During the last decade, the Caribbean Hispanic islands experienced accelerated demographic aging, representing the fastest aging region within Latin America. Age-related non-communicable diseases, including dementia, are now reported at high prevalence. The Caribbean islands share similar genetic ancestry, culture, migration patterns, and risk profiles, providing a unique setting to understand dementia in the Caribbean-Hispanics. This perspective article aimed to describe the impact of dementia in the Caribbean, at a local and regional level and reflect on research strategies to address dementia. We report on 10/66 project findings, described research projects and regional plans for the region. According to our results, the prevalence of dementia in the Caribbean is the highest in Latin America, with 11.7% in Dominican Republic, 11.6% in Puerto Rico, and 10.8% in Cuba. Preliminary data from new waves of the 10/66 study shows increasing numbers of dementia cases. Furthermore, dementia is expected to be one of the most serious medical and social issues confronted by Caribbean health systems. However, there is a scarcity of knowledge, awareness, and health services to deal with this public health crisis. In light of the new evidence, local and regional strategies are underway to better understand dementia trends for the region and develop policies aimed to decrease the impact of dementia. Implementation of our national plans is critical to deal with an aging population with high dementia rates. Current recommendations include emphasizing public health prevention campaigns to address modifiable risk factors and expand support to caregiver and family interventions. Epidemiological studies show a rapid increase in dementia in Hispanic populations , 2. ApprIn relation to other Latino populations, Caribbean-Hispanics have several differences in customs, traditions, nutritional patterns, risk behaviors, and genetic admixtures, which may differentially impact the prevalence of dementia and expression of its symptoms. Furthermore, during the last decade, the Hispanic Caribbean islands  experienced accelerated demographic aging, representing the fastest aging region within Latin America, which posed a unique challenge for aging and dementia , 7. The This perspective article aimed to examine the associations of genetics and socioeconomic determinants with dementia and describe the impact of dementia in the Hispanic Caribbean islands at a local and regional level. In addition, we present current research and describe future projects in the region. Finally, we share current dementia strategies aimed to address this epidemic within the Caribbean area.The Caribbean is undergoing increasingly rapid population aging with the proportion of older persons (60 and over) increasing from 10% in 2000 to 14% in 2015, and projected to reach 25% by 2050 , 9. ConsAs number and proportion of older persons increase, a central question to the Caribbean region is whether the aging of this population will be accompanied by sustained or improved health, better quality of life, and sufficient social and economic resources. To date, the increase in life expectancy has not been coupled with improvements in lifestyle; risk behaviors such as consuming high fat and carbohydrate diets, smoking, and sedentarism are becoming more common. Likewise, population aging has been associated with a sustained increase in non-communicable diseases, including neurodegenerative disorders, like dementia , 12. DemSince 2003, the three islands in a coordinated effort led a major epidemiological study  to detern = 2813) and the Dominican Republic (n = 2011) from 2003 to 2007, and in Puerto Rico (n = 2009) from 2006 to 2008. The interviews were done in a single phase, and the methodology allowed for the diagnosis of dementia and its subtypes, as well as other mental disorders. Data obtained included sociodemographic characteristics, physical health, anthropometric measures; information about risk factors, disability, frailty, utilization of health services, income, lifestyle (including nutrition and exercise), care characteristics, caregiver burden, and genetic risk markers (including ApoE-4); implemented together with a physical and neurological examination. An interview with a reliable informant was required. An incidence phase (second wave) was conducted 3 to 4 years after cohort enrollment, 2007 to 2011. A third wave of the 10/66 protocol is currently underway (details of this new wave are provided below).In the Caribbean region, the 10/66 project surveyed 6833 residents, aged 65 and over, in Cuba , we are revisiting the original catchment areas to generate a revised representative estimate of the prevalence of dementia for the residents who are 65 years old or older; the updated sample includes those individuals who, since the first wave of the survey have been incorporated into the age group of interest, either from having aged into it or from having moved into the areas. The new, third-wave prevalence survey includes a comprehensive assessment of health status, which will consist of spirometry, body mass index, visual acuity, grip strength, and tests to determine the presence of hearing impairment . For the new survey  , 26. Thiportions , 29.APOE-ϵ4 is weaker in Caribbean Hispanics compared to Caucasian populations. In the three Caribbean islands being an APOE-ϵ4 carrier increases the risk to develop dementia by two folds (SHR 2.03 [1.32-3.11]); in Caucasian populations the risk among APOE-ϵ4 carriers is 10 to 15 times higher than in non APOE-ϵ4 carriers  may have a differential risk among the Hispanic population; therefore the influences of genetic risk factors in Caribbean Hispanics should be explored in future studies.According to our studies, the association between dementia and carriers , 30, 31.R 2.03 1.-3.11); icarriers  like APOPSEN1_L174M) affecting several families (280 family members) in the Cuban western provinces. Furthermore, two decades ago, a family study was developed in New York that included multiplex relatives affected with AD in Dominican Republic and Puerto Rico, leading to the discovery of novel PSEN1 pathogenic variant (PSEN1_G206A) (The frequency of Dominantly Inherited Alzheimer disease (DIAD) is relatively high in the Caribbean countries compare to Europe and US; , 34 whic1_G206A) . Since 21_G206A) , 39. SevAs mentioned early, the prevalence of dementia is high among use Caribbean-Hispanics, yet knowledge, health services, and awareness of the disease are scarce . AwareneUnfortunately, cognitive impairment and dementia diagnosis depend mostly on clinical suspicion, and ~50% of dementia cases are missed in primary care, delaying detection until later in the disease course. Furthermore, awareness about dementia diagnosis and care is not only relevant for health care professionals, but also for the general population, especially among those caring for someone with dementia; and these are unmet needs in LMIC , 45. ConSeveral local and regional strategies have been implemented to address these issues. The Alzheimer's Research Center in Cuba has developed a multidisciplinary Master's degree program to improve early diagnosis, treatment, and support of people living with dementia . ThroughAnother need is to expand the training of caregivers. The regional Alzheimer Associations have led a caregiver awareness program to improve caretaking abilities and expand caregivers' support. A full training video-series featuring relevant topics regarding caretaking in dementia and advice from health care professionals are available to view and share free of charge. “Conversando con Los Cuidadores”—new series in Spanish- includes eight videos focusing on critical issues related to the work and role of those who provide care for relatives with dementia. Training of caregivers will enhance their management ability and enables them to handle disruptive situations. The Dominican Alzheimer's Association has been developing online and on-site training sessions to support and decrease the burden associated with caretaking. The 10/66 “Helping Caregivers to Care, train-the-trainer” intervention was tested in a randomized control trial in the DR, providing evidence on the relevance of caregiver training to decrease the burden associated with care. This intervention continues to be widely used by the Dominican Alzheimer's Association to train caregivers . The University of Puerto Rico, Geriatric's Education Center, offers specialized courses to train caregivers in different sites, including a special arrangement with the Dominican Republic Embassy in PR, to train Dominican caregivers working in Puerto Rico. Likewise, there are multiple caregiver training programs in private universities on the island.In 2015, Cuba and the Alzheimer's Research Center announced the National plan for dementia, and it was adopted as a National dementia strategy by the Cuban Ministry of health in 2016 . The maiAlzheimer's disease Action Plan in response to the World Health Organization's (WHO) call to action, by rating AD and other dementias as a public health priority. The focal areas of the PLAN were divided into seven pillars: (1) Public Policy (2) Public Health Efforts and Epidemiologic Surveillance (3) Home and Community Caregiving Services (4) Education and Training (5) Diagnosis and Treatment, (6) Long Term Care Services, and (7) Long-Term Care Financing. Each area includes specifics goals to be accomplished by 2025.Similar to Cuba, in 2015, Puerto Rico launched the Finally, in July 2020, the Dominican Republic's Ministry of Public Health, in close partnership with Asociación Dominicana de Alzheimer, announced a national strategy for dementia with similar action points as Cuba and Puerto Rico. Future efforts in the region are needed to achieve a coordinated response and plan to address the rising number of dementia cases.The Caribbean American Dementia and Aging Study (CADAS) is new research study funded through the National Institute of Health (R01AG064778) aimed to support regional research efforts to expand the previous waves of 10/66 studies to rural populations and facilitate a nationwide dementia estimate . ResultsTo better understand disease pathology and AD progression in Caribbean-Hispanics, a new Brain Bank is already in place in DR, another one is being organized in PR. Our final goal is to develop a Hispanic Caribbean Coordinating Center for neuropathology sample collection and storage. To maintain a coordinated research program, Caribbean researchers are meeting several times per year, promoting the work's continuity and keeping the international collaborations. More recently, the Ministry of Higher Education Science and Technology of the Dominican Republic awarded the Brain Bank research funding to expand research on AD pathology, with a special focus in Tau patho-physiology. This project is a joint consortium with a local university, “Pontificia Universidad Catolica Madre y Maestra” and Universidad Autonoma de Mexico (UNAM).Dementia represents a challenge for public health in developing countries with rapid demographic transitions, such as those occurring in the Caribbean. We propose expanding our research efforts to rural areas and explore gene by environment interactions on dementia risk. Emphasis on genetic family studies and admixture and new clinical trials with Hispanic representation, should continue. The development of brain banks and the implementation of biomarker studies are current research priorities. Implementation of our national plans is critical to deal with an aging population with high dementia rates, ideally a true public health-focused approach should aim at social change and emphasize on public-health prevention campaigns directed to addressing modifiable risk factors and developing caregiver and family interventions.https://1066.alz.co.uk/.Publicly available datasets were analyzed in this study. This data can be found here: The studies involving human participants were reviewed and approved by Universidad Nacional Pedro Henriquez Ureña, Universidad de Ciencias Médicas de la Habana, University of Puerto Rico. The patients/participants provided their written informed consent to participate in this study.All authors had full access to all the data in the study and take responsibility for the integrity of the data and the accuracy of the data analysis. JL-R, JL-G, IJ-V, and DA: study concept and design. JL-R, IJ-V, and DA: acquisition, analysis, or interpretation of data. JL-G and DA: drafting of the manuscript. All authors critical revision of the manuscript for important intellectual content. JL-G: project administration. JL-R: study supervision.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.


