Also, health policymakers could benefit from this scale—by assessing the people's knowledge and attitudes, they may gain better insights on their policy making.One limitation of this study is that this research was only performed in Iran, despite the appropriate number of samples. Since it was mentioned that cultural factors might also affect the results, it is better to also include other countries and cultures to prove its validity and reliability. Due to the disease's current situation, the construction and distribution of this scale were done electronically; thus, there was a decreased opportunity to send it to lower economic status groups who do not have access to cyberspace. Moreover, the mentioned economic issues also affect the results.It is crucial to attract attention toward factors that engage people's knowledge and attitudes toward COVID-19 and how to break the transmission chain using the Persian version of the KA-C scale. We designed and developed a scale using multifaceted variables through this study, operationalizing them on the validated scale. The final scale consisted of 18 items and two factors that explained 66.05% of the total variance. This scale could bring robust suggestions for health policies toward COVID-19 transmission control across the world. Furthermore, the factors of “health literacy” and “home health empowerment” are a new basis for capturing such infrastructures' interaction in providing important information for focused health policies. Altogether COVID-19 prevention, with different transmission methods, requires collaboration among different sectors, public and governmental institutions.The data sets generated for this study are available at reasonable request to the corresponding author.The studies involving human participants were reviewed and approved by This study was approved by the Ethics Committee of Mazandaran University of Medical Sciences by the code of IR.MAZUMS.REC.1399.596. All participants voluntarily participated in this study. Names of the participants were not mentioned in the study, and instead of their names, codes were used in the interview texts. The participants' statements were not discussed with other research team members to maintain the information's confidentiality. Also, in the initial set of data gathering, participants were ensured that all answeres remain confidential, and involvement in the study is voluntary. Written informed consent to participate in this study was provided by the participants' legal guardian/next of kin.All authors contributed to the study conception and design. Material preparation, data collection performed by MA, MM, MS, ShagK, and ShahK. PR and HS analyze data. The first draft of the manuscript was written by MA, MM, PR, ShagK, and ShahK and all authors commented on previous versions of the manuscript. All authors read and approved the final manuscript.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.


Pinus halepensis) is widely distributed in the Mediterranean region and in other areas of the world, where it has been introduced due to its adaptive capacity to xerothermic conditions. The giant pine scale Marchalina hellenica often infests Aleppo pine, as well as other pines, in several southeastern European countries, causing pine declines. When combined with the expected intensified heat and drought events in eastern Mediterranean, the impact of this biotic parameter on the host pines may be exacerbated. The importance of understanding the defense mechanisms of Aleppo pine is emphasized by the recent invasion of the pine scale in new regions, like Australia, lacking the insect’s natural enemies, where more intense negative effects on pine species may occur. To date, Aleppo pine’s physiological responses to the infestation by M. hellenica are largely unknown. This study aimed at assessing the responses of Aleppo pine to the giant pine scale attack, both on an ecophysiological and a metabolic level. For this purpose, gas exchange, needle water status, and carbon and nitrogen content were measured during 1 year on healthy and infested adult trees. M etabolic profiling of Aleppo pine needles was also performed before, during, and after the high feeding activity of the insect. The maintenance of stable relative water content, δ13C signatures, and chlorophyll fluorescence in the needles of infested pines indicated that infestation did not induce drought stress to the host pines. At the peak of infestation, stomatal closure and a pronounced reduction in assimilation were observed and were associated with the accumulation of sugars in the needles, probably due to impaired phloem loading. At the end of the infestation period, tricarboxylic acids were induced and phenolic compounds were enhanced in the needles of infested pines. These metabolic responses, together with the recovery of photosynthesis after the end of M. hellenica intense feeding, indicate that in the studied region and under the current climate, Aleppo pine is resilient to the infestation by the giant pine scale. Future research should assess whether these promising defense mechanisms are also employed by other host pines, particularly in regions of the world recently invaded by the giant pine scale, as well as under more xerothermic regimes.Aleppo pine ( Pinus halepensis Mill.) is a widespread species that is native to the Mediterranean basin  may be a key parameter affecting the performance of Aleppo pine, especially in the light of climate change. M. hellenica is a sap-sucking insect that feeds on adult trees of several pine species , but also on fir   . The insa D. Don .M. hellenica is highly valued in apiculture since honey bees are feeding on its honeydew to produce most of the world’s pine honey  has been reported to cause desiccation, attributed to tracheid injury and blocked water transport  to impaired needle gas exchange and water status and (b) to metabolic disorders in response to this biotic stress.In the present study, we analyzed a range of physiological parameters to assess the gas exchange patterns and water balance of healthy and infested M. hellenica in Sane, Halkidiki, Greece  as the infested study site. According to macroscopic observations before the initiation of the study, most pines at this site were to some extent infested by the giant pine scale and it could not be ensured that the few unaffected individuals would not get infested during the study period. In addition, field surveys at neighboring low-elevation Aleppo pine stands with similar growth conditions and biometric traits revealed that infestations by the insect were extended and completely unaffected forest stands could not be identified. For this reason, a 58-year-old Aleppo pine plantation in the experimental station of the Forest Research Institute (FRI), Thessaloniki, Greece  comprising healthy trees was used as the control site of the study. The height and diameter at breast height (DBH) of the studied sites are presented in In northern Greece, Aleppo pine is naturally distributed solely at the region of Halkidiki . Thus, wTo characterize the growth conditions during the study period, air temperature  and precipitation  were recorded on an hourly basis and data logged by automated meteorological stations were established close to the study sites.M. hellenica attack as indicated by the cotton-like wax secreted by the scale that was found on the tree branches. Measurements and sampling were conducted at both study areas for 1 year (August 2018–July 2019) on a monthly basis, on two consecutive sunny days. If cloudy conditions occurred on the second measuring day , measurements and sampling took place on the next sunny day.Within an area of c. 0.5 ha at each site, six and seven dominant, non-neighboring Aleppo pine trees were used at the control and infested site, respectively. Selection of the infested trees in the field depended on the macroscopic evaluation of Amax) and stomatal conductance (gs) were measured on 1-year-old needles using the Li6400XT portable gas exchange analyzer coupled to a Li6400–40 