Biological age measurements (BAs) assess aging-related physiological change and predict health risks among individuals of the same chronological age (CA). Multiple BAs have been proposed and are well studied individually but not jointly. We included 845 individuals and 3973 repeated measurements from a Swedish population-based cohort and examined longitudinal trajectories, correlations, and mortality associations of nine BAs across 20 years follow-up. We found the longitudinal growth of functional BAs accelerated around age 70; average levels of BA curves differed by sex across the age span (50–90 years). All BAs were correlated to varying degrees; correlations were mostly explained by CA. Individually, all BAs except for telomere length were associated with mortality risk independently of CA. The largest effects were seen for methylation age estimators (GrimAge) and the frailty index (FI). In joint models, two methylation age estimators (Horvath and GrimAge) and FI remained predictive, suggesting they are complementary in predicting mortality. Everyone ages, but how aging affects health varies from person to person. This means that how old someone seems or feels does not always match the number of years they have been alive; in other words, someone’s “biological age” can often differ from their “chronological age”.Scientists are now looking at the physiological changes related to aging to better predict who is at the greatest risk of age-related health problems. Several measurements of biological age have been put forward to capture information about various age-related changes. For example, some measurements look at changes to DNA molecules, while others measure signs of frailty, or deterioration in cognitive or physical abilities. However, to date, most studies into measures of biological age have looked at them individually and less is known about how these physiological changes interact, which is likely to be important.Now, Li et al. have looked at data on nine different measures of biological age in a group of 845 Swedish adults, aged between 50 and 90, that was collected several times over a follow-up period of about 20 years. The dataset also gave details of the individuals’ birth year, sex, height, weight, smoking status, and education. The year of death was also collected from national registers for all individual in the group who had since died.Li et al. found that all nine biological age measures could be used to explain the risk of individuals in the group dying during the follow-up period. In other words, when comparing individuals with the same chronological age in the group under study, the person with a higher biological age measure was more likely to die earlier. The analysis also revealed that biological aging appears to accelerate as individuals approach 70 years old, and that there are noticeable differences in the aging process between men and women.Lastly, when combining all nine biological age measures, some of them worked better than others. Measurements of methylation groups added to DNA (known as DNA methylation age) and frailty (the frailty index) led to improved predictions for an individual’s risk of death. Ultimately, if future studies confirm these results for measures from single individuals, DNA methylation and the frailty index may be used to help identify people who may benefit the most from interventions to prevent age-related health conditions. Biological aging is a process characterized by progressive deteriorations occurring simultaneously at multiple levels, that is molecular, cellular, tissue- and organ-specific, in an organism . Due to Previous studies found low to moderate correlations among different types of BAs, for example telomeres, epigenetic clocks, clinical biomarkers, and frailty . HoweverTherefore, the present study aims to add to this knowledge gap in the field by examining longitudinal trajectories, correlations and associations to all-cause mortality using nine BAs in the Swedish Adoption/Twin Study of Aging (SATSA) . Here, tIn the present study, 845 individuals in SATSA , with atSATSA was initially designed to study individual differences in aging within twin pairs reared apart. The present study, however, did not focus on twin-oriented aspects, such as quantifying the magnitude of familial effect in exploring the BA-mortality associations. Instead, we made statistical inferences for the general population, in which relatedness within twin pairs was accounted for by introducing random effects in the mixed models and estimating robust errors in Cox regressions.telomere length was measured by quantitative polymerase chain reaction (qPCR) and presented as a relative telomere length  were then calculated using established algorithms based on the methylation levels of 353, 71, 513, and 1030 CpG sites, respectively  measured by blood tests and physical examinations  ability, spatial (fluid) ability, memory, and perceptual speed using a cognitive testing battery. The four specific domains were combined into an overall score by principal component analysis , pulmonary, strength (grip strength), and movement/balance (gait speed). The four indicators were standardized separately and summed to create a composite score , 60.4 years for Horvath DNAmAge, 65.2 years for Hannum DNAmAge, 63.8 years for DNAmPhenoAge, 69.4 years for DNAmGrimAge, 64.7 years for physiological age, 51.5 for cognitive function, 48.3 for FAI, and 0.10 for FI. The baseline characteristics in individuals with complete measurements were similar to those in all individuals.  was 25.7 kg/mviduals. .On average, BAs were assessed between 2.5 times (telomere length) to 4.2 times (FI) in each participant with information on the respective BAs.  