As an increasing number of deaths occur in the intensive care unit (ICU), studies have sought to describe, understand, and improve end-of-life care in this setting. Most of these studies are centered on the patient’s and/or the relatives’ experience. Our study aimed to develop an instrument designed to assess the experience of physicians and nurses of patients who died in the ICU, using a mixed methodology and validated in a prospective multicenter study.Physicians and nurses of patients who died in 41 ICUs completed the job strain and the CAESAR questionnaire within 24 h after the death. The psychometric validation was conducted using two datasets: a learning and a reliability cohort.p = 0.002).Among the 475 patients included in the main cohort, 398 nurse and 417 physician scores were analyzed. The global score was high for both nurses [62/75 ] and physicians [64/75 ]. Factors associated with higher CAESAR-Nurse scores were absence of conflict with physicians, pain control handled with physicians, death disclosed to the family at the bedside, and invasive care not performed. As assessed by the job strain instrument, low decision control was associated with lower CAESAR score  versus 63 , Factors associated with higher CAESAR-Physician scores were room dedicated to family information, information delivered together by nurse and physician, families systematically informed of the EOL decision, involvement of the nurse during implementation of the EOL decision, and open visitation. They were also higher when a decision to withdraw or withhold treatment was made, no cardiopulmonary resuscitation was performed, and the death was disclosed to the family at the bedside.We described and validated a new instrument for assessing the experience of physicians and nurses involved in EOL in the ICU. This study shows important areas for improving practices. As an increasing number of deaths occur in the intensive care unit (ICU), studies have sought to describe, understand, and improve end-of-life care in this setting –5. Most Among end-of-life research projects, tools have been developed to evaluate the quality of dying and death in the ICU, such as the Quality Of Dying and Death (QODD) instrument . QualityThus, the French CAESAR project aimed to develop an instrument to assess the experience of relatives of patients who died in the ICU as well as the experience of healthcare professionals, physicians, and nurses. The relatives’ instrument was validated in a previous publication and presents valuable psychometric properties . This 15Here, our objective was to develop and validate two tools specifically designed to assess the overall experience of physicians and nurses caring for patients who died in the ICU. We used a mixed methodology to create the instrument that was then validated in a multicenter prospective study in 41 French ICUs.The study was approved by the institutional review board of the Paris-North Hospitals , Paris 7 University. Informed consent was obtained from each relative before study inclusion.Physicians and nurses were invited to participate and to complete the CAESAR tool on a voluntary basis. They received information about the study, and those who did not wish to participate could decline. No written consent was required.Before inclusion of patients, in each participating center, the local investigator, together with the head nurse, completed a description of the ICU.Development of the CAESAR instrument was described in a previous publication . A 33-itIn each ICU, the intensivist included consecutive adults who died after at least 48 h in the ICU and for whom the relatives had visited at least once. For each patient, the physician and the nurse in charge of the patient were asked to complete the instrument within 24 h after the death. At the end of the instrument, participants were also asked to complete information such as age, sex, years of experience in the ICU, and religious beliefs. As described elsewhere, relatives completed the tool 21 days after the patient’s death .After full data completion, 7 investigators  for the physicians’ instrument (CAESAR-P), and 6 investigators  for the nurses’ instrument (CAESAR-N), worked to reduce the instrument from 33 to 15 items, based on clinical relevance and item distribution, discrimination, and redundancy –16. InteWe also collected ICU, healthcare professionals’, patients’, and end-of-life characteristics.Physicians’ and nurses’ job strain was evaluated using an instrument that explores three domains: job demand (3 questions), control (5 questions), and social support (4 questions) –21. HighStatistical analyses were performed using the R 3.1 package with the “psy” package .p values of 0.05 or less were considered significant.Quantitative data were described as median ; comparisons between groups were tested using ANOVA test. Binary and categorical data were described as number and percentages. No imputation for missing data was performed. To assess the number of dimension identified in the survey, a principal component analysis and the corresponding scree plot were used. The cutoff was determined using a set of simulations that illustrated the amount of variance that may be expected by chance alone . The numPatients’, ICUs’, and management of end-of-life characteristics are summarized in Supplemental Table Among the 475 patients included, 398 nurse scores and 417 physician scores were analyzed.The response rates were 441/475 (92.8%) for nurses and 446/475 (93.9%) for physicians. Among these, respectively 398/441 (90.2%) and 417/446 (93.5%) were fully completed, allowing score calculation and analysis.p 0.28), but a center effect was detected for physicians’ scores (p < 0.001).The median global CAESAR score was 62/75  for nurses and 64/75  for physicians; the proportion of surveys with scores higher than 60/75 were 248/398 (62.3%) for nurses and 313/417 (75.1%) for physicians. There was no center effect for nurses .Job strain was evaluated among 231 physicians and 379 nurses. Ten percent of physicians had a positive job strain. Among the 3 domains , the most frequent complaint for physicians was a high job demand (38.2%), but there was no association between the CAESAR score and job strain for physicians. Twenty-seven percent of nurses had a positive job strain. The most frequent complaint for nurses was a low decision control (54.2%) and a high job demand (44.5%). Low decision control was associated with lower CAESAR score for nurses  versus 63 , This multicenter study allowed us to develop and validate two 15-item CAESAR questionnaires (CAESAR-P and CAESAR-N) designed to measure self-reported experience of caregivers about patients’ end-of-life in the ICU, as we did previously for relatives [Both tools (for nurses and for physicians) showed good internal consistency and a single dimension. The global score was high for both nurses [62/75 ] and physicians [64/75 ], reflecting global satisfaction regarding dying and death in the ICU, in keeping with previous studies . The nurPrevious studies aiming at evaluating quality of dying and death , 9, 25 uIn France, nurses’ perception of dying and death was evaluated over 10 years ago in hospitals  and in IIn our current study, nurses’ experience of the patient’s end-of-life in the ICU was worse in case of conflict with physicians. In a previous European study about ICU conflicts , end-of-For both nurses and physicians, a better experience of end-of-life was associated with an absence of invasive care (considered aggressive in this setting): for nurses, absence of invasive care such as surgery, chest tube, or bronchoscopy; for physicians, decision to withhold or withdraw treatment, and absence of cardiopulmonary resuscitation—factors concordant with other study results , 9. AbseFinally, good communication with family members and their presence in the ICU were associated with a better experience of patients’ end-of-life for nurses and physicians  and are key elements in end-of-life care, as shown in previous studies , 25, 29.This study has several limitations. First, all participating ICUs were in France and the findings may not be pertinent in different cultural settings. However, the large number of participating ICUs and clinicians, the very high response rate, and the validation of the results in a reliability cohort support the robustness of the data. Second, authors are aware of the potential issues with using an older dataset, specifically the potential changes of EOL practices across the world. However, use of the tool will permit to compare and describe changes across time and countries. Third, the global score was high in both groups of clinicians and the score differences were low and may not be meaningful to clinicians. However, this is true for other end-of-life tools used in Europe, such as the QODD . Lastly,In summary, we described and validated two new instruments for assessing nurses’ and physicians’ experience of end-of-life in the ICU. Our study shows factors associated with a better experience that include quality communication, both with family members and inter-professional communication and collaboration; family presence in the ICU; and avoidance of aggressive care. These results will help design future interventional studies aimed at improving end-of-life care in the ICU.Additional file 1: Supplemental Table 1. Characteristics of ICU, end-of-life management and patients.Additional file 2: Supplemental Table 2. Characteristics of physicians and nurses.Additional file 3: Supplemental Table 3. Psychometric validation of the physician questionnaire: distribution of individual item scores.Additional file 4: Supplemental Table 4. Psychometric validation of the nurse questionnaire: distribution of individual item scores.Additional file 5: Supplemental Table 5. Psychometric validation of the physician questionnaire: Measurement error: internal consistency.Additional file 6: Supplemental Table 6. Psychometric validation of the nurse questionnaire: Measurement error: internal consistency.