fluorescence chamber . Fully expanded needles of each cut branch were placed in the 2-cm2 cuvette, and the cuvette was firmly closed to ensure stable measurements. When no firm sealing of the cuvette is achieved, Blu-tack is frequently applied . Although optimal PAR may vary seasonally, we chose to keep the same PAR level throughout the study for comparability reasons. CO2 flow rate in the chamber was set to 300 μmol s–1 and reference CO2 at 400 ppm. Depending on the seasonal fluctuation of ambient air temperature, chamber temperature was maintained at a range of 17–25°C. P. halepensis was acclimated to the seasonal changes in growth temperature, as long as no heat waves occurred. We generally avoided hot summer days and initiated summer measurements the latest till 10:00 a.m. Likewise, very cold winter days were avoided and measurements in winter days were initiated later. In general, measurements were performed between 10:00 and 12:00 in summer and spring and between 11:00 and 13:00 in winter and autumn days. During each measurement, Amax and gs values were monitored and, when stable, 5–8 recordings were logged at c. 3- to 5-s intervals, and used to calculate mean values. On average, stable values of gas exchange parameters were achieved within 10–20 min after the needles were put in the cuvette. Intrinsic Water Use Efficiency (WUEi), was calculated by the ratio of Amax to gs. A pre-test performed with sunlit low canopy branches of three Aleppo pines grown at the control site (FRI) showed that Amax remained stable half an hour after abscission . RWC was calculated according to the formula of where FW is the needles’ fresh weight, DW is the needles’ dry weight after oven-drying , and TW is the turgor weight after the needles were saturated in water .For chlorophyll fluorescence measurements, the branches were placed in black bags for dark adaptation, transferred in cool boxes, and stored in a cold room (5°C) in the lab for 24 h. Maximum quantum efficiency of PSII (Fv/Fm) was measured in the dark-adapted needles using a Pocket PEA .13C signature, needles were collected from one of the two cut branches per tree and five trees per site were used as replicates (n = 5). The needles were oven-dried  and ground with a ball mill. An aliquot of approximately 2 mg of the fine powder produced per sample was used for the determination of total N and C content and δ13C composition according to 13C; ‰) of each sample was then determined asFor the determination of the total N and C content and δRsample is the 13C/12C ratio of the sample and Rstandard is the 13C/12C ratio of the Vienna Pee Dee Belemnite (VPDB) standard.where M. hellenica. Each branch was put in a separate plastic bag and stored in a cold room (5°C) until measurements. The stored samples were examined macroscopically and stereoscopically when needed. The number of nymphs was counted and the size (length and diameter) of each branch was measured to calculate its bark area. Finally, the number of nymphs was expressed on a bark surface area (cm2) basis, according to At the infested site (Sani), one additional branch per tree was collected using a telescopic scissor to assess the level of infestation by M. hellenica number of nymphs  was added as internal standard, samples were incubated at 70°C for 15 min and shaken at 1400 r.p.m. Two hundred microliters of chloroform was added and samples were incubated for 5 min at 37°C and 1400 r.p.m. and, subsequently, centrifuged for 5 min and 15,000 g. An aliquot of 80 μl of the supernatant was lyophilized overnight. For derivatization, 20 μl of a 20 mg ml–1 solution of methoxyamine hydrochloride in anhydrous pyridine  was added to the dried extracts. Samples were incubated for 90 min at 30°C and 1400 g before 35 μl of N-methyl-N-(trimethylsilyl)-trifluoroacetamide  was added and samples were derivatized at 37°C for 30 min and with shaking at 1400 r.p.m.Metabolites of needles were extracted and derivatized using a modification of the method described by Derivatized samples of needle extracts were analyzed with a G.C.–M.S. system , equipped with an autosampler  and controlled by the Agilent MASSHUNTER software (Agilent Technologies). For this purpose, 1-μl aliquots were injected in splitless mode into the system and metabolites were separated on a capillary column . Run conditions as well as MS settings were as described by Amax, gs, WUEi, RWC: n = 6 or 7, for the control and the infested site, respectively). For the metabolic analyses and the needle δ13C composition and total N, C content, five trees were used as replicates for both sites (n = 5) and samples were collected from one branch per tree.Two branches per tree were used for the physiological measurements and one branch per tree was used for determining the infestation level. In the former case, a mean of the values obtained by the two branches per tree was calculated and the individual trees were used as replicates .However, a difference between the sites was observed for the summer rainfall that was almost equally distributed in June and July at the control site, but took place almost exclusively in July at the infested site. Finally, mean daily potential evapotranspiration (PET) during the study period at the two sites, calculated according to M. hellenica nymphs was constantly close to zero until March 2019 (The number of rch 2019 . Nymph nAmax) showed a similar seasonal increase from August 2018 to January 2019 , then peaked in May, and declined during summer months.In both study areas, maximum net photosynthetic assimilation  exhibited small seasonal fluctuations and no substantial differences between needles of infested and healthy Aleppo pines . Fv/Fm wgs and Amax , opposite responses of RWC were observed between control and infested trees. Similarly, with the exception of May, needle δ13C did not differ significantly between infested and healthy trees  tended to be higher in infested vs. healthy trees throughout the study period, except from May 2019, but the differences among treatments were not significant . On the a nymphs . Finallya nymphs .M. hellenica, compared to healthy Aleppo pine trees, were tested at five different time points , based on the observed changes in the number of the insect’s nymphs  cycle, 12 sugars and derivatives, six amino acids, four phenolic compounds, and phosphoric acid . The inds nymphs  and the s nymphs . April, s nymphs . In addiAmax. It decreased when the number of M. hellenica nymphs peaked and increased before and after this peak in March and June  and during recovery .M. hellenica. The impacts of the insect’s attack range from pine growth declines and partial canopy desiccation to individual tree necroses, when combined with other abiotic and biotic stress factors  and in mid-summer (July) (Amax rates during winter months (December and January), in line with the species’ ability to fully exploit warm and sunny winter days in terms of assimilation  . Both hemilation . Indeed,es occur . Moreovees occur . The decm values  and closm values  in infess nymphs  and the s nymphs . Thus, oAmax (Amax during the intense infestation period. Limitation of Amax was probably mediated by partial stomatal closure (Amax and gs (M. hellenica may lead to structural modifications in the tracheids, which eventually could hinder water transport and lead to desiccation – as observed by other insects (13C signature between infested and healthy Aleppo pines (The limited photosynthetic rates during the insect’s intense feeding and the negative effect of the insect’s abundance on Amax  contradiAmax . ReducedAmax  and is cAmax . It is,  closure , as suppx and gs . Petraki insects . We, howpo pines . The sampo pines .M. hellenica results in impaired gas exchange in Aleppo pine is partly verified. However, we also observed compensatory responses in Aleppo pine, achieved by adjustments in its seasonal fluctuation of Amax. In addition, although related to stomatal closure, the limited assimilation during the intense infestation of Aleppo pines was not due to desiccation. Partitioning of carbon resources