LongitudTo examine the possible effect modification, we next introduced an interaction term between CA and sex in the mixed models. We found no evidence that the shape of the BA curves differed between men and women, except for in physiological age (p<0.001 for sex interaction). However, the difference was mainly observed at the end of the CA spectrum . .BAs were broadly categorized into four groups according to the main biological structural levels where the BA measurements were implemented, that is telomere length, DNAmAges, physiological age, and functional BAs  comparable, we rescaled all BAs and BA residuals by their standard deviation (SD), so that HRs express changes in mortality risk due to a one-SD increment of the respective BA or BA residual. Fully adjusted models controlled for sex, educational attainment, smoking status, and BMI. All models were stratified by participants’ birth year  and implicitly adjusted for CA through the choice of attained age as the underlying time scale. When repeated measurements were available, only the first BA assessment was included in the survival analysis and referred to as the baseline BA.First, we looked at the effect of baseline BAs separately . During a median follow-up time of more than 15 years, we observed statistically significant BA-mortality associations for DNAmAges  and functional BAs , where a one-SD increase of BAcorresponded to a change in mortality risk by 39, 26, 17, 32, 27, and 15%, respectively. Increased mortality risks were also found with changes in Hannum DNAmAges and physiological age, although the effects were generally weaker and came with a slightly higher level of imprecision, with HRs (95% CI) being 1.17  and 1.13 . We found no evidence for telomere length as associated with mortality risk.  The corrSecond, we explored the baseline BAs in relation to mortality risk by taking all BAs into consideration simultaneously in 288 individuals with complete measurements . To avoid biased estimation due to collinearity , BA residuals instead of BAs were used. During a median follow-up time of 16.5 years, we observed that three BA residuals, Horvath DNAmAge, DNAmGrimAge, and FI, out of nine were associated with the risk of all-cause mortality independently of all other BAs. A one-SD increment of Horvath DNAmAge, DNAmGrimAge and FI residuals was associated with a 31%, 43% and, 58% higher risk of death, respectively, independent of the level of CA, all other BA residuals and common risk factors. The HRs for the other DNAmAges (Hannum and PhenoAge), physiological age, and the other functional ages (cognitive function and FAI) were attenuated towards one after controlling for all BAs in the same model .We replicated the first part of the survival analyses  in subgroups classified by sex, age group, and smoking status, as well as in the sub-cohort of individuals with complete measurements. In the subgroup analyses, we observed evidence for potential effect modification. The associations of BAs with mortality risk were generally stronger in women , more pronounced in the younger individuals , and a bit stronger in current or ex-smokers (for Horvath DNAmAge and DNAmGrimAge) compared to those in men, older individuals, and non-smokers, respectively.  FurthermThe present study investigated nine different BAs in a middle and old-aged Swedish population-based cohort. We observed longitudinal growth of functional BAs accelerated around age 70 and sex differences in the average levels of BAs across the age spectrum. All BAs were correlated to varying degrees and the correlation coefficients were attenuated after adjusting for CA. Of nine BAs, four DNAmAges, physiological age, and three functional BAs were associated with the risk of mortality individually, with the strongest effect size observed for DNAmGrimAge and FI. In particular, Horvath DNAmAge, DNAmGrimAge, and FI remained predictive of mortality in a multivariable model including all BAs.As biological aging does not proceed at a constant pace across time, monitoring longitudinal trajectories of BAs using repeated assessments to depict the characteristics of the aging course is of great importance. We observed an accelerated change in the aging pace in functional BAs, which are in parallel with previous studies focused on trajectories of cognition, physical function, and frailty . This reIn contrast to functional BAs, DNAmAges and physiological age accrued at a relatively constant rate. Due to their mode of construction, DNAmAges and physiological age were transformed into an age-calibrated scale in the unit of year. Thus, these BAs allow individuals to perceive an intuitive impression on their biological aging status by comparing BA with CA and have the potential to serve as a straightforward-to-use metric in the geriatric practice. However, in order to be clinically meaningful, they need to be validated on the individual level data as well. In addition, a drawback of this mode of construction is that it largely explains the linear shapes of the BA trajectories, making longitudinal BA characteristics less informative as the changes are constant. Another feature of physiological age is the strong correlation with CA; hence, it allows limited biology perception in prediction models on top of CA. This is again due to its mode of construction using a composite score of several biomarkers where each one, by definition, strongly correlates with CA. To improve construction models for BA, composite scores should balance the reliance on biological aspects and weight on CA.Sex differences in aging are observed at multiple biological levels . Most noAging is a complex process proceeding through