PD patients had more frequent previous ICDs history, premorbid impulsive personality and antidepressant treatment (p < 0.05) compared with CS. In PD, patients with ICDs/CBs presented younger age at disease onset, more frequent history of previous ICDs and premorbid personality (p < 0.05), as well as higher comorbidity with nonmotor symptoms, including depression and poor quality of life. Treatment with dopamine agonists increased the risk of ICDs/CBs, being dose dependent (p < 0.05). As conclusions, ICDs and CBs were more frequent in patients with PD than in CS. More nonmotor symptoms were present in patients with PD who had ICDs/CBs compared with those without. Dopamine agonists have a prominent effect on ICDs/CBs, which could be influenced by dose.The study was aimed at analysing the frequency of impulse control disorders (ICDs) and compulsive behaviours (CBs) in patients with Parkinson’s disease (PD) and in control subjects (CS) as well as the relationship between ICDs/CBs and motor, nonmotor features and dopaminergic treatment in PD patients. Data came from COPPADIS-2015, an observational, descriptive, nationwide (Spain) study. We used the validated Questionnaire for Impulsive-Compulsive Disorders in Parkinson's Disease-Rating Scale (QUIP-RS) for ICD/CB screening. The association between demographic data and ICDs/CBs was analyzed in both groups. In PD, this relationship was evaluated using clinical features and treatment-related data. As result, 613 PD patients  and 179 CS  were included. ICDs and CBs were more frequent in PD (ICDs 12.7% vs. 1.6%,  Within the ICDs spectrum, pathological gambling, compulsive shopping, hypersexuality and compulsive eating are included. CBs are those related to the performing of aimless repetitive stereotyped complex tasks, known as punding, hobbysm, excessive and aimless wandering  and dysregulation dopaminergic syndrome (DDS)5. Comorbidities such as tobacco consumption and depression are well-established factors linked to ICDs. However, the directionality of the association between ICDs and affective disorders is not still clear8. Epidemiologic factors such as sex, age and age at onset could influence the incidence of ICDs and CBs1. Nevertheless, it is controversial whether these disorders are exclusively linked to the first years of disease evolution when patients still have an integrum ventral striatum2, or whether they are associated with longer exposure to dopamine treatment3. On the other hand, estimated prevalence rates across countries range from 10 to 39%4. This wide range depends on the study characteristics, but also on the population features, being therefore important the assessment of specific populations.The etiopathology of these disorders is not simple. Demographic factors, premorbidities, comorbidities and genetic factors can influence the risk of ICDs and CBs8. Thus, under external dopaminergic stimulation by DRT, the regulation of mesocorticolimbic pathway activity might be inappropriate, inducing an aberrant response to reward tasks and impulsivity traits9. Despite this fact, not all patients undergoing DRT treatment have ICDs or CBs, suggesting the action of a neurobiological substrate leading to individual susceptibility5.In addition, it is known that dopaminergic replacement therapy (DRT) plays an important role in ICDs and CBs, given their prevalence in untreated de novo patients is the same as in control subjects (CS)11. However, there is controversy regarding DA dose and ICDs, given it is not fully understood whether the ICDs are associated with the DA dose alone or in combination with the dose of levodopa and other dopaminergic drugs12.Dopamine agonists (DA) have been shown to be a principal drug involved in ICDs, and the association appears to be dose dependent14.Parkinsonian features have also been associated with ICDs. Not only can cognitive performance in patients with ICDs be impaired, but executive dysfunction as well as impairment in the reward-related decision process and set shifting can be presentTaking into account the factors influencing ICDs and CBs in PD, we sought to assess clinical features linked to these disorders in the COPPADIS-2015 population of PD and CS. We especially aimed to evaluate the impact of DRT and the influence of clinical features on these disorders in the PD population.15. The project is conducted in accordance with the standards for Good Clinical Practice and the fundamental ethical principles established in the Declaration of Helsinki. Approval of the Ethics Committee at each center was obtained  as a Post-authorization Prospective Follow-up study with the code COH-PAK-2014-01.The participant data was drawn from the baseline assessment of the COPPADIS-2015, an observational, descriptive, 5-year follow-up, nationwide (Spain) evaluation study15.A total of 694 patients with PD  and 207 CS  were included in the COPPADIS study16. For DDS, we used the investigator criterion, given there is no established cut-off.The Questionnaire for Impulsive-Compulsive Disorders in Parkinson’s Disease Rating Scale (QUIP-RS) was assessed in both patients with PD and CS for screening of ICDs and CBs Demographic data concerning educational level, marital status, smoking, alcohol intake, premorbid impulsive personality, previous ICDs and family history of ICDs were collected for both groups (patients with PD and CS).15. Baseline data were used for the analysis.In the PD group, a cross-sectional assessment was performed for each patient according to the study methodologySubanalyses of the group with an age at disease onset younger than 50 years and of the DA treatment group were also performed.Parametric univariate tests (t-test and chi-squared test) and nonparametric univariate tests (Wilcoxon test) were used to compare demographics and QUIP-RS results between the CS and the patients with PD. A logistic regression analysis (controlling for sex and age) was also used.In the PD group analysis, univariate and multivariate analysis were used to compare demographic variables, motor and nonmotor symptoms and dopaminergic treatment variables between patients with PD with and without ICDs/CBs. While controlling for age, age at disease onset and disease progression time, a multivariate linear regression analysis was used for the quantitative variables and a logistic regression analysis was employed for the categorical variables. Finally, the association between ICDs severity and treatment was analysed using the QUIP-RS score as the outcome variable and dopaminergic treatment variables  as independent variables in multivariate linear regressions . All the statistical analyses were conducted in R for Statistical Computing, version 3.5.1.p < 0.001; CBs 7.18% vs. 1.67%, respectively, p = 0.01). The most frequent ICD was compulsive eating in both groups and hobbysm-punding in terms of CBs . In the CS, the presence of hypersexuality was higher in the male participants, but it was not significant .The analysis split by sex showed a higher presence of hypersexuality in male patients with PD  and were more often men . Premorbid impulsive personality  and previous ICDs  were more frequent among patients with PD than CS, in addition to them having a higher rate of antidepressant treatment .The demographic characteristics of the sample are shown in Table p = 0.01) and premorbid impulsive personality   and at disease onset   (Table p < 0.001). In addition, when the NMSS subdomains were analysed, the cardiovascular, sleep/fatigue, mood/cognition, attention/memory, urinary, sexual and miscellaneous domains had a higher impact on the patients with ICDs . In terms of mood, patients with ICDs showed higher rates of depression . Analyzing the subtype of depression according to the Beck Depression Inventory, patients with ICDs showed a higher ratio of subclinic and minor depression compared to those without ICDs. Anxiety, euphoria and irritability were the features specifically associated with patients with ICDs.Concerning the PD features according to the presence of ICDs, the motor characteristics were similar in both groups, except for a higher score in the freezing of gait (FOG) scale in patients with ICDs .Apart from the NMSS sleep score, the Parkinson’s Disease Sleep Scale showed that patients with ICDs presented with poorer sleep quality, as shown by a lower score , we did not find differences between the patients with and without ICDs.p < 0.001). Analysing the domains of the PDQ-39, ICDs had a higher impact on patients in terms of mobility, activities of daily living, body discomfort, communication, cognition and emotional well-being   was poorer in patients with ICDs   and Ropirinole the highest  . When we compared the type of DA with the non-use of DA and the severity of ICDs/CBs using the QUIP-RS total score, Ropinirole was associated with the greatest significant increase in severity . This result remained significant after correcting for confounding variables such as age, age at disease onset and disease progression time . Moreover, the increased QUIP-RS total score in patients with ICDs undergoing Ropinirole treatment was significant , but not for patients with other types of DA . On the other hand, we observed that the QUIP-RS total score was significantly increased according to DA equivalent dose  and daily levodopa equivalent dose , but not with levodopa dose alone   and differences on the cultural level. Other features such as a premorbid impulsive personality, non-motor symptoms according to NMSS and BDI and poorer quality of life were similar to the entire sample. In the same line with the global analysis, a higher rate in the FOG scale was shown in the group under 50 years old . The proportion of ICDs found in our sample was similar to that reported in the DOMINION study2, which is the largest PD prevalence study to date, in which they found ICDs in 13.2% of cases, in accordance with our study and with other European works17. This result is in contrast to the high rate of ICDs found in some other European studies, including in the Spanish population11. The prospective Italian study, ICARUS, had found a rate of ICDs at the basal evaluation of 29.3%, which was higher than in our baseline assessment in COPPADIS3. These discrepancies could partially be explained by the sociocultural differences in both groups and by the differing tools used for ICDs screening. In the ICARUS study, the Minnesota Impulsive Disorders Interview (MIDI) and QUIP were used, whereas in COPPADIS, the QUIP-RS was applied. Following the MDS recommendations, QUIP-RS has more accurate clinimetric properties for application in PD, being “recommended,” whereas MIDI lacks some criteria validity when compared with the QUIP-RS18. In addition, ICDs screening using QUIP could overestimate ICDs frequency since Papay et al. found that, up to 40% of patients who scored positive for ICDs on the QUIP did not fulfil the diagnosis of ICDs19. However, the QUIP-RS used in COPPADIS has stricter ICDs diagnostic criteria; thus, the proportion found in our study could be more accurate for the identification of ICDs.The objective of the present study was to assess ICDs and CBs in patients with PD and CS, as well as the clinical features linked to these disorders in both groups. We found that in the patients with PD, ICDs and CBs were more frequent compared with the CS . It would be more accurate to evaluate the whole UPDRS part II to establish the handicaps in the performance of the daily activities according this scale. To partially compensate this issue, we assessed the PDQ-39 that also included activities of daily living.This study has some limitations. The presented data correspond to the cross-sectional evaluation at baseline of COPPADIS cohort, nevertheless, the assessment of the ICDs/CBs behaviour over time would be more informative. This issue would be overcome in the future since the follow up of this cohort is on-going and we will obtain the longitudinal dataICDs and CBs are common nonmotor features in patients with PD compared with CS. Some premorbid features such as a previous impulsive personality act as risk factors for developing these disorders. DRT had a large impact on ICDs, and DAs were the drugs with the greatest effect. However, patients with ICDs had a high burden of nonmotor symptoms belonged to mood, cardio-vascular, sleep, urinary domains among others as well as a poor quality of life. Hence, apart from treatment, patients might have a genetic background that could influence the presence of a prominent non-motor phenotype.Supplementary Information.