to needles may explain the decline in photosynthesis as explained in the next section.Therefore, our first hypothesis that infestation by Amax in the needles of infested Aleppo pines (M. hellenica mainly feeds on the phloem sap of the trunk and branches (Sucrose, the main end product of photosynthesis, follows the same seasonal pattern as po pines . This repo pines . On the po pines . Given tbranches , partiti2 concentration and mediate stomatal closure (e.g., The mechanism that explains the high carbohydrate induction in the needles on infested Aleppo pines may be similar to that imposed by environmental stresses. Drought resulted in reduced mobilization of carbon in the leaves . Droughtre e.g., , as suppre e.g., , 5A, 6.The high concentration of the three carbohydrates raffinose, D-cellobiose, and D-sequoyitol underlines the idea that the plant’s defense mechanisms were activated because they play important signaling roles in plant innate immunity . The indM. hellenica nymph accumulation (The level of tricarboxylic acids, like malic, fumaric, and citric acid, were indeed augmented during different phases of mulation , 6. Thesmulation  and are mulation . MoreoveHowever, most amino acids identified in our study were down-regulated during the high infestation period (April–June; 2 concentration (Ascorbic acid was depleted in needles of infested vs. healthy trees, almost throughout the infestation period , 6. Enhantration . HoweverIn summary, our second hypothesis that infestation by the giant pine scale induces metabolic disorders in Aleppo pine is rejected. Although ascorbic acid and amino acids were depleted, other metabolic changes included the accumulation of carbohydrates, TCAs, and phenolics, which can be seen as a successful defense of Aleppo pine against the insect’s herbivory.M. hellenica. Aleppo pine exhibited a cascade of defense mechanisms. The core included the limitation of photosynthesis during the peak of infestation and its compensatory stimulation in periods preceding and following the insect’s high feeding activity, under otherwise suboptimal environmental conditions. Defense reactions of both primary and secondary metabolism were observed. Accumulation of carbohydrates in the needles of infested trees signaled stomatal closure and photosynthetic inhibition, while induction of TCA intermediates enabled the production of phenolics, necessary in defense against biotic stress. The employment of defense responses by Aleppo pine during intense attack by the pine scale and recovery potential at its end may enable re-establishment of physiological functions and long-term resilience. Such a defense strategy may explain why the long-term occurrence of M. hellenica in eastern Mediterranean countries may have negative impacts on Aleppo pine, but does not seem to threaten its existence under the current climate. Future research should focus on diverse Aleppo pine forests and pine species to elucidate whether these responses are site- or species-specific and if they ensure defense of host pines in regions of the world recently invaded by M. hellenica, as well as under a drier and warmer climate.This is a first record on the ecophysiological and metabolic responses of Aleppo pine against the attack by the sap-sucking insect The raw data supporting the conclusions of this article will be made available by the authors, without undue reservation.MF and KR conceived this research. MF and DA designed this research. MF, FL, DA, and DM collected the data. MF, FL, GS, DM, and AP carried out the data analysis. MF, FL, and GS contributed to the data visualization. MF, HR, AP, and KR provided resources. MF wrote the manuscript. HR, AP, and DA reviewed 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.


The aim of this study was to assess the criterion validity of a new screening instrument, the Early Language Scale (ELS), for the identification of young children at risk for developmental language disorder (DLD), and to determine optimal age-adjusted cut-off scores. We recruited a community-based sample of 265 children aged 1 to 6 years of age. Parents of these children responded on the ELS, a 26-item “yes-no” questionnaire. The children were assessed with extended language tests . A composite score out of these tests (two tests below – 1 SD or one below − 1.5 SD) was used as reference standard. We assessed the validity of the ELS, measured by sensitivity, specificity, predictive values, and AUC. The optimal sensitivity/specificity age-dependent cut-off ELS score was at 15th percentile. Sensitivity and specificity were 0.62 and 0.93, respectively. Positive predictive value was moderate (0.53), negative predictive value was high (0.95), the positive likelihood ratio was 9.16, and negative likelihood ratio was 0.41. The area under the ROC curve was 0.88. The items covered the increasing language development for the ages from 1 to 6.Conclusion: The ELS is a valid instrument to identify children with DLD covering an age range of 1 to 6 years in community-based settings. Developmental language disorder (DLD) is one of the most common developmental problems in children and has a negative effect on children’s emotional functioning, academic success, and social relationships –5. Its eEarly identification of DLD in well-child care requires short, valid, and reliable instruments, that preferably cover the age range in which language develops. A widely used short instrument for delineated age categories in a large age range is the Ages and Stages Questionnaire (ASQ) . The ASQWe developed an Early Language Scale (ELS)—that measures language development in 26 easy observable items by parents, covering the full range of early language development (1–6 years). However, evidence on its validity and scoring is lacking. The aim of this study was therefore to assess the criterion validity of the ELS, and to determine optimal age-adjusted cut-off scores.This study was the validation part of a cross-sectional study on the development of an Early Language Screening instrument (ELS) for the measurement of language development in children . In the current study, we compare the results of the ELS with a reference standard in a community-based cohort of children aged 1 to 6 years. This study was approved by the Medical Ethical Committee of the University Medical Center of Groningen (M13.134252 / NL45253.042.13).We recruited parents and their children from 1 to 6 years of age in a two-step procedure. In the first step, we recruited well-child clinics, kindergartens, and schools. Second, these institutions recruited parents by distributing our folder, and collecting and returning reply forms. We asked them to do so according to predefined guidelines on the sample required , in order to create a well-balanced sample regarding age and sex distribution. Exclusion criteria were significant sensory impairments , or mental disorders . In total, 1231 parents of children from 1 to 6 were recruited via 162 institutions and provided normative data. In this study, we validated the Early Language Scale (ELS) in a subsample of the normative sample. For this subsample, all participating parents were invited to let their child participate in the validation part of the study , items were selected from an item bank of 75 items based on the criteria of Mokken scaling, which is based on the nonparametric Item Response Theory [H = 0.83) with Item H coefficients between 0.62 and 0.90 [The Early Language Scale (ELS) is a parental questionnaire consisting of 26 yes/no questions on the language development of children in the age of 1 to 6 years old (Appendix 1). The items were administered in Dutch. We used forward-only translation of the original items to English for international publication. A yes score results in one point; the ELS total score represents the sum of all items a child has acquired, this score varies from 0 to 26. The ELS consists of items on language development, within the domains of vocabulary (semantics), sentences/grammar (syntax and morphology), and communication (language use). In