interconnected biological mechanisms. BAs, in essence, are underlain by different sets of these mechanisms. Correlations of BAs could provide an idea of how one BA changes in accordance with another. We observed moderate to strong correlations between CA and all BAs, with the exception of telomere length, which was only weakly correlated with CA and the other BAs. These results are in line with previous research observing similar correlation pattern for 11 BAs, as well as low correlations between telomere length and other types of BA . As someAfter regressing out the effect of CA, we still observed weak to moderate correlations between DNAmAges. These DNAmAges were initially trained to capture methylome differences associated with various aging-related phenotypes, namely CA for Horvath and Hannum, Phenotypic Age for PhenoAge, and mortality risk for GrimAge . EmployiFunctional BA indicators included cognitive function, FAI, and FI, which quantify cognitive, physical, and frailty-related functioning. Intriguingly, the present analysis found comparable levels of correlation for these BAs as well as for their residuals, suggesting that most of these correlations were not simply driven by CA. Previous studies have also reported correlations between physical function, cognitive function, and health-related quality of life . As funcA majority of the included BAs are well-established aging indicators and were found to be associated with mortality in previous studies . Our resFew studies have focused on the comparison of the predictive aspect of BAs. Physiological age is a composite biomarker of aging, developed from 10 blood and clinical markers. Levine et al. applied a similar method to develop a BA indicator and found it associated with mortality beyond CA in a large US population . We alsoDNAmGrimAge is derived via a two-step development method, in which methylation data were first used to predict a set of biomarkers (plasma proteins and smoking pack-year), and the methylation-based biomarkers were then used in the prediction of mortality risk. As a result, the DNAmGrimAge is explicitly trained to be a mortality predictor; it also includes the largest number of genomic sites compared with the other three DNAmAges, and allows the methylation level of a single CpG site to contribute to mortality risk via different intermediate biomarkers, as the methylation-based biomarkers used somewhat overlapping CpG sites in the two-step training phase. The FI is developed using clinical information and is likely to capture an organismal perspective on aging that is closer to the clinical end-point of death compared to molecular-based BAs. In addition, the FI incorporates information in multiple health domains, in contrast to the other two functional BAs, cognitive function, and FAI, which exclusively measure cognitive or physical performance. The comprehensive nature of the FI could partly explain the strong and robust mortality association observed here and in previous studies . FunctioIn the subgroup analysis, we found moderate effect modification from age and smoking status. A previous study in a large Swedish cohort from our research group also observed that the effect size of FI with mortality was attenuated with increasing age . DNAmGriIt is noteworthy that in comparing the strength of relative risk of mortality related to each BA, we were not aiming to propose any superior BA, as mortality association alone is by no means the gold standard for a qualified BA in aging research. Evidence exists that some BAs have advantages in predicting other aging-related outcomes such as entry into care  , which mThe present study has several strengths. First, we measured nine BAs in the same population from the molecular to the organismal level, which provided us an opportunity to examine BA characteristics from several molecular and functional aspects. Second, the BAs were assessed in a longitudinal manner to allow for descriptive and correlational analyses considering changes over time. Third, through the linkage to the Swedish national registers, we were able to keep track of study participants’ vital status for nearly 20 years, thus being able to predict the risk of death in relation to BA status prior to the occurrence of most aging-related outcomes.Several limitations should be taken into consideration when interpreting the present findings. First, only individuals with complete measurements were included in the nine-BA survival models. Complete observations may not be a representative subgroup of the entire study population and lead to selection bias. However, IPT occasions with incomplete BAs were mostly due to administrative reasons, such as methylation data were only assessed in five waves since IPT3, and these factors are likely not related to subject-specific features. In addition, we replicated the one-BA survival models in individuals with complete measurements, and the results did not show major discrepancies. Second, the birth year of SATSA participants spanned a long period of time, from the year 1900 to 1948, thus raising concerns for cohort effects. During this time, improved medical conditions or emergent public health events could affect both BA and mortality risk and cannot be controlled merely through the age scale in the present analysis. All survival models were stratified by participants’ birth year in 10 year intervals to in part alleviate bias due to the cohort effect.In summary, biological age is a construct built on molecular, cellular, and functional aspects of an individual´s health status and has the potential to change the way risk prediction is performed in the clinic. In the era of personalized medicine, individual