Ultrasound imaging measurements who reported statistically significant differences were EI (p = 0.001) and standard deviation (SD) (p = 0.001). MTh and EV variables did not show statistically significant differences . Moreover, reliability values for the MTh (ICC = 0.911), EI (ICC = 0.982), SD (ICC = 0.955) and EV (ICC = 0.963). Based on these results the intra-rater reliability was considered excellent. Conclusion: Athletes with a central tendon injury of soleus muscle showed a lower EI when they were compared to healthy athletes. The echogenicity showed by the quantitative ultrasound imaging measurement may be a more objective parameter for the diagnosis and follow-up the soleus muscle injuries.Background: the aim of this study was to compare the echotexture of patients with soleus muscle injury and age matched controls. Methods: a sample of 62 athletes was recruited at the private clinic and was divided in two group: a healthy group (n = 31) and a soleus pathology group whose athletes had soleus muscle injury, located in the central tendon (n = 31). The muscle thickness (MTh), echointensity (EI) and echovariation (EV) were analyzed. An intra-rater reliability test (Intraclass Correlation Coefficient-ICC) was performed in order to analyze the reliability of the values of the measurements. Results: Sociodemographic variables did not show statistically significant differences ( The connective tissue plays an important role within the lower limb muscles, mainly rectus femoris, biceps femoris, gastrocnemius and soleus. This structure may be named in different terms, as intramuscular tendon, connective tissue, central tendon or aponeurosis and also present variability between individuals .Soleus muscle pathologies were presented in several sports modalities . The soleus muscle is part of the posterior region of the leg . HistoriBalius et al. argued that the location of five possible points in the soleus muscle where injuries may be identified: myofascial areas (anterior and posterior stress areas) and musculotendinous junction locations  . In all Recent research suggests that sometimes soleus injuries do not have a proper diagnosis which has consequences on its evolution, increasing the athletes’ risk of reinjury ,8. SeverUltrasonography is an accessible, noninvasive and economical tool that allows us to obtain images for the diagnosis of musculoskeletal injuries by clinicians . EchogenWe hypothesize that the athletes with soleus pathology, located in the central tendon (CT), may show differences in soleus echotexture (EI and EV) variables with respect to healthy athletes because of the appearance of disrupted intra-muscular structure . TherefoA prospective, cross-sectional, case-control study was carried out in an athletic population according to the Strengthening the Reporting of Observational Studies in Epidemiology Statement (STROBE) recommendations.The Declaration of Helsinki was respected throughout the study and also the study was approved by the Ethics Committee of University Hospital Virgen Macarena-Virgen del Rocio (Reference code: 06/2019). In addition, all the individuals signed written informed consent form before participating in this study.The sample size calculation was carried out with G*Power software by the difference between healthy group and patient group using the EI variable of a pilot study (n = 14) divided into two groups (mean ± SD), 7 subjects with soleus pathology (49.43 ± 13.59) and 7 individuals for the healthy group (59.55 ± 17.37). An α error of 0.05, a power of 0.80 and effect size of 0.64 with 1 tailed hypothesis were used for the sample size calculation. Finally, a total sample of 62 subjects was calculated.A total sample of 62 subjects were recruited and divided based on the echogenicity of the soleus muscle in both groups. Precisely, a soleus muscle injury, located in the central tendon, was considered in the present study, according to the following parameters: had symptoms for at least six months; and a self-rated progressive tightness score greater than or equal to 4 cm during sport activities or during the soleus stretching, measured using the numeric rate scale (NRS) . The recThe selection criteria for the patients were (a) age greater than 18 years old; (b) athletes who participated in sports that involve lower limbs; (b) at least of five-years practicing the sport; (c) training at least two hours, four days a week; (d) non-acute lower limb pathology at least six months ago; (e) non low back musculoskeletal disorders. Furthermore, inclusion criteria to be included for the healthy group was being a healthy athlete; and for the pathological group, having a clinical diagnosis of soleus muscle injury, characterized by muscle stiffness and lack of elasticity when stretching the muscle and corroborated by an experienced clinician.Clinical and demographic variables were recorded, such as age, height, weight, body mass index (BMI), sex, dominance side, injured side, sport training (years) and average sport hours per week. Severity of progressive tightness during sport activities or during the soleus stretching and pain level at palpation in the injured location was evaluated using NRS .All US imaging evaluation was carried out by an experienced therapist, who was blind to the subjects’ muscle status, with more than 15 years of experience in US imaging. A high-definition US with a 6-15 MHz linear transducer  was employed to perform all US images. For the classification into either the healthy or pathology group, US examination was carried out in a prone position with the feet hanging over the end of the table. In addition, the transducer was located in the transverse plane at the central tendon in order to describe the echogenic pattern . US imagQuantitative US variables, including thickness, EI and EV were taken in both groups. The muscle thickness (MTh) was described as the distance between superficial and deep fascia of the muscle. The imaging analysis was developed offline with the ImageJ software  by a blinded examiner. Moreover, according to research by Fukumoto et al. , an ROI The ROIs’ size were the same in all subjects and the position was the one that best represented the characteristics of the muscle. According to Caresio et al., the authors knew that the muscle echo-intensity depends on different factors such as ROI size, shape and location . To minit test and median, interquartile range (IR) with Mann-Whitney U test were carried out for parametric and non-parametric data, respectively. In addition, Levene’s test was employed to assess the equality of variances. Moreover, the intraclass correlation coefficient (ICC) was performed to evaluate the intra-rater reliability of all the ultrasound measurements.The statistical analysis was performed using the SPSS package v.22.0 . First, Kolmogorov-Smirnov was performed to check the normality assumption. Second, a descriptive analysis was employed for all the individuals and separately in the two groups. Finally, a comparative analysis between healthy group and patient group was developed. Mean, SD with the Student-s p > 0.05). Ultrasound imaging measurements who reported statistically significant differences were EI (p = 0.001) and SD (p = 0.001). MTh and EV variables did not show statistically significant differences . Moreover, reliability values for the MTh (ICC = 0.911), EI (ICC = 0.982), SD (ICC = 0.955) and EV (ICC = 0.963). Based on these results, the intra-rater reliability was considered excellent.Considering the In the present cross-sectional study, the assessment of the echoestructure and echotexture, as well as the intra-rater reliability, of each variable was performed in the soleus muscle of athletes with and without pathology. The main finding of the present study was that echotexture features differentiated athletes with a CT injury of soleus muscle from the healthy athletes. Specifically, the results showed that a lower EI in the pathology group compared with the healthy group and also no differences were observed regarding MTh and EV. In addition, an intra-rater reliability test was performed with the aim of increasing the reliability of the values of the measurements. Our scores showed a high reliability in all variables of the present study for this sample.Results reported in the present study are in line with previous studies which analyze the EI parameter in different muscle conditions. EI of the selected regions has been considered a useful biomarker reflecting musculoskeletal quality and disturbances related to pathological conditions. Heckmatt et al.  comparedCurrently, there is a high interest in studying soleus muscle characteristics due to its importance during sporting activities as well as for a better understanding of its pathological features ,6,31,32.Regarding the EV and MTh parameters, this study did not show differences between groups. Several authors hypothesized the possibility of developing a muscle biomarker with ultrasonography, for example, Martínez-Payá et al. found a decrease in the EV of the quadriceps femoris, tibialis anterior, biceps brachialis and forearm flexor group in patients with amyotrophic lateral sclerosis. Almazán-Polo et al.  observedThe most important clinical application of the present study highlighted the benefits of the EI as a more objective guideline for the classification of the echogenic pattern applied to the muscle. In addition, EI may be used in future studies for the management and follow up of musculoskeletal disturbances, as well as in preventive and management exercise programs.Several limitations were observed for the present study. Despite the excellent intra-rater results for the reliability, no inter-rater procedure was developed and future inter-rater reliability studies are still needed in order to standardize protocols and methods for quantitative ultrasound examinations in muscle and soft tissues. Thus, further research could be considered in musculoskeletal disturbances and other areas. Moreover, future studies are needed for EI and EV variables as descriptors of the soft tissues features.The use of this new echotexture parameters measured with ultrasound imaging is a promising biomarker in musculoskeletal disorders. The authors suggest that EI could be employed to differentiate athletes with soleus muscle pathology from healthy athletes due to soleus tear being characterized by a minor EI. The echogenicity reported by the quantitative US measurement may be an objective parameter for the diagnosis, management and follow-up of soleus muscle injuries.