a prior study, using the data from a community-based sample , and communication. The reference tests measured the same aspects of language development; therefore, the reference standard is a combination of language tests that assess the aspects language comprehension and production (of words and sentences), and communication. Table Lexilist Comprehension (LLC) [Lexilist Production (LLP) [The on (LLC)  and Lexion (LLP)  to measuon (LLP) , 25. Inton (LLP) .Schlichting tests for Language Comprehension (SLC) [Word Production (SWP) [Sentence Production (SSP) [The on (SLC) , Word Pron (SWP) , and Senon (SSP)  to measuon (SSP) , 28. Theon (SSP) , 28.Language Standard (LS) [The ard (LS)  is a 20-ard (LS) . We extrChildren’s Communication Checklist – 2 – NL (CCC) [The NL (CCC)  is a 70-NL (CCC) . The subNL (CCC) .. Education level was classified into three categories: low , middle (secondary education), and high .We obtained data by parent report on the following background characteristics: age of the child, gender, birthweight, length of pregnancy, language situation at home, highest level of education achieved by the mother, maternal age at birth of the child, and family history  of language delayn = 1231). We chose 3 months age groups to be able to differentiate within an age year, as children achieve several language milestones per age year. We also performed ROC analyses determining the area under the curve (AUC), by calculating sensitivity and specificity for the various cut-off values. Finally, we assessed whether the performance of the ELS differed by background characteristics.First, we selected children with two or more reference test scores − 1 SD or more below the mean or one reference test score (on language comprehension or production) − 1.5 SD or more below the mean in order to identify these children as children with atypical language development. Second, we assessed the criterion validity of the ELS. To visualize our data, we plotted the ELS total score against age, and added smoothed splines, representing group means, one for the children with typical language development and one for children with atypical language development, based on the combined reference standard. Next, we calculated sensitivity and specificity indices, positive predictive value and negative predictive value, and positive likelihood ratio and negative likelihood ratio, for various age-adjusted cut-offs of the ELS against the reference standard. The age-adjusted cut-off values of the ELS were calculated for 3 months age groups regarding 5th, 10th, 12th, 15th, 25th, 50th, 75th, and 100th percentiles within our norm sample  of the children. These 29 children were from all ages . One child spoke Dutch as first language and another language at home, one child had a birthweight below 2500 g, five children were born at a pregnancy duration of less than 37 weeks, and two children had mothers with a low education level.ELS total scores for children with a typical language development, based on the reference standard, were higher than for children with an atypical language development. ELS total scores for both groups increased across ages, with a ceiling effect at the maximum score of 26 on the ELS  was built using various age-adjusted cut-offs of the ELS. The area under the receiver operating curve (AUC) in Fig. F = 1.19, p = 0.315), Length pregnancy  = 2.47, p = 0.06), maternal age  = 1.32, p = 0.27) assessed using one-way ANOVA. There was also no significant effect of gender  = 0.69, p = 0.88). Maternal education and language could not be tested as these had too little observations in one or more subgroups. Maternal education low is more prevalent in the true positive group, confirming that children from low educated mothers have slower language development. Regarding language situation at home, most of the children with Dutch and another language spoken at home had typical language development and were not identified by the screening, indicating that the language situation did not affect the performance of the ELS. Finally, results were similar if the 17 included preterm children were excluded . In general, results confirmed expectations.Finally, we assessed whether the performance of the ELS differed by background characteristics, i.e., for the four subgroups: true positives, false positives, false negatives, and true negatives of the ELS. We found no significant effect of birthweight  in children aged 1 to 6 using a composite score of language tests as reference standard. We found that the ELS has a good validity making use of only a limited set of items feasible for an age range of 1 to 6 years.We found that the ELS has a good validity, i.e., an AUC of .88, comparable with the CDI and LDS with reported AUC’s of .86 and .85, respectively , 33. HowWe found a sensitivity of .62 and a specificity of .93, which is suitable for community-based screening. A high specificity, of at least .90, minimizes over-referrals with its negative effects, e.g., discomfort for parents and children, and costs . On the The properties of the ELS, e.g., a limited set of 26 items reflecting a broad definition of language development that is also feasible for a large age range, make the ELS an addition to other valid language screening instruments. The ELS exceeds the age range of the MacArthur-Bates Communicative Development Inventory (CDI) short forms  and the The scale has been developed in Dutch, but consists of items that are very common in related languages, such as English and German. For instance, an item like “says two-word sentences” (item 11) is rather similar in those languages. Use of the scale in other languages requires additional research to obtain language-specific norms and a validation in that language.p values were < 0.001, indicating that the results of the ELS are not based on coincidence, and therefore, the sample of n = 265 was sufficient to assess validity. However, a limitation of this study is that despite its size, the number of cases was still relatively small. This limited the potential to analyze outcomes per age year. Another limitation of our study is that the sample had some overrepresentation of mothers with a high education level. This resulted in cut-off norms that might be slightly low for children from disadvantaged families, as evidence suggests that children of highly educated mothers (and fathers) have better language [A major strength of our study is that we used a large, community-based sample, equally distributed over gender, and age groups of 1 to 6 years, with state of the art reference tests. We tested the power of our sample based on the assumption of a prevalence of 11% atypical language development and 89% typical language development (according to the reference standard) in our sample, i.e., the prevalence under the independence. Both resulting language . HoweverThe results of this study show that the ELS is a valid instrument for the detection of children at risk for developmental language disorder in a population-based sample. The ELS can provide support to professionals working in well-child care and early educational settings to quickly identify children (1–6) with an atypical language development. This identification of children supports timely referral to proper diagnostics and intervention. This may improve these children’s emotional functioning, academic success, and social relationships.The validity of the 26-item ELS is satisfactory for a broad age range. Further research is recommended to investigate its age-specific validity and long-term predictive value, as well as the applicability of its set-up to other languages. The use of the ELS in routine community settings deserves further study regarding feasibility and effects in a clinical setting.The ELS is a valid instrument to identify children at risk for DLD covering an age range of 1 to 6 years in community-based settings. The ELS is promising for identifying children with atypical language development in a public health setting as it does not yield a high proportion of false positives.