health assessments are warranted and should rely on better methods than simply using the CA. In this study, we present the most comprehensive analysis of biological age in humans to date. We explain their correlations and longitudinal patterns. We highlight the sex-specific effects and point to the importance of providing sex-specific estimates in aging investigations and acknowledging sex-specific care in geriatric practices. Furthermore, we found BAs had the potential to provide mortality-relevant information independently of CA and independently of other types of BAs. Further studies are needed to investigate if the present findings could be extrapolated to a younger population, what the specific mechanisms underlying the BA correlations are, and possible modification effects on and between BAs.The Swedish Adoption/Twin Study of Aging (SATSA) was a population-based study consisting of reared apart and reared together twin pairs . In-persTelomere length was measured from DNA extracted from leukocytes in peripheral blood . A quantGenome-wide methylation levels were measured from leukocytes using Illumina’s Infinium HumanMethylation 450K BeadChip according to the manufacturer’s protocol and quantified by beta-values . DNAmAgePhysiological age considered a set of physiological biomarkers assessed from the immediate blood test, blood test in lab, urine strip test, and physical examination data that were available in all waves of IPT. First, we included one measurement for each individual to form a sub-sample in which one measurement was randomly selected when repeated measurements for a single individual were available. Pearson correlations were examined using measurements of age and candidate biomarkers in the sub-sample. As a result, nine and five eligible age-associated biomarkers (Pearson correlation >0.1) were included in the development of physiological age for men and women separately. Four cognitive domains were assessed through a battery of in-person cognition testing, including crystallized, fluid, memory, and perceptual speed abilities . Scores Four types of specific functional measurements were taken into consideration in the development of FAI . Vision FI was introduced to conceptualize the vulnerability of a given person to a range of age-related adverse outcomes. FI in SATSA was constructed from 42 self-reported health deficits, such as symptoms, diseases, disability, mood, and activities in daily living. FI was calculated as the ratio of the number of deficits presented in a given person to the total number of deficits considered in the study (n = 42 in SATSA). Details of FI items are described in We constructed BA residuals by regressing out the CA-related part from respective BA. As BAs were assessed in a longitudinal manner and BA levels within twin pairs were assumed to be related due to shared familial factors, we adopted mixed models with fixed effects for sex and CA, the latter as a natural spline term with three degrees of freedom, to allow for non-linear relationships, and random intercepts at the twin-pair and subject level:2. Smoking status was categorized as non-smokers, ex-smokers, and current smokers. Among the included individuals and measurements, 28 out of 846 individuals without education information were assigned into an unknown group, and another one and two individuals who missed the BMI and smoking status were imputed with the corresponding information collected from their nearest available IPTs.Sex, educational attainment, and baseline information of BMI, and smoking status were considered as covariates in the survival analyses. BMI was assessed through physical examination and other covariates were acquired through self-reported questionnaire data. Educational attainment was classified as primary education, lower secondary or vocational, upper secondary education, and tertiary education. BMI was derived as the body mass divided by the square of body height, expressed in units of kg/mSince the participants in the present analysis were born over a long period of time (1900 to 1948), we estimated the mortality associations with stratification on the calendar year of the participants’ birthday to avoid confounding from cohort effects. Individuals’ birth year was treated as a categorical variable with five groups in an interval of 10 years.All-cause mortality data, including vital status and dates of deaths, were obtained from linkages between the STR and Swedish Population Register through the personal identification number assigned to all residents. All-cause mortality data were updated on August 16, 2018.Characteristics of baseline measurements were presented as means  and proportions. Longitudinal changes of BAs were presented in plots with individual level measurements illustrated by dots, lines, and broken lines. Average trajectories of BAs were estimated through mixed linear regression models as described above, each of which included fixed effects of the intercept, CA, and sex, and a random effect of intercept at the individual and twin-pair level. P-values for sex effects were obtained from fixed effects within the mixed linear model. In addition, an interaction term between each BA and CA in natural spline was introduced to the model, and P-values for sex interaction were determined by the likelihood ratio test comparing the models with and without the interaction term.We estimated the repeated measures correlation coefficients between age and nine BAs using all complete measurements . The corWe used the Cox regression model to estimate the association between baseline BAs and the risk of all-cause mortality. When repeated measurements were available, only the first BA assessment was