The purposes of this study were to explore the effect of obstructive sleep apnea-hypopnea syndrome (OSAHS) on the voice by analyzing the acoustic parameters between patients with OSAHS and those without OSAHS and to compare acoustic analyses performed by two software programs (MDVP and Praat).Patients with OSAHS (n = 75) and normal controls (n = 46) were asked to produce a sustained sound of the vowel /i/ and were analyzed with electroglottography (EGG), MDVP, and Praat software. A self-rated scale  and acoustic parameters were compared.There were no statistically significant differences in the fundamental frequency (F0), jitter, shimmer, noise/harmonic ratio (NHR), contact quotient perturbation (CQP), or contact index perturbation (CIP) between the patient group and the normal group. The VHI-10 values were significantly increased in patients with OSAHS. The receiver operating characteristic (ROC) analysis suggested that the shimmer obtained from MDVP and Praat possessed relatively high accuracy in differentiating patients with OSAHS from healthy individuals. The results for F0, jitter, shimmer, and NHR were significantly different between MDVP and Praat in OSAHS patients. In normal persons, there was a significant difference in NHR; however, no significant differences were found for F0, jitter, or shimmer between the two software programs. The results demonstrated that high correlations were found between values obtained by both software programs.Patients with OSAHS were prone to vibration irregularity, incomplete glottal closure, hoarseness, and other vocal problems. The two acoustic software programs present different values of acoustic measures. There was a strong correlation and consistency between the parameters calculated by the two software programs. Obstructive sleep apnea-hypopnea syndrome (OSAHS) is characterized by the narrowing and collapse of pharyngeal structures during sleep, resulting in intermittent hypoxemia and sleep disruption. One or more sites of obstruction may include the soft palate, tongue, lateral walls, and/or epiglottis . OSAHS hOSAHS patients have structural abnormalities in the upper airway , which lead to structural changes, and these changes affect voice production and resonance and contribute to abnormal voice features , 11. It In this study, we focused on acoustic analysis software to assess OSAHS patients’ voice disorders by comparing the results of the acoustic analyses performed by two software programs: MDVP (Kay Elemetrics) and Praat (Boersma and Weenink). The two software programs are commonly used for acoustic analysis in clinical and research settings, while MDVP is the most used and cited acoustic analysis software . Praat iThe aim of this study was to assess acoustic features of voice in OSAHS patients and to identify whether the results obtained from the same voice samples are comparable and/or correlative between the MDVP and Praat acoustic computer programs.This was a prospective study performed at Tianjin First Central Hospital. Two groups of subjects were recruited. The patient group consisted of 75 patients who were diagnosed with severe OSAHS (apnea-hypopnea index > 30/h)  by polysEach voice sample was recorded while the examiner was under identical conditions in a sound-treated room with ambient noise below 50 dB. The person took a seat in front of the microphone placed 15 cm from their mouths. To reduce intrasubject variability, 5 samples of sustained vowel /i/ sounds at a comfortable pitch, constant amplitude, and flat tone were used for each subject. All voice samples were obtained using EGG and Speech Lab Model 4500 (Kay Elemetrics Corporation) at a sampling rate of 50 kHz and 16-bit quantization. Acoustic voice analysis was performed twice: once using MDVP and once using Praat. The fundamental frequency (F0), jitter, shimmer, and NHR were compared between MDVP and Praat. Additionally, CQP and CIP were obtained by EGG.The Turkish version of the Voice Handicap Index-10 (VHI-10) was used to evaluate voice quality. The VHI-10 is a questionnaire that assesses a patient’s perception of the physical, emotional, and functional aspects of their voice . SubjectIn the present study, all acoustic voice parameters were evaluated in the morning, which prevented the voices of the participants from being affected by circadian changes .U test was used to compare the four markers between normal persons and patients. The receiver operating characteristics (ROC) approach was also applied to evaluate the diagnostic value of voice parameters in OSAHS patients, and the results were estimated by calculating the area under the ROC curve (AUC), sensitivity, and specificity according to standard formulas. Spearman’s rank correlation coefficient (Spearman’s rho) and Bland-Altman analysis were performed to assess the correlation and consistency of F0, jitter, shimmer, and NHR between the two acoustic voice programs: MDVP and Praat.Statistical analysis of the data was performed using IBM SPSS statistics 19. The Mann-Whitney The results of the acoustic analysis performed using the two software programs in normal persons and OSAHS patients are shown in Table In the MDVP analysis, there were significant differences in jitter, shimmer, and NHR between the two groups. The ROC curve analysis is shown in Fig. For Praat, significant differences were also found in jitter, shimmer, and NHR between the normal group and patient group. The ROC curve analysis showed that the AUC of jitter was 0.736 (95% CI 0.648–0.825), and the cutoff was 0.568, with a sensitivity of 71.05% and specificity of 71.7%. The AUC of shimmer was 0.869 (0.804–0.934), and the cutoff was 3.47, with a sensitivity of 81.5% and specificity of 80.1%. Shimmer possessed relatively high accuracy in differentiating OSAHS patients from healthy individuals. The AUC of NHR was 0.689 (0.590–0.789), and the cutoff value was 0.035, with a sensitivity of 73.6% and specificity of 63.0% Fig. .P < 0.001).VHI-10 scores and acoustic parameters from EGG are also depicted in Table The results of the acoustic analysis performed using the two software programs for the two study groups are presented in Table In the normal group, statistical analysis revealed a significant difference between the results obtained from the two software programs for NHR. No significant differences were found between the two programs for the F0, jitter, or shimmer Table . HoweverIn the OSAHS patient group, there were significant differences between the results obtained from the two software programs for F0, jitter, shimmer, and NHR. The F0, jitter, shimmer, and NHR determined with MDVP showed a strong correlation with those determined with Praat Fig. .Fig. 3PaThe results of the MDVP and Praat consistency tests are shown in Fig. It is known that the upper airway structure plays an important role in voice resonance and articulation . ObstrucThis study shows that the vocal cord vibration of OSAHS patients is irregular, and the fundamental frequency perturbation, amplitude perturbation, and vibration-related acoustic parameters, such as jitter, shimmer, NHR, CQP, and CIP, are significantly higher than those of the normal group. Jitter and shimmer reflect the grade of hoarseness and roughness of the speech spectrum. NHR measures the aperiodic noise present in the analyzed signal, which can effectively reflect glottal closure. The CQP and CIP could reflect the regularity of the vocal cord vibration and the periodic change of the vocal cord contact section. A reduction in voice quality was also found in patients by using the Voice Handicap Index-10 (VHI-10).Our study also suggests that the different software programs present different values of acoustic measures. Previous reports have shown that the MDVP and Praat analysis produce equivalent mean F0, whereas jitter and shimmer differ significantly –23. HoweThe results of the study showed that MDVP appeared to differentiate the OSAHS group better than Praat. However, we also found a strong correlation between the values calculated by the two software programs. Previous studies have attempted to compare values of perturbation measures that were calculated using MDVP with those calculated using Praat. They also found a strong correlation between the values obtained by the two software programs.  ConsisteIn conclusion, OSAHS patients experience long-term mouth breathing and changes in the structure of the articulation area, which result in irregular vocal cord vibration, poor glottis closure, pronunciation difficulties, hoarseness, and other vocal problems. Therefore, in clinical treatment, attention should be paid to the patient’s voice changes, and mouth-breathing habits should be corrected to avoid changes in vocal cord tissue biomechanics and vocalization due to vocal cord dehydration. The two acoustic software programs present different values of acoustic measures, and MDVP appeared to differentiate the OSAHS group better than Praat. In addition, there is a strong correlation and consistency between the values calculated by MDVP and Praat. In some cases, either of the two software programs can be used to analyze voice data.


Staphylococcus aureus (SA) sequence type ST59 is the most commonly isolated sequence type in Eastern Asia. The origins and evolution of this strain type remains unclear and therefore we gathered a collection of ST59 isolates from Canada and mainland China for a detailed genetic analysis of the lineage. Bayesian inference phylogenomic analysis of our isolates, along with previously published ST59 sequences indicated that the lineage could be divided into 6 distinct subgroups (WGS-1 thorough 6), each having distinct molecular characteristics. Analysis also demonstrated the concurrent but separate evolution of North American and East Asian lineages, as well as the extensive diversification of the East Asian lineage. The presence of a mobile element structure (MES) was found to be the major difference between these two continental lineages, absent in all North American isolates, and present in all East Asian ones. Other mobile genetic elements, such as the Immune Evasion Complex (IEC), Panton Valentine Leukocidin (PVL), and Staphylococcal Cassette Chromosome mec (SCCmec), showed significant variability within each sub-group and likely represents local selective pressures rather than major characteristics defining the groups. Our analysis also demonstrated the existence of a more ancient ST59 sub-lineage from North America, which was MES negative and contained some of the earliest reported ST59 isolates. Combined with the existence of a MES negative isolate from Taiwan, predicted to have appeared prior to diversification of the East Asian lineages, these results hint at the possibility of a North American origin for the lineage, which gained hold in Eastern Asia following acquisition of MES, and subsequently diversified.Despite initially being described in North America,  Staphylococcus aureus (MRSA) is an important human pathogen that causes a wide variety of infections ranging from skin and soft tissue infections to endocarditis and sepsis  type IV, and predominantly negative for Panton-Valentine Leukocidin (PVL) gene carriage , scn  and sak (encoding staphylokinase), and is responsible for protecting the bacteria from the host’s immune response. Differing combinations of the IEC genes are suggested to be present in the differing lineages of ST59, and can be carried on ϕSa3 as in the Asian-Pacific clone, or on νSaβ as in the Taiwan clone  from Geneva and three isolates  from Malaysia were present in our strain collection and added to the study. Canadian PFGE reference strains CMRSA1-10 were provided by the National Microbiology Laboratory, Health Canada, Winnipeg, Manitoba, Canada.Fifty-eight spa typing  after digestion with Smal . Fingerpa typing , MLST , while PacBio reads were assembled by the McGill University Génome Québec Innovation Centre using the HGAP workflow, whereby short reads were used to correct long read, which were subsequently assembled  , whereby at least one representative was selected from each major branch in the PFGE tree, with major branches defined as sharing a 78% or greater Dice coefficient of correlation (DCC). Long sequence reads for 5 randomly selected isolates were obtained with Pacific Biosciences (PacBio) RSII sequencing technology.  