Health-related physical fitness is vital for children with intellectual and developmental disabilities (IDD) to gain healthier lives. The adapted rhythmic gymnastics (ARG) program was designed for children with IDD and is aimed at testing the effects of the exercise program on children's physical fitness.  Participants were recruited from two special needs schools in Beijing of China. Twenty-two children with IDD were assigned to an ARG experimental group or a traditional control group. The experimental group took part in a 16-week ARG program consisting of three 50 min sessions each week. And children's body composition, aerobic capacity, and musculoskeletal functioning were measured by the Brockport Physical Fitness Test (BPFT) before and after the program. p = 0.025 < 0.05) and upper limb strength (dumbbell press test: p = 0.038 < 0.05). Compared to the pretest, most of the physical fitness parameters improved significantly in the experimental group except BMI, and flexibility of the experimental group children showed a substantial increase.  The between-group analysis revealed great improvements for the experimental group in abdominal strength (curl-up test:  Most of the physical fitness parameters of children with IDD in the experimental group improved significantly, especially on abdominal strength and upper limb muscle strength when comparing to the control group. It is recognized that children with IDD are poorer at all kinds of physical activities than typical development children, which in turn renders them to be less active, to have poor motor abilities, and to attempt a sedentary lifestyle. Such conditions will lead them to many chronic diseases, such as obesity or cardiovascular disease. Low physical fitness also brings negative impacts on one's ability to integrate into society . HoweverSince poor fitness seems to be the threatening factor for exacerbating the disabilities associated with children with IDD , aerobicThe ARG program was designed based on the basic gymnastics exercise and dance elements with small apparatus, which was used to develop participants' self-control, motor skills, and physical fitness. Simple, single, and combined aerobic rhythmic movements are more suitable for children with different abilities to learn, because the sense of rhythm is children's innate ability, which makes each movement more natural. It is easy to master the ARG program for young-aged participants, which makes them become confident and initiative. The most valuable finding is that the rhythmic movements combined with music are more appealing to encourage the participants to engage in various kinds of movements than other activities. None of these programs have been designed or included children with IDD before.The purpose of our study is to examine whether participation in an ARG program could have a positive impact on levels of physical fitness among children with IDD.N = 12; ASD = 4, ID = 5, and DS = 3; mean age = 7.2 years; and IDD level moderate) came from School A; through the fundamental motor skill test , 12 children out of 25 who performed higher scores got chosen. The FMS test indicates children's retardation of gross motor abilities and of a sufficient cognitive competence to adapt to the ARG program. In this study, the FMS test was only used to select participants, hence no FMS data analysis in this study. In School B, the FMS test was also executed to select 10 participants for the control group. The control group  was matched for age, IDD level, and disability types with the experimental group  from two special needs schools in Beijing of China . Participants were identified as active and healthy. They did not have cardiac respiratory conditions that could be dangerous during aerobic rhythmic exercise, and they were able to understand basic visual and verbal instructions. The experimental group . Considering IDD children's low levels of moderate to vigorous physical activities (MVPA), this program was designed to be stimulating, interesting, and easy to manipulate and is based on children's motor skills and cognition. Because of the program is fun, understanding and mastering the exercise could overcome the children's participation barriers affecting their physical fitness . The ARGN = 3), rehabilitation teachers (N = 2), dance teachers (N = 2), and university professors (N = 3) whose research field was adapted physical education and who had ample knowledge about children with IDD. After three rounds of questionnaire, the adapted motors identified by experts got chosen. To further confirm the feasibility of the ARG program, the preexperiment was conducted before the program. The teachers taught the experimental group children with expert-identified ARG movements for about seven 20 min classes and judged their acceptance of the contents from children's learning states. Finally, 96% expert-identified contents got chosen for the final program. Music acted as an important element in the ARG program. Music offered children both amusement and comfort. Researchers suggested that music was the signal to guide children's movements [Zunea-Zunea was chosen to stimulate children's enthusiasm and cue them to step on the tempo. Children could shake their heads and bend their knees consistently within the rhythm. Slow-paced music was played while stretching and relaxing.The Delphi technique used to testify the appropriateness of the ARG movements was used to collect data from experts . Expertsovements  and alsoovements . All movN = 4) of the ARG program, majoring in gymnastics from Beijing Normal University, were supervised by an instructor with expertise in working with disabilities. Children in the experimental group participated three times a week in a 50 min class for 16 weeks, each class  to promote the interaction between children and teachers; stage two, warm-up (5 minutes) to reduce the possibility of injury and keep up to speed at the exercise; stage three, core activities (35 min) including basic rhythmic gymnastics skills such as imitating animals and several exercise games such as crawling to compete with other peers; stage four, cool-down (5 min) to explore the children's satisfaction level; and last stage, feedback (3 min) to end up the class.After the preexperiment, the ARG program was finalized. Teachers . Participants' weights and heights were measured using a standard protocol. Body composition would unlikely be changed in a short period of time; there were no disability-specific standards for body composition [Body composition was reflected by BMI. BMI was calculated as the weight (kg) divided by the square of height ; participants sat down at the test apparatus. One leg was fully extended with the foot flat against the testing instruments; the other knee was bent. The participants reached directly forward with both hands and held the position for at least 1 s. After 3 trials, the maximum value was used to reflect flexibility. Each leg's flexibility was tested separately.The pretest was performed one week prior to the initiation of the study, and the posttest was completed within one week of its conclusion. All tests were undertaken by the students of the Department of Physical Education from BNU, who were trained in the administration of the BPFT and supervised by an