included and referred to as the baseline BA. In all models, we used attained age as the underlying time scale and the five groups of birth year as strata. Left truncation and right censoring were accounted for in the estimations; that is individuals entering into the cohort is conditional on their survival at baseline ages, and follow-up time may stop before death occurs. Follow-up time started from the first available measurement (baseline measurement) and ended at the date when they died or were censored on August 16, 2018. In addition, robust standard errors were introduced to adjust for relatedness within twin pairs and subjects. To achieve a direct comparison of BA effects in relation to mortality risk, we standardized all nine BAs to the mean of zero and SD of one across all available measurements and replicated the transformation in the BA residuals, such that the estimated hazard ratios (HRs) could be interpreted as the relative risk of death associated with a one-SD increase in the level of BA or BA residual. In the first part of the survival analyses, all models accounted for only one BA (one-BA models). For those who had their BAs assessed for multiple times, the first available measurement was treated as the baseline measurement. For each BA, we estimated the association of BAs and BA residuals with mortality risk using two models, a univariate model and a multivariate model with sex, education, BMI, and smoking status as additional covariates. In the second part of the survival analyses, all nine BAs were accounted for altogether in the same survival model (multi-BA models). Similarly, two models with or without adjustment for common risk factors were estimated. Only BA residuals were taken into consideration in the second part of survival analyses to avoid biased results caused by collinearity within BAs.P values were two-sided, and statistical significance was defined as p<0.05. All analyses were conducted using Stata 15.1 and R 3.6.0. In the interests of transparency, eLife publishes the most substantive revision requests and the accompanying author responses.Acceptance summary:This is an important and timely study for the aging research field and more broadly for biomedical researchers. The study addresses the important question about validating promising human biomarkers of aging within a robust investigational design.Decision letter after peer review:eLife. Your article has been reviewed by three peer reviewers, and the evaluation has been overseen by Diane Harper, as Reviewing Editor, and Eduardo Franco, as Senior Editor.Thank you for submitting your article entitled "Longitudinal trajectories, correlations and mortality associations of nine biological ages across 20-years follow-up" for further consideration by There are some issues that need to be addressed before acceptance, as outlined below:Reviewer #1:I think this is an important and timely study for the aging research field and more broadly biomedical researchers. The study addresses the important question about validating promising human biomarkers of aging. The study stands out in terms of its scope (profiling many different biomarkers ranging from DNA methylation to frailty index), its large sample size: longitudinal assessments from n=845 individuals, long follow up times, and its rigorous statistical approach (linear mixed modeling for adjusting for dependencies). It carefully evaluates the latest molecular biomarkers (e.g. recent methylation based biomarkers) relates them to a hard outcome: time to death. The sensitivity analysis illustrates robustness. The discussion seems to be fair and is grounded on the results. The statistical analysis appears to be sound.Reviewer #2:The advantages of this work are longitudinal (20-year) studies of several approaches to biological age estimation (nine measurements) at once, including data on mortality, in the same population.Not sufficiently discussed is why a combination of these 9 marker groups was chosen? What are their mean absolute errors? How many persons have different longitudinal time points with all the nine? If such patients exist, is it possible to give individual correlations of different BAs with age?Is it possible to conclude that longer telomeres or shorter DNAmAge are practically not associated with physiological indicators of functioning ? Could the authors emphasise this aspect of the results in the Discussion?Reviewer #3:In this paper the authors assessed longitudinal measurements of multiple biological age in a population-based cohort from Sweden with long term follow-up. The dynamic changes of 4 types of 9 BAs over 20 years, mutual correlations and individual/joint associations with mortality of the BAs were explored. The study stands out in terms of longitudinal design, repeated measures of multiple types of BAs, and adequate statistical analysis. The results further confirmed the validity of multiple BAs as aging indicators and their potential complementary roles in aging process, and highlighted the variations with respect to gender. The following comments are mainly to improve the clarity.1) Longitudinal changes of BAs over CA were investigated, whereas longitudinal changes of BAs in relation to mortality were not explored.2) Follow-up time for cognitive function and physiological age (and even FAI and FI) were obviously longer than follow-up time of other BAs, it therefore may not be appropriate to compare the strength of different BAs with mortality given that only the first BA assessment was included in the survival analysis. What are the results if truncating follow-up time for different BAs comparable?3) What are the health status of participants at baseline (first BA