Canada) . ST59 sede novo assembly was done and then the PVL genes identified with Virulence finder 2.0  was used to search for and identify the Immune Evasion Complex (IEC) genes, as well as Mobile Element Structure (MES) genes. For downloaded sequences with no isolate physically available, nder 2.0 , spa typr v. 1.0  and SCCmr v. 1.2 . The genommended .1. The default minimum coverage parameter value of 10 was used for threshold variant calling. The SNP-substituted genomes were run through Gubbins  modal estimate being 2005 (logP = −3) using it and all 173 dated genomes in 10M iterations of a gamma site model with an HKY substitution model and relaxed log normal clock.Tree convergence was confirmed using BEAST’s Tracer 1.7.1 program  using thC. elegans infection model, following established techniques . Killing rates were calibrated as [% deathIsolate −% deathM92(–ve control)]/[% death8325(+ve control) −% deathM92(–ve control)], with mean killing rates determined as the mean of 3–5 experimental replicates.Virulence was assessed on representative ST59 isolates using the chniques . Modificoli OP50 . L4, nemS. aureus, and for identifying different strain types. Within our complete collection of S. aureus strains, we identified 224 ST59 isolates, from which we generated a ST59 group specific PFGE figure , IEC, and MES, which were characterized for all the strains, with the results summarized in Because of the molecular heterogeneity seen within each of the PFGE groups, 47 representative isolates were selected for whole genome sequencing and a more detailed analysis. Several mobile genetic elements are proposed to be associated with evolution of the lineage, including SCCmec type, PVL carriage, and IEC type, with MES type being more random (summarized in Previous studies have suggested that ST59 evolved concurrently in both North America (NA), previously referred to as the United States lineage and East Asia (EA), with strains from each region belonging to separate and distinct sub-lineages , with moIn silico genome to genome distance calculations were done on the most closely positioned MRSA/MSSA pairs to demonstrate their high degree of relatedness, with the results summarized in As seen in C. elegans infection model, with the results presented in spa type, SCCmec type, PVL presence an IEC type, didn’t appear to correlate with C. elegans toxicity. While strains in WGS-1 with higher toxicity carried the sea gene, other strains with higher relative toxicity (such as GD1171) did not carry the gene, and PRE carried it but had low toxicity.Having defined the distinct ST59 sub-lineages and determining that they each have relatively homogenous molecular characteristics, we sought to determine if there were any differences in virulence between the groups. All 47 of our isolates, with multiple representatives from each of the 6 sub-lineages, had virulence assessed using the S. aureus CC59 has received a significant amount of interest in recent years due to its high prevalence in Asian countries. Previous studies have attempted to explain evolution of the lineage, not only identifying separate sub-lineages, but also identifying factors that contributed to expansion and success of the each of the sub-lineage. Studies by mec, and PVL contributed to diversification of the lineage. However, an important finding made by Ward et al. was the identification of two major clades of S. aureus ST59 (one from the North America and a second from East Asia), which were calculated to have evolved separately but concurrently, with no direct exchange between them. This dispelled the notion that the EA clade descended from the NA one, which had previously been suggested based on the earlier reporting of USA1000 (The origins and evolution of  USA1000 . To expaBayesian inference phylogenomic (BEAST) analysis of our 47 isolates, combined with previously described representative global isolates, allowed us to prepare an updated evolutionary hypothesis for the lineage, which is presented in T/IEC-C/PVL(+)], as is WGS-5 [MRSA IVa/IEC-C/PVL(+) or MSSA/IEC-C/PVL(−)] (A recent study by /PVL(−)] . Where omec and PVL are shared by isolates in both groups. The presence of MES in EA lineages has been attributed to the high levels of non-prescription antibiotic use seen in many Asian countries . However, as the ST59 lineage evolved the complexes appeared to contain progressively fewer of these genes. It has been suggested that IEC associated genes on ϕSa3 are associated with strain virulence (T-PVL(+); WGS-6) is documented to cause more severe infections (scn and chp), suggesting that its success is unrelated to the complexity of IEC. PVL may in fact be more tied to the virulence of the Taiwan sub-lineage, as was previously suggested , concluding that the acquisition of specific SCCmec types helped the clades achieve dominance in their local regions.A deeper look at factors that may be responsible for the dominance of ST59 in EA has focused on mobile genetic elements such as IEC, MES, SCCge in EA . Our resirulence , which cfections , represefections , and 71.fections . It is, uggested , as mostmec types (or lack of one) typified many of the sub-lineages, in each group where both MSSA and MRSA were seen, and often multiple SCCmec types were encountered. In fact, both MRSA and MSSA were identified as coexisting amongst the earliest reported isolates , in the earliest sub-lineage to have evolved (WGS-1). Genome to Genome distance calculation between MRSA and MSSA pairs within each sub-group indicated that they were closely related to each other, suggesting that they share the same genetic background and that SCCmec is readily being gained/lost from the strain. As such, our data suggests that while mobile genetic elements such as IEC, PVL, and SCCmec have types commonly associated with each of the sub-lineages, the traits cannot be used as rigid definers of the lineages, as was suggested by Our results indicated that while certain SCCC. elegans infection model. We were surprised to note that there was very little difference in toxicity between the ST59 sub-groups, despite the differences in IEC related virulence gene carriage. Overall, we found that the lineage had a low level of C. elegans toxicity, particularly when compared to other lineages such as ST398 (including isolates from the same region in Guangdong)  , USA300 angdong) , ST30 anangdong) . No discirulence . IEC is,mec, showed significant variability within each sub-group and likely represent local selective pressures, rather than major defining characteristic for each group. And finally, while our data cannot directly show that ST59 evolved in NA and was then transported to EA, the fact that the earliest appearing sub-group had NA characteristics [MES (-)] and was predominantly isolated in NA with some of the earliest recorded ST59 isolates, combined with the presence of a MES(-) isolate in Taiwan, predicted to have appeared prior to diversification of the EA lineages, hints that this may be the case. Further studies will of course be needed to confirm this supposition.Our study was able to construct an updated evolutionary story for the ST59 lineage. Our model not only included the co-evolution of North American and East Asian sub-lineages previously described in other studies, but also identified and older North American sub-lineage that was calculated to have emerged prior to the previously described NA/EA groups. We identified MES as being the major difference between NA an EA lineages, with NA sub-lineages showing minor expansion, while EA lineages showed significant diversification. Mobile genetic elements, such as IEC, PVL, and SCCThe datasets presented in this study can be found in online repositories. The names of the repository/repositories and accession number(s) can be found in the article/The protocols were approved by the University of Calgary Conjoint Health Research Ethics under the Certification No: REB13-0219, and the Ethics Committee of the First Affiliated Hospital/School of Clinical Medicine of Guangdong Pharmaceutical University under the Ethics No. 2011(1), respectively. Written informed consent was obtained from all participants.KZ conceived, designed, and supervised the work. J-AM, SL, AN, GD, and OO performed the experiments and analyzed data. PG constructed Bayesian inference phylogenomic tree. SC and JC collected and provided the clinical isolates. J-AM and KZ structured and drafted the manuscript. JC and KZ edited the manuscript. All authors reviewed and approved the 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.


Suicide is one of the most critical public health concerns in the world and the second cause of death among young people in many countries. However, to date, no study can diagnose suicide ideation/behavior among university students in the Middle East and North Africa (MENA) region using a machine learning approach. Therefore, stability feature selection and stacked ensembled decision trees were employed in this classification problem. A total of 573 university students responded to a battery of questionnaires. Three-fold cross-validation with a variety of performance indices was sued. The proposed diagnostic system had excellent balanced diagnosis accuracy (AUC = 0.90 [CI 95%: 0.86–0.93]) with a high correlation between predicted and observed class labels, fair discriminant power, and excellent class labeling agreement rate. Results showed that 23 items out of all items could accurately diagnose suicide ideation/behavior. These items were psychological problems and how to experience trauma, from the demographic variables, nine items from Post-Traumatic Stress Disorder Checklist (PCL-5), two items from Post Traumatic Growth (PTG), two items from the Patient Health Questionnaire (PHQ), six items from the Positive Mental Health (PMH) questionnaire, and one item related to social support. Such features could be used as a screening tool to identify young adults who are at risk of suicide ideation/behavior.  According to the World Health Organization’s (WHO) report , the ratAs one of the leading causes of death for young people, suicide, and measures to prevent it are among the most critical health priorities in the “Mental Health Action Plan 2013–2020” . To prevExposure to trauma and post-traumatic stress disorder (PTSD) is yet another psychological issue that is significantly associated with suicide ,16,17,18A time gap between suicide ideation and suicide attempt is critical in mental healthcare. Many researchers found a relationship between experiencing a traumatic event and post-traumatic stress disorder (PTSD) with higher risk of suicide ideation, and there is an association between the number and type of traumatic events and suicidality . SeveralAlthough some studies attempted to predict the risk of suicide by investigating risk and protective factors, there is little knowledge of clinical prediction rules to predict university students’ suicide ideation/behavior. Since suicide is a multi-faceted phenomenon that has complex risk and protecting factors to predict suicide ideation/ behavior, we need to use algorithms that “model complex relationships among a large number of factors” . RecentlMoreover, a reliable medical diagnosis system must meet the following simultaneous conditions, namely sensitivity, specificity, positive predictive value, and diagnostic odds ratio, higher than 80%, 95% , 95% 4646, and 1p = 26%) , and 0.661 , for PMH and MSPSS, respectively. The Suicide Behaviors Questionnaire-Revised (SBQ-R)  containsEthics approval of the research was granted by the University of Isfahan’s Ethic Committee with the number IR.UI.REC.1399.008 . To ensure confidentiality, the questionnaires were anonymous. Respondents could refrain from answering anytime they wished. There was no incentive for participation. However, participants could write an email and receive their test interpretations.Choosing informative, discriminating, and stable features