instructor expert in adapted physical education.t-tests were used to test the pre- and postperformances of the two groups, which are aimed at finding if there were some progress made by children in the two groups after physical exercise. Independent-samples t-tests were used to test whether there were significant differences in pre- and postperformances between the two groups. The assumption of homogeneity of variances (Levene's test) was violated for the 10 m PACER run test; the assumption of normality (Wilcoxon) was violated for the standing long jump test and the 10 m PACER run test; hence, the standing long jump test and the 10 m PACER run test were assessed by the Mann Whitney U test. There were no significant differences between groups before the intervention.Data were analyzed using the SPSS 20.0 statistical software. The statistical significance level was set at 0.05. Descriptive statistics were conducted to describe the participants in this study and to calculate the mean and standard deviation. Paired-samples The attendance rate of the two groups of children in the program was 97%  and 95.5% (control group) separately, which made the results convincing.p < 0.05). The curl-up test , trunk lift test , dumbbell press test , standing long jump test , and 10 m PACER run test  had significant differences compared to pretest, except the BMI test and sit-and-reach test (p > 0.05). Only the standing long jump test  was significantly improved in the control group, while other parameters were not significantly different from the pretest. The curl-up test  and sit-and-reach test (t(L) = 2.216, p(L) = 0.054 > 0.05; t(R) = 1.608, p(R) = 0.142 > 0.05) even decreased slightly from the pretest.From p > 0.05). Hence, it could be concluded that participants in both groups were similar in relevant variables before the intervention began (p = 0.025 < 0.05), dumbbell press test (p = 0.038 < 0.05), and sit-and-reach test (p(L) = 0.043 < 0.05, p(R) = 0.047 < 0.05). However, the BMI test (p = 0.109 > 0.05), trunk lift test (p = 0.957 > 0.05), standing long jump test (p = 0.346 > 0.05), and 10 m PACER run test (p = 0.674 > 0.05) showed no significant differences between the two groups.In order to eliminate the interference of other factors on the experimental results, the physical fitness between the two groups was compared. At pretest, there was no significant difference in all parameters of physical fitness between the two groups.  of participants can be improved through aerobic activities such as aerobic dance [Aerobic capacity is one of the qualities to be developed in rhythmic gymnastics which requires a long time of technical and physical exercises. The muscular endurance and aerobic capacity of children in the experimental group were improved. Through continuous vertical and horizontal jumping in different directions in the ARG program, children's lower limb strength and dynamic balance have been improved, and standing long-distance jumping has also been improved. In addition, long-term aerobic exercise can promote children's aerobic function. Compared with routine tests, children can run longer distances at a stable speed. Studies have shown that moderate to intense aerobic exercise can improve the cardiovascular system and respiratory system . In the ic dance –44. The Upper limb motor function is an essential component to complete the fundamental motors required in daily life, such as moving, lifting, and holding stuffs. In our study, the upper limb strength (dumbbell press test) of children in the experimental group may be improved by crawling using the upper limbs to control the body. The muscles of the extremities continuously overcome resistance and contract continuously through repeated exercise. Through rhythmic exercise and manipulation of equipment, the muscles can be contracted to increase the strength of the upper limbs. Abdominal strength can be enhanced by imitating animal movements, such as frog balance, monkey jumping, and various core strength training exercises. Stimulating abdominal strength will also make the child's motor skills more proficient. Lower limb explosive strength can be measured by the standing long jump test. Both groups of children improved significantly in this test, which reflected the increased control, coordination, and explosive movement of the children. It can be inferred that various functional motor skills in the control group, such as squats, standing vertical jumps, and other simple body movements, can enhance children's lower limbs. Skopal et al.'s and Douda's research studies show that gymnastics has a positive effect on participants' lower limb strength [Muscle strength is related to intelligence. Usually, children with IDD shows weak muscle strength, but muscle strength is important for health, for keeping good body shape, and for gaining independence in activities of daily living . In our strength , 49. Abastrength . Piazza strength . Douda astrength . AlthougThe sit-and-reach test was used to determine the flexibility. The results showed that the children in the experimental group performed better than those in the control group, and there were significant differences between the two groups. Although the progress of the children in the experimental group was small, the control group children were less flexible than before. This means that according to ARG practice, the experimental group children kept their flexibility stable and had a substantial increase. Gymnastics plays an important role in promoting the flexibility of the participants' body , 54. VarAt present, most of the studies on BMI and aerobic fitness mainly target nondisabled groups. In the study of Montosa et al., girls and adolescents had low BMI, waist circumference, and body fat percentage after aerobic exercises . Gerstl Results of our ARG program support the idea that properly selected exercise can improve the development of physical fitness in children with IDD. Our study has limitations. First, the sample size is small. In order to distinguish between different forms of disability that may exist between accurate diagnoses, a larger sample size is required, and the popularization of the results requires further examination through a large sample size. Second, there are not many schools with special needs participating in the program. It is necessary to verify the promotion of the ARG program in more schools, and when all schools have obtained the same effect, this can reflect more stable and reliable conclusions.This research demonstrated that after participating in a 16-week ARG program which targeted the development of rhythmic movement skills and basic gymnastics exercise, the levels of health-related physical fitness of children with IDD improved. Moreover, significant improvements were reported for most of the physical fitness parameters, such as muscle strength, aerobic capacity, and explosive strength when compared to the pretest. The ARG program provided rhythmic movements that were safe and feasible for children with IDD as evidenced by the progress participants made. Long-term and feasible exercise participation needs to be emphasized to promote physical fitness. However, future research could be needed to examine how well these physical fitness improvements are maintained.