assessment), which may affect mortality risk and is suggested to be controlled in regression models.4) Categories of BAs are suggested to be described explicitly in the Introduction. Reviewer #2:The advantages of this work are longitudinal (20-year) studies of several approaches to biological age estimation (nine measurements) at once, including data on mortality, in the same population.We thank the reviewer for this positive feedback and the following helpful comments to improve our work.Not sufficiently discussed is why a combination of these 9 marker groups was chosen? What are their mean absolute errors?a) The main objective of the present study is to investigate different types of biological ages (BAs) jointly. Therefore, we intended to include as many BAs as possible so that a range of BA types could be compared. As such, the nine BAs are selected mainly based on data availability and we have added the following sentence to the manuscript:“Nine BAs were included in the present study based on data availability in SATSA.”i and BA2i denote the levels of a pair of BAs in individual i, and n is the total number of individuals.b) In The pairwise MAEs are presented graphically in As MAE quantifies the absolute difference between two continuous variables, it would be an efficient indicator if all the BAs were assumed to measure the same underlying process, i.e., if the expected relationship between each pair of BAs is the identity line y=x. However, both prior biological knowledge and our results suggest that telomere length, DNA methylation age estimators, physiological age, and functional BAs are likely to measure different aspects of the aging process. Thus, assuming all the BAs are growing in the same manner would be a rather unrealistic assumption and makes MAE a less informative measure to indicate the difference between the two BA measures of the different typesConsidering that we have already reported correlation coefficients to provide the information of how different BAs agree with each other, we will not include the MAE results in the manuscript and only present the figure in the response letter for the reviewer’s reference.How many persons have different longitudinal time points with all the nine? If such patients exist, is it possible to give individual correlations of different BAs with age?a) In total, 288 individuals had at least one complete measurement occasion, of which 105, 87, 60, 27, and 9 individuals had 1, 2, 3, 4, and 5 times of complete measurements.Author response table 1: Number of individuals by number of complete measurementsb) For the correlation analysis, we included all the 612 complete measurements and estimated the repeated measures correlation (rmcorr) coefficients, which accounts for non-independence among observations of the same individual.Therefore, the reported correlation coefficients have already taken repeated measurements into consideration and are essentially measures of overall intra-individual correlation. We have expanded the methods description of the repeated measurement correlation in the manuscript to clarify this point:“Using all complete measurements, we estimated repeated measures correlation  between age and all nine BAs, which accounts for non-independence among observations and captures the common intra-individual association between age and each of the BAs in turn.”Is it possible to conclude that longer telomeres or shorter DNAmAge are practically not associated with physiological indicators of functioning ? Could the authors emphasise this aspect of the results in the Discussion?It is indeed true that our results suggest molecular BAs are practically not associated with indicators of functioning  when we controlled for the age effect . It can be interpreted that among the individuals with the same CA, molecular BAs and functional BAs are only weakly associated, and largely reflect different aspects of the aging process.We have added the following sentences to emphasize this point in the Discussion section. Further interpretation regarding the characteristics of functional BAs in contrast to molecular BAs is detailed in the Discussion section .“In other words, among individuals with the same CA, molecular BAs and functional BAs are only weakly associated. We interpret this as an indication that these categories largely reflect different aspects of the aging process.”Reviewer #3:In this paper the authors assessed longitudinal measurements of multiple biological age in a population-based cohort from Sweden with long term follow-up. The dynamic changes of 4 types of 9 BAs over 20 years, mutual correlations and individual/joint associations with mortality of the BAs were explored. The study stands out in terms of longitudinal design, repeated measures of multiple types of BAs, and adequate statistical analysis. The results further confirmed the validity of multiple BAs as aging indicators and their potential complementary roles in aging process, and highlighted the variations with respect to gender. The following comments are mainly to improve the clarity.We thank the reviewer for this positive feedback and the following helpful comments to improve our work.1) Longitudinal changes of BAs over CA were investigated, whereas longitudinal changes of BAs in relation to mortality were not explored.We agree with the reviewer that investigating the association between BA changes and mortality risk is an interesting and important topic. However, the current data is underpowered to test such effects as the number of individual with at least two measurements are quite limited, especially when it comes to telomere length and DNAmAge estimators (Table 2). Investigating the longitudinal