is a crucial step for designing a classification model . In thisEnsemble learning is a concept that multiple weak learners are trained to solve the problem. They are then combined to achieve better performance. The stack ensemble methods use various weak learners separately, and a meta-model is learned to predict outputs on top of all weak learners. In this paper, decision tree (DT) C4.5 was used as a weak learner, and selected features were used for training a meta DT classifier, which was created of stacked ensembled DT . Each DTThe demographic variables, and the questionnaires PCL-5, PTGI, MSPSS, PHQ-9, and PMH were used as the inputs of the prediction system, while the suicidal high and low-risk groups  were used as the binary output.Three-fold cross-validation was used to guard against testing hypotheses suggested by the data (Type III errors) . The folfficient , the Cohfficient , diagnosfficient , respectfficient , the CI fficient ,81.M = 24.45 (SD = 6.651) years, and 72.1% were female. In addition, 63% were undergraduates, while 37% were graduate students. Furthermore, 60.7%, 5.8%, 23.6%, and 7.2% were liberal arts, basic sciences, engineering sciences, and medical sciences students. Among the 573 participants, 25% were at high-risk of suicide (SBQ-R ≥ 8). The characteristics of participants belonging to the low and high-risk groups are shown in The average age of all participants was The selected features, along with their importance, are shown in The performance of the prediction system is displayed in The performance of the proposed diagnosis system could be provided based on the interpretations of the overall AUC, MCC, DP, and K(C) scores as the following. The proposed prediction system has excellent balanced diagnosis accuracy with a high correlation between predicted and observed class labels, fair discriminant power, and excellent class labeling agreement rate.Moreover, a medical diagnosis system is clinically reliable if Se ≥ 80%, Sp ≥ 95% , PPV ≥ 9To the best of our knowledge, this is the first study of its kind to make an accurate diagnosis of suicide ideation/behavior using robust ensemble learning in a university student sample in the MENA region. According to the results, we were able to identify 23 items out of 92 items that could be used to create a screening tool to distinguish young adults with high and low suicide ideation/behavior. The screening tool may have important implications for universities and policymakers that are responsible for university students’ suicide prevention and treatment. Based on our results, exposure to trauma, history of psychological illness, PTSD symptoms (9 items out of 20 items in PCL-5), positive mental health (6 items out of 9 in PMH), depression symptoms (3 items out of 9 in PHQ), post-traumatic growth (2 items out of 21 in PTGI), and social support (1 out of 12 items in the MSPSS) were recognized as the main variables for predicting suicide ideation and attempt.The first important findings showed that exposure to trauma and PTSD symptoms could diagnose suicide ideation/behavior. The main items here were the type of exposure to trauma (first-hand or witnessing trauma) and PTSD symptoms. These findings complement previous findings showing that there is a strong link between trauma exposure and suicide risk ,36,85,86Psychological illness is another input feature in three prediction systems that could predict suicide ideation/behavior. This is in line with previous studies that found a strong relationship between psychological distress and suicide ideation ,90,91,92In our study, a lack of positive mental health, i.e., psychological and subjective well-being, was selected by the proposed diagnostic system, as another main predictor for suicide ideation/behavior. From the nine items of the PMH scale, six items were selected by the system as effective factors on suicide ideation/behavior. This is in agreement with some previous studies that have shown that positive mental health moderates the effects of stressful life events on suicidal thoughts . In seveOne of the notable factors in our research was post-traumatic growth (PTG) items. According to the literature, PTG moderates suicide risk among students who experienced traumatic events , and lowOur system did not select social support as one of the well-studied buffering factors in the suicide literature ,96 as a The advantage of our study is the high accuracy of the proposed statistical model. It is the first study to analyze the comprehensive factors related to suicide ideation/behavior and to design a diagnostic system for university students at risk of suicide ideation/behavior in the MENA region. However, since a pure student sample was studied with the majority of the participants being female, one should refrain from generalizations to other samples. Furthermore, all variables were assessed using self-report measures. This is of great advantage with regard to the development of an easy-to-administer screening instrument. At the same time, bias effects cannot be ruled out. Therefore, it might be beneficial to cross-validate the described diagnostic model using interviewer-based assessments in future studies. Another limitation of the study is related to the distribution of gender and major among the sample. According to the Statistical Centre of Iran , the numAs mentioned before, the findings of this study have practical implications for policymakers and universities. Concerning the findings that trauma exposure and PTSD symptom were selected as the main variables in diagnosis of suicide ideation/behavior, routine assessment of cognitive distortion and PTSD symptoms among students at risk is important. The majority of the Iranian universities have free-of-charge counseling service. Having a trauma-informed counseling service to identify at-risk students is essential. As a lack of positive mental health and a lack of some of the post-traumatic growth’s items were selected as other predictors of suicide ideation/behavior, creating national programs to enhance PMH and PTG for the at-risk students is recommended. Based on the findings of this research, a screening tool will be designed, and the application can be used in universities to screen high-risk and low-risk students for suicide ideation/behavior for future prevention and intervention programs. Prospective studies should follow to further investigate the efficiency of the described screening tool.To the best of our knowledge, this study is the first study to use robust ensemble learning to reliably and accurately diagnose suicide ideation/behavior among university students in the MENA region. Based on our results, we found that trauma exposure and PTSD symptoms, psychological problems , low mood, and low self-esteem can diagnose students that are at high risk for suicide ideation/behavior. We have also found that positive mental health can strongly affect suicide ideation/behavior, meaning that individuals that had high scores in PMH had less suicide ideation/behavior. People who appreciate life also showed less suicidality. Therefore, two items related to post-traumatic growth will be included in the final screening tool. Social support was only necessary for terms of family support in our prediction system. This study was performed to create a screening tool to identify university students at risk of suicide ideation/behavior and to help policymakers and universities to make appropriate on-time prevention and early intervention programs.


TBC1D24 and ATP6V1B2 genes, both of which are involved in endolysosomal function. Because of its extreme rarity, to date, no detailed neuropathological assessment has been performed to establish clinicopathological relationships and, thereby, understand better the neurobiology of this disease in aged cases. Accordingly, the aim of the current study was to highlight the clinicopathological characteristics of a novel case with a presumable de novo mutation in the ATP6V1B2 gene from a neuropathological point of view. This Caucasian male patient, who died at the age of 72 years, presented all the typical cardinal signs of DOORS syndrome. In addition, behavioral alterations, pyramidal signs, and Parkinsonism were observed. The p.R506X pathogenic mutation identified in the ATP6V1B2 gene was responsible for the clinical phenotype. The detailed neuropathological assessment revealed a limbic-predominant tauopathy in the forms of argyrophilic grain disease, primary age-related tauopathy, and age-related tau-astrogliopathy. In summary, we present the first detailed clinicopathological report of a patient with DOORS syndrome harboring a pathogenic mutation in the ATP6V1B2 gene. The demonstrated tauopathy may be considered as a consequence of lysosomal and/or mitochondrial dysfunction, similar to that found in Niemann–Pick type C disease, which is another lysosomal disorder characterized by premature neurodegenerative disorder.DOORS  syndrome can be caused by mutations in the  TBC1D24 gene (ATP6V1B2 (p.Arg506X de novo heterozygous mutation), in DDOD syndrome was published in 2014 , or in clinical syndromes overlapping with the three entities mentioned above in patients harboring the p.E374Q or p.L398V mutations in a heterozygous state  guidelines , and seizures, in addition to specific craniofacial anomalies . The mosD24 gene . The fir in 2014 . Interesus state . Althougus state , there aidelines , with spThe Caucasian male patient reported here was born at term and his post-natal period was uneventful. Although the available data are limited, in addition to deafness, intellectual disability, and structural abnormalities (as detailed later), no other major alteration was reported with regard to his early childhood. He was not able to attend school, lived with his parents, and needed some assistance in daily activities in his later life. His family history was negative for the condition described above and no consanguinity was reported in the family. His medical history was relatively uneventful until the age of 52, when he was admitted to the neurological department after two generalized tonic–clonic seizures. No provoking factors could be identified. Subsequently, he was free of seizures until the age of 55, when he was admitted again to the neurological department after two additional generalized tonic–clonic seizures. Although electroencephalography (EEG) did not find any abnormalities, a diagnosis of epilepsy was established based on the presence of repeated unprovoked seizures, and carbamazepine treatment was initiated accordingly. Thereafter, only one seizure was reported. At the age of 63, resting tremor was identified during a regular check-up. At the age of 65, he was admitted to the psychiatric unit because of impulsive behavior and self-harm. Clonazepam and risperidone were initiated, providing appropriate symptom control. Between the ages of 65 and 70, he was admitted several times to the surgical unit because of incomplete or complete ileus events, which were managed conservatively, with the exception of one occasion. At the age of 66, he was admitted to the Department of Internal Medicine with the suspicion of slight cardiac decompensation. Echocardiography demonstrated a dilated aortic root, mitral prolapse, and minimal pericardial fluid, but an overall good myocardial function. The leg edema responded well to diuretics. At the age of 67, carbamazepine was changed to valproic acid without any known reason . At the age of 69, he was referred to the emergency department with a general malaise. The patient was admitted to the neurological department for a diagnostic workup of his unknown syndrome.On inspection, craniofacial dysmorphism  did not show parenchymal or vascular deposits (image not shown). Immunostaining for phospho-tau (AT8) revealed few neurons with diffuse fine granular immunoreactivity, similar to pretangles, and occasionally with small fibrillar conglomerates, similar to early neurofibrillary tangles (NFTs). Small amounts of neuropil threads were also detected. In the cingulate, there were no ballooned neurons and grain-like tau positivity was observed only occasionally. In