Development of a core outcome set (COS) for clinical trials for COVID-19 is urgent because of the pandemic wreaking havoc worldwide and the heterogeneity of outcomes in clinical trials.A preliminary list of outcomes was developed after a systematic review of protocols of clinical trials for COVID-19. Then, two rounds of the Delphi survey were conducted. Stakeholders were traditional Chinese medicine (TCM) experts, Western medicine (WM) experts, nurses, and the public. Patients with confirmed COVID-19 were also invited to participate in a questionnaire written in understandable language. Then different stakeholders participated in a consensus meeting by video conference to vote.Ninety-seven eligible study protocols were identified from 160 clinical trials. Seventy-six outcomes were identified from TCM clinical trials and 126 outcomes were identified from WM clinical trials. Finally, 145 outcomes were included in the first round of the Delphi survey. Then, a COS for clinical trials of TCM and WM was developed. The COS included clinical outcomes (recovery/improvement/progression/death), etiology , inflammatory factor (C-reactive protein), vital signs , blood and lymphatic-system parameters , respiratory outcomes , clinical efficacy , and symptoms . Outcomes were recommended according to different types of disease. Outcome measurement instruments/definitions were also recommended.Though there are some limitations for the research, such as insufficient patients and the public involvement, and the unbalanced stakeholders' region, the COS for COVID-19 may improve consistency of outcome reporting in clinical trials. It also should be updated with research progression. Since December 2019, a novel coronavirus causing pneumonia has been spreading around the world. It was temporarily named “2019 novel coronavirus” on 2 January, 2020, but the World Health Organization (WHO) officially named it “coronavirus disease 2019” (COVID-19) on 11 February 2020. The coronavirus that causes COVID-19 was termed “severe acute respiratory syndrome coronavirus 2” (SARS-CoV-2) on 11 February 2020 by the WHO.As of 10:00 CET on 2 April 2020, 896,450 cases of COVID-19 had been reported to the WHO, and 45,526 of these cases have died . COVID-1The first clinical trial of COVID-19 was registered on 23 January 2020 . Since tPreviously, we found several problems regarding the protocols of clinical trials of COVID-19  that maA COS is an agreed standardized set of outcomes that should be measured and reported, as a minimum, in all clinical trials in specific areas of health or healthcare . When reThis COS was based on: (i) a population with confirmed COVID-19 cases of “mild”, “ordinary”, “severe”, or “critical” types; (ii) interventions that include TCM and WM; and (iii) the COS being applied in randomized controlled trials (RCTs) and observational studies.This COS has been registered on the Core Outcome Measures in Effectiveness Trials (COMET) database . This reA steering group was formed by a TCM expert, WM expert, methodologist, nurse, and statistician. They conducted the research protocol, made decisions if there was confusion, and attended the consensus meeting to facilitate COS development.The stakeholders in the Delphi survey included TCM experts (clinicians and researchers), WM experts (clinicians and researchers), nurses, patients, and the public.COVID-19 is a new infectious disease that is spreading rapidly. In China, many clinicians have been trained to face emergencies, irrespective of whether they are on the “frontline” of the “battle” against COVID-19. More than 40,000 clinicians and nurses from other areas of China moved to Hubei Province to support the local medical system. Not all of these clinicians and nurses were trained in respiratory medicine or critical care. To obtain perspectives on a larger scale, we used “snowball” sampling to extend the sample size. We invited members from the Clinical Research Information Association of China and the Information Association for Traditional Chinese Medicine and Pharmacy to participate in the Delphi survey. We asked them to send the questionnaire to their colleagues.via social media  to invite the public to participate.We believe that the perspectives of patients and the public are important. Hence, we sent the questionnaire To obtain patients' perspectives, frontline clinicians of Dongzhimen Hospital, Beijing University of Chinese Medicine  invited and helped patients who consented to complete the questionnaire.The stakeholders in the consensus meeting were TCM clinicians, WM clinician, nurse, methodologist, evidence-based medicine researcher, and staff from the Chinese Clinical Trials Registry.All the databases of ICMJE-accepted platforms of clinical-trial registries  which outcomes do you think are important but were not included in the questionnaire? (ii) what is your opinion of this questionnaire?The questionnaire for patients was sent by smartphone, too. It included outcomes/outcome domains that were understood readily by patients. Patients were asked to vote on which outcomes/outcome domains were important to them. There was one open-ended question: which outcomes do you think are important but were not included in the questionnaire?The questionnaire for professionals and the public employed a nine-point scoring system, which has been used in previous COS studies . A scoreFor the Delphi survey administered to professionals and the public, the consensus definitions were: (i) consensus in: ≥70% of participants in all stakeholders scored the outcome as 7–9, and <15% of participants in all stakeholders scored the outcome as 1–3; (ii) consensus out: ≤50% of participants in TCM experts and WM experts scored the outcome as 7–9; (iii). no consensus: anything else.The voice of patients should be considered. Hence, for the patients' survey, the consensus definition was outcomes that were voted by >50% of patients.For the consensus meeting, the consensus definitions were: (i) consensus in: outcomes that were voted by ≥70% of participants; (ii). consensus out: outcomes that were voted by <70% of participants.The consensus meeting was held by teleconference. The contents of the consensus meeting covered: (i) the reporting background and methods of the research; (ii) reporting the results of the Delphi survey of professionals and the public, and the results of the patients' questionnaire; (iii) discussing the candidate outcomes and their instruments/definitions; and (iv) voting on the outcomes and reaching a consensus.The entire project is part of a clinical trial of COVID-19, which was approved by the Ethics Committee of Dongzhimen Hospital (DZMEC-KY-2020-09). Because of the special circumstances of the COVID-19 pandemic, participants who completed the questionnaire were assumed to have provided consent for their data to be used.A total of 160 protocols from 19 platforms of clinical-trial registries were searched. After reading the titles and study details, 63 non-relevant or ineligible study protocols were excluded. Finally, 97 eligible study protocols were included from ChiCTR and ClinicalTrials.gov. Thirty-four clinical trials were for TCM therapy and 63 clinical trials were for WM therapy. All clinical trials will be conducted in China. These clinical trials comprised 75 RCTs (53 for WM and 22 for TCM) and 22 non-RCTs (10 for WM and 12 for TCM). For 34 protocols of TCM clinical trials, there were 76 individual outcomes from 16 outcome domains after the merging and grouping of outcomes. For 63 protocols of WM clinical trials, there were 126 individual outcomes from 17 outcome domains after merging and grouping. The list of outcomes can be obtained from the systematic review . There wAfter removing duplicated outcomes, we developed the questionnaire for first round of the Delphi survey. After review by the steering group, 145 outcomes were included in the questionnaire.We had incentive measures to improve the response of