change of functional BAs  in relation to health-related outcomes could be less affected by the power issue and there are, in fact, some on-going works from our research group devoted to this topic. Therefore, we have decided to not touch upon the topic in the present analysis.Author response table 2: Number of individuals by number of BA measurements2) Follow-up time for cognitive function and physiological age (and even FAI and FI) were obviously longer than follow-up time of other BAs, it therefore may not be appropriate to compare the strength of different BAs with mortality given that only the first BA assessment was included in the survival analysis. What are the results if truncating follow-up time for different BAs comparable?We agree with the reviewer that different follow-up times might interfere with the comparability of multiple BAs in relation to mortality risk. In one of the sensitivity analyses, we replicated the individual BA-mortality association among the participants with the complete measurements, in which all baseline BA measures were investigated with the mortality risk during the same follow-up time, a median of 16.5 years. The results were present in Supplementary Table 10 in Supplementary file 1 and didn’t show major discrepancies compared to those in Table 4.3) What are the health status of participants at baseline (first BA assessment), which may affect mortality risk and is suggested to be controlled in regression models.We thank the reviewer for suggesting us to consider potential confounding effect due to baseline health status. We analyzed the self-reported data on prevalent diseases and found 4.0%, 1.7%, 5.5%, and 3.9% of the participants have reported previous diagnoses of heart failure, stroke, diabetes, and cancer, respectively, at their first available measurement occasion.As a consequence, we have included Supplementary Table 11 in Supplementary file 1, and added the following sentence to the Results section.“We also adjusted for the presence of the previous diseases, including heart failure, stroke, diabetes, and cancer in the survival model and the observed results were largely unchanged.”4) Categories of BAs are suggested to be described explicitly in the Introduction.We thank the reviewer for pointing this out and have now explicitly described the BA categories at the end of the first Introduction paragraph.“One of the potential BAs, telomere length, is the length of a nucleotide sequence at the chromosomal ends, and represents the capacity for cell division . Epigenetic clocks comprise aging-related DNA methylation information from various genomic loci  . Another composite score derived from clinical measurements and blood biomarkers can be viewed as a physiological age measure . Furthermore, organismal functioning status could reflect the biological aging process from various aspects, such as cognitive performance, physical functioning, and overall frailty .”


Ananas comosus waste (ACP) for the adsorption of CEB. The SEM, EDX, FTIR, XRD, BET, and TGA characterizations showed the successful presence of ZnONPs on the biomass to form a nanocomposite. The uptake of CEB was enhanced by the incorporation of ZnONPs on ACP. A faster CEB adsorption onto ZnO@ACP (120 min) compared to ACP (160 min) was observed. The Langmuir (R2 > 0.9898) and pseudo-second-order (R2 > 0.9518) models were most appropriate in the description of the adsorption process. The impregnation of ZnONPs onto the biomass enhanced the spontaneity of the process and displayed endothermic characteristics. High CEB desorption of 81.3% from the dye loaded ZnO@ACP as well as efficient reusability showed the efficacy of the prepared nanocomposite for CEB adsorption.Despite the efficiency of ZnO nanoparticle (NPs) composite adsorbents in the adsorption of various pollutants, there is presently no report on the combo of ZnONPs with biomass for adsorption. Besides, there is a dearth of information on the biosorption of celestine blue (CEB), a dye used in the nuclear and textile industry. In this study, biogenic-chemically mediated synthesis of a composite (ZnO@ACP) was prepared by the impregnation of ZnONPs onto  Most dyes are used in the textile and paper industries and are frequently encountered in the effluents subsequently released into the environment3. These dyes present in the environmental waters can be carcinogenic, mutagenic, and result in chronic illnesses in humans and aquatic organisms4. Apart from that, they reflect or absorb light entering the water thus hindering the photosynthetic activity of aquatic plants1. Therefore, the removal of dyes from wastewater is necessary to abate their harmful effects on the ecosystem6. Most dye removal studies have focused on the removal of dyes such as methylene blue, congo red, malachite green, rhodamine B, methyl orange, methyl violet, reactive black, basic blue, acid yellow, brilliant green, and crystal violet8. However, research on the removal of celestine blue (CEB) is rare despite the wide use of CEB in the nuclear and textile industries. Therefore, the removal of CEB from wastewater is important. Several techniques have been harnessed for the remediation of dye-polluted water, such as filtration, coagulation, oxidation, precipitation, reduction, photocatalytic degradation, solvent extraction, and adsorption9. The adsorption technique is the most efficient and adsorption on bio-waste (biosorption) is promising due to the simplicity, low cost, degradability, reusability, and efficiency10. Hence, the use of several bio-waste for the biosorption of dyes has received much attention, but pineapple (Ananas comosus) peel waste (ACP) is seldom reported, despite its abundance and