the temporal cortex, a more pronounced neuronal tau pathology was detected and that was accompanied occasionally by granular/fuzzy astrocytes in the gray matter and several oligodendroglial coiled bodies in the white matter. A few grain-like structures were also observed. The parietal cortex showed a single neuronal tau pathology, whereas the occipital cortex did not show any tau pathology. No Aβ deposition was detected in these regions.The hippocampus and amygdala showed a prominent tau pathology in the form of NFTs , pretangles , and neuropil threads and grains , as well as granular/fuzzy astrocytes in the gray matter , thorn-shaped astrocytes in the white matter in the subependymal  and subpial areas, and oligodendroglial coiled bodies and threads in the white matter . The amyDespite the presence of a reduced vascular reserve capacity demonstrated by SPECT, the basal ganglia, and thalamus did not exhibit prominent vascular lesions apart from widened perivascular spaces; furthermore, there was a lack of inflammatory infiltrations. In the nucleus accumbens, several pretangles, and threads were observed, but no grains. Thorn-shaped astrocytes accumulated in the frontobasal white matter. The basal nucleus of Meynert showed a few NFTs.In the brainstem and cerebellum, the anatomical structures were well-preserved and there was no significant neuronal loss or vascular lesions. With the exception of a few NFTs and pretangles in the locus coeruleus, there was no tau pathology or α-synuclein pathology. The regional distribution of tau pathology is presented in Method used for immunohistochemistry. Formalin-fixed, paraffin-embedded tissue blocks (2.5 × 2.0 cm) were applied. The presence of astrogliosis/microgliosis/neuronal loss and the degree of deposition of various proteins were semi-quantitatively  evaluated in the following anatomical regions: frontal, temporal, parietal, occipital, and premotor cortices; basal ganglia; cerebellum; thalamus; amygdala; hippocampus; and the mesencephalon, pons, and medulla oblongata in the brainstem. In addition to Hematoxylin & Eosin and Luxol Fast Red staining, the following monoclonal antibodies were used for immunohistochemistry: anti-phospho-tau AT8 , anti-phospho-TDP-43 , anti-α-synuclein , anti-Aβ , and anti-p62 . The Dako EnVision Detection System, Peroxidase/DAB, Rabbit/Mouse , was used for the visualization of antibody reactions.Although the postmortem delay resulted in a considerable decrease in sample quality for transmission electron microscopy (TEM), the images obtained demonstrated a relatively preserved mitochondrial  and synaMethod used for electron microscopy. For ultrastructural examination, tissue specimens were fixed with 2% glutaraldehyde in phosphate-buffered saline containing 2.25% dextran (20 kD) overnight at 4°C. After fixation, the specimens were embedded in Embed 812  using a routine TEM embedding protocol. Ultrathin (70 nm) sections were prepared on an Ultracut S ultramicrotome  for TEM examination. After staining with uranyl acetate and lead citrate, the sections were observed using a Jeol 1400 plus TEM (Japan). Measurements of the largest diameter of mitochondria and synaptic vesicles were obtained and the average size was calculated.The clinical picture suggested the presence of DOORS syndrome underlying most of the alterations observed in the patient. After obtaining written informed consent from the legal guardian, genomic DNA was extracted from peripheral blood leukocytes via a standard protocol. Sanger sequencing was performed to screen for pathogenic mutations in the TBC1D24 gene, but no alterations were detected. Mutations in the ATP6V1B2 gene were assessed next, which led to the identification of the previously reported p.R506X pathogenic mutation in a heterozygous form, which is mainly known to cause DDOD syndrome  and storAlthough the majority of cases of DOORS syndrome are caused by mutations in the TBC1D24 gene . HoweverRegarding the establishment of clinicopathological relationships, the pyramidal signs observed were probably related to the generalized atrophy observed on MRI, which evolved with age as a part of the syndrome, or to the presence of small lacunes, as demonstrated by macroscopic neuropathological examination; however, their prominent role was not confirmed by either MRI or microscopic neuropathological assessment. No territorial ischemic stroke or any other reason was found to underlie the slightly left-sided corticospinal symptomatology.Drug-induced movement disorder , the functional consequence of the reduced vascular reserve capacity of the basal ganglia, and a late-onset symptom of DOORS syndrome may all account for the observed Parkinsonism. If considering Parkinsonism as a drug-induced movement disorder, the presence of resting tremor before the known administration of possible initiating agents represents a major contradiction. Nevertheless, the halting of risperidone therapy  led to a remarkable improvement, with almost complete cessation of the tremor. However, subsequently, the tremor recurred and the rigidity worsened, and these signs could only be moderately controlled by low-dose levodopa treatment (100 mg t.i.d.). In line with the SPECT findings, this may support a vascular etiology; however, no prominent underlying vascular lesions were identified either by MRI or by microscopic neuropathological assessment. Regarding the etiology of the neurodegenerative presentation, no α-synuclein or tau pathology could be demonstrated by detailed neuropathological assessment of the nigrostriatal system. Therefore, a combined secondary etiology  is suggested as an explanation for the Parkinsonian syndrome, and its relationship with DOORS syndrome cannot be proved. Of note, Parkinsonism was reported previously to be associated with mutations in the most common causative gene for DOORS syndrome . In addiATP6V1B2 gene , can initiate an NFT pathology that resembles that detected in AD and in the present case, albeit with a somewhat different regional distribution pattern [ the presence of age-associated tauopathies is demonstrated in DOORS syndrome in the forms of AGD, PART, and ARTAG, which may eventually be a consequence of lysosomal and/or mitochondrial dysfunction as discussed also in NPC, eventually in the frame of an accelerated aging process; (2) the cognitive and behavioral alterations that were present from childhood in the form of intellectual disability may be augmented by the evolution of the limbic-predominant tauopathy.In conclusion, the current study was the first to present a detailed clinicopathological relationship regarding p.R506X pathogenic mutation in the The limitation of this study was that only one case was reported; therefore, the findings need to be confirmed in additional cases. Accordingly, we propose future prospective studies using a series of patients and controls (including patients with AD and NPC) to elucidate the characteristics of endolysosomal dysfunction at different levels including a fine ultrastructural analysis and to obtain unbiased comparative quantitative measures of subregional tau pathologies.Written informed consent was obtained from the patient/participant's guardian/next of kin for the publication of this case report, including any identifiable data or images included within the study.DZ, LS, PK, and LV performed the neurological workup of the patients. ZR and GK performed the neuropathological assessment of the patient. NA and PC performed the genetic analysis. RM-T performed the determination of ApoE genotype. DZ wrote the first draft of the manuscript. DZ, LS, ZR, GK, and PC wrote sections of the manuscript. All authors contributed to manuscript revision and read 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.


In another cohort of patients with melanoma receiving mixed ICI treatment, the benefit score was also positively associated with higher overall survival (p = 0.022) and outperformed the TMB alone, with a significance of p = 0.089. The benefit score showed a positive correlation with clonal TMB, a reported immunotherapy marker, and exceeded it in immunotherapy response prediction. Besides, a high benefit score was found to be associated with higher proportions of natural killer cells, lower proportions of M2 macrophages and elevated CD8 T cells, all of which favor ICI therapy. In summary, tumor heterogeneity combined with the TMB showed superior efficacy in predicting the response to ICI therapy. This might further help to delineate the mechanisms of immunotherapy in patients with melanoma.The tumor mutational burden (TMB) has been reported as a predictive marker of the response to immune checkpoint inhibition (ICI) therapy in previous melanoma clinical trials. However, the TMB alone is not sufficient to accurately predict immunotherapy benefit. Additional biomarkers are needed for better stratification of immunotherapy-sensitive patients. In the present study, mutation data and survival information of patients with melanoma were collected from several immunotherapy studies, and tumor heterogeneity was estimated using mutant-allele tumor heterogeneity (MATH). The benefit score was defined as the ratio between the TMB and tumor heterogeneity, and optimal critical values were selected to group patients and evaluate their response to ICI treatment. The benefit score significantly improved the performance of stratifying the overall survival of patients compared with the TMB alone as a predictor in two independent cohorts (p = 0.0068  In recent years, immune checkpoint inhibition (ICI) therapy has benefited patients with advanced cancers, as exemplified for lung cancer  and melaAmong all identified markers, the TMB has proven its potential to predict the response of ICI in melanoma, based on the hypothesis that an elevated TMB corresponds to more neoantigens –12. HoweIn the present study, the TMB and tumor heterogeneity were estimated using somatic mutation data collected from several studies, in which patients with melanoma were treated with checkpoint inhibitors. Next, we defined the benefit score as a metric to integrate the TMB and tumor heterogeneity, and evaluated its predictive performance for the response to ICI therapy.After excluding studies without sufficient data, three melanoma cohorts that employed ICI therapy were brought into the analysis: The Van Allen cohort , the MiaWe defined the benefit score as the ratio between the TMB and tumor heterogeneity, where the TMB denoted the number of nonsynonymous somatic mutations and heterogeneity was determined using mutant-allele tumor heterogeneity (MATH), as calculated using the R package, maftools . For theThe clonal TMB, representing the number of clonal nonsynonymous somatic mutations, is a measure used to evaluate the clonal tumor mutation load of patients. In the Riaz cohort, cancer cell fraction (CCF) of mutations was estimated using PyClone , and muthttps://tcia.at/home)  were calculated. Fraction data of each tumor immune cell type in the Riaz cohort were downloaded from The Cancer Imaging Archive (TCIA) (at/home) . Cell frA total of 321 patients with melanoma from three cohorts, the Van Allen cohort, the Riaz cohort, and the Miao cohort, were included in our investigation Figure 1In addition to the benefit score, the clonal TMB is also simultaneously related to the TMB and tumor heterogeneity, and was reported to be associated with the response of patients treated with checkpoint blockers. We compared the benefit score with the clonal TMB based on the clonal results collected from the included studies and the correlation result is shown in vs. p = 0.1, HR = 0.546; vs. p = 0.13, HR = 0.596; vs. p = 0.089, HR = 0.629; vs. p = 0.092, HR = 1.52; vs. p = 0.33, HR = 0.664; vs. p = 0.037, HR = 0.624; The cut-off value for each marker was determined based on the principle of comparing their best performance to stratify the patients  cells and a lower proportion of M2 macrophages compared with those in the low benefit score group  Co., Ltd.The remaining 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.The handling editor declared a past co-authorship with one of the authors YY.