the Delphi survey (randomized rewards after completing and submitting the questionnaire). The time planned for round 1 of the Delphi survey was from 4 March to 12 March 2020. As of March 9, 2020, 176 participants had completed the questionnaire. After review, 51 questionnaires were found to be invalid. On March 8 and 9, 2020, ≤5 questionnaire/day were completed, and almost all of them were invalid. Most of the invalid questionnaires had been completed by the public within 5 min (it was not possible for people who were unfamiliar with COVID-19 to complete the questionnaire) or who had chosen the same score for all outcomes. After discussion with the steering group, we decided to stop the Delphi survey.Finally, 125 valid questionnaires were evaluated. The characteristics of participants in the round 1 of the Delphi survey are shown in Only 15  participants were in Hubei Province. However, the Internet Protocol (IP) address that the electronic questionnaire obtained showed that 30  participants were in Hubei Province. Only one person  was from outside of China (Canada). The regions of participants are shown in More than 20 participants provided outcomes or significant proposals for round 1 of the Delphi survey. After discussion with the steering group, six of them were added to round 2 of the Delphi survey. There are some “consensus out” outcomes in different stakeholders, but no “consensus out” outcomes by all of the stakeholders were noted, so we did not delete any outcomes in the round 2 of Delphi survey.According to the significant proposals from participants in round 1 of the Delphi survey, the steering group decided to add more personal information. To reduce the risk of invalid questionnaires, participants would receive a random reward if the completed questionnaire was considered to be valid. Participants were also asked if they agreed to be mentioned in the “acknowledgements” section when the research was published. Computed tomography and magnetic resonance imaging of the hip were grouped as “hip imaging”. There were 150 individual outcomes in round 2 of the Delphi survey.The feedback from participants in round 1 of the Delphi survey showed that scoring for some outcomes was difficult. Hence, in round 2 of the Delphi survey, participants had the opportunity to choose “unclear” for any outcome that was difficult to determine. The median score of each outcome from each stakeholder group was shown in round 2 of the Delphi survey. The steering group wanted more people to participate in the Delphi survey. Hence, the questionnaire was sent to potential participants (irrespective of whether they completed round 1 of the Delphi survey) and they were asked to invite colleagues who might be interested in this research. Round 2 of the Delphi survey was conducted from 11 to 13 March 2020.A total of 110 questionnaires were completed, but seven of them were invalid, so 103 valid questionnaires were assessed. The characteristics of participants in round 2 of the Delphi survey are shown in The IP address that the electronic questionnaire obtained showed that 28  participants were in Hubei Province. The regions of participants are shown in After the results of round 2 of the Delphi survey had been reviewed by the steering group, outcomes that achieved “consensus out” by TCM experts and WM experts were excluded. Outcomes that achieved “consensus in” from stakeholders were grouped and presented according to the classification of disease and interventions. They were presented to consensus-meeting participants with “no consensus outcomes” before the consensus meeting was held.Results suggested that nurses and the public may find it difficult to score outcomes because they may misunderstand the terminology. Hence, we developed a simple questionnaire with understandable language for patients. There were 43 outcomes/outcome domains in the questionnaire. The list of outcomes is in The consensus meeting was held on 18 March 2020 and was a video conference. Six frontline clinicians  as well as one frontline nurse, one methodologist, and one researcher who participated in the design of clinical trials of COVID-19 were invited to attend the consensus meeting. The participants were from Shanghai (one), Beijing (five), Tianjin (two), and Guizhou (one) and all were voting participants. All clinicians and nurses had worked in Hubei Province after the COVID-19 outbreak. Two additional participants  attended the meeting but did not participate in the discussion or voting.After reporting the results of the Delphi survey and patients' survey, participants discussed some outcomes they believed should/should not be measured in clinical trials. After discussion, voting participants were invited to vote on which outcomes should be included in the COS of COVID-19. The outcomes voted by ≥70% of participants were included in the COS. The voting results are shown in This COS was conducted rapidly and rigorously to report on an emergency in a specific environment. It can be used for any type of disease, intervention, and design. There is a specific outcome for TCM clinical trials: clinical symptom score. Researchers can measure the clinical symptom score according to different TCM syndromes. For some individuals, there are several measurements because there is no evidence to show which measurement is the best one. We hope researchers of clinical trials can use this COS to reduce heterogeneity in outcome reporting. Furthermore, our COS may help decision-makers to approve new agents for COVID-19 if researchers report important outcomes. However, researchers can report other outcomes according to the purpose of their research.Our study had four main limitations. First, due to the highly infectious nature of SARS-CoV-2, patients and the public did not participate in the design or development of the preliminary list of outcomes, but they participated in the process of Delphi survey. Second, the preliminary list of outcomes was developed from protocols of clinical trials when there were knowledge gaps in the prevalence, therapy, prognosis, and clinical characteristics of COVID-19. With the research progression, new important outcomes may be reported. Hence, the COS must be updated in the future. Third, the number of patients was small and all of them were from Hubei Province, so their perspectives may not reflect those of other regions in China or overseas. Fourth, almost all stakeholders were from China. Though one participant in round 1 of the Delphi survey was from Canada, his/her opinion reflected a Chinese perspective because the questionnaire was written in Chinese.https://medrxiv.org/cgi/content/short/2020.03.04.20031401v1. This manuscript was included in a priprint server https://www.medrxiv.org/content/10.1101/2020.03.23.20041533v2.The data is from public databases and does not include identifiable patient data. All of the primary data was searched from different registry databases before it was analyzed, and the results can be found in another manuscript: The studies involving human participants were reviewed and approved by Ethics Committee of Dongzhimen Hospital, Beijing University of Chinese Medicine. Written informed consent for participation was not required for this study in accordance with the national legislation and the institutional requirements.RQ, JC, and HS designed the research. RQ drafted the manuscript. CZho and XW conducted the systematic review. TL, XH, and YH conducted the Delphi survey and patients' questionnaire. XZ, ZC, and MZ developed questionnaire. CZha, JH, ML, SH, TH, and YS revised the questionnaire. JC and HS revised the manuscript. All authors read and approved the final manuscript.This work was supported by the National High-level Personnel of Special Support Program (W02020052) and the COVID-19 Project, Dongzhimen Hospital, Beijing University of Chinese Medicine (No.2020-dzmyy-lczx-yj001).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.