viable potentials11.The pollution of environmental water from industrial effluents contaminated with industrial by-products is on the rise. This is attributed to the rapid growth of industries, hence the increase in industrial activities across the globe. About 700,000 tons of over 100,000 commercial dyes are produced annually in industries, which makes dye the most common water pollutant13. Much attention has been given to the use of magnetic NPs and the composites for water treatment when compared to other NPs, attributed to the easy of recovery of magnetic NPs from solution by magnetic attraction15. However, the problem encountered in the recovery of other NPs from the treated water due to their small sizes has limited their application. Therefore, the impregnation of these NPs on a substrate (support) to form composites adsorbents to aid the recovery has become popular. Also, these composites adsorbents have better efficiency for water treatments due to the presence of various kinds of active sites for pollutant uptake, especially when composed of inorganic and organic sites16. Furthermore, among nanocomposite adsorbents, zinc oxide (ZnO) NPs composites have been reported to be highly efficient for the treatment of dye polluted water23. However, these hybrids are mainly a combo of ZnONPs with materials like activated carbon, graphene oxide, chitosan, graphene, polyaniline, and clay minerals28. Despite the desirable characteristics of biosorbents mentioned, there is presently no report on the combination of ZnONPs with biomass for water treatment. This study, therefore, reports for the first time the synthesis of a ZnONP-biomass composite (ZnO@ACP) and its application for the adsorption of CEB. This research is also significant as it provides insights into the biosorption behavior of CEB on materials, which is presently not established. The thermodynamics, kinetics, and isotherm of the biosorption process were evaluated. In addition, the mechanism of CEB uptake, as well as the regeneration and reuse of the materials, was investigated.Meanwhile, recent researches have focused on the use of nanoparticles (NPs) for water treatment owing to their adsorption efficiency and potent catalytic activities3), sodium hydroxide (NaOH), zinc acetate hydrate (Zn(CH3COO)2), and celestine blue (C17H18ClN3O4) were all obtained from Sigma-Aldrich. These chemicals were all analytical grade and were utilized in the experiments without any purification. The structural representation of celestine blue dye is shown in Scheme Ananas comosus) was obtained from Checkers mall, Mimosa, Nelson Mandela Drive, Bloemfontein, South Africa.The hydrochloric acid (HCl), nitric acid  for 24 h. The dried peels were pulverized into a fine powder with a laboratory pestle and mortar. To ensure the removal of water-soluble components, which could contaminate the water during the adsorption process, the sample was pretreated. Dilute acid treatment was carried out by contacting the pulverized mass with 0.1 M HNO3 in a beaker with constant stirring for 1 h. Thereafter, diluted with excess distilled water and decanted, the dilution was repeated until the pH was 7.0. The residue was dried at 70 °C for 24 h in an oven, pulverized, and sieved through the 100-µm mesh. The prepared Ananas comosus peel waste (ACP) was kept in a desiccator until use.Manual removal of the peels of 3COO)2 to 100 mL of distilled water in a beaker with vigorous stirring using a magnetic stirrer at 30 °C for 30 min. Thereafter, 10.0 g of pulverized Ananas comosus (without acid treatment) was added with continuous stirring for 15 min, after which 0.2 M NaOH solution was added dropwise until pH 11.0. The solution was stirred for 2 h, allowed to settle for 30 min, before centrifuging at 8000 rpm for 1 h. The obtained combo was washed repeatedly with distilled water until neutral pH and then centrifuged. The ZnONP biomass hybrid was dried at 80 °C for 24 h, pulverized using the mortar and pestle, sieved through a 100-µm screen, and designated as ZnO@ACP.One-pot synthesis by incipient-wet impregnation technique was applied in the preparation of the ZnO-biomass composite. The reducing action of the biogenic waste biomass in combination with chemical reduction was employed. This was carried out by adding 4.0 g of Zn(CH2). The pore properties and surface area (SBET) were examined by the surface area analyzer (Micromeritics ASAP 2020 model) and results refined by MicroActive VI.01 software. The thermo-gravimetric analyzer  with a 10 °C min−1 heating rate and 200 mL min−1 nitrogen flow rate analyzed the thermal stability. The pH drift method was used to determine the pH point of zero charged (pHpzc)11. The surface functionality was examined by the Fourier transform infrared spectrometer . X-ray diffraction (XRD) was determined using an X-ray diffractometer (Brucker Model) with Cu radiation of 1.54 Å. Diffraction patterns were in the 2θ range of 10–80°, with a step size of 0.1° and 2 s counting time per step.The ACP and ZnO@ACP were characterized to determine the surface properties. The morphology and elemental composition were analyzed by the field emission scanning electron microscopy  coupled with energy-dispersive X-ray spectroscopy , pH (2.0–9.0), temperature (300–323 K), and CEB concentration (10–50 mg L−1). The solution pH was adjusted with 0.1 M solution of HCl or NaOH. In a typical experiment, 0.05 g of the material was added to 10 mL of a given CEB solution, agitated for 5 min and left for a contact time of 180 min. Thereafter, the solution was centrifuged at 8000 rpm for 30 min, analyzed for residual CEB concentration at the maximum wavelength of 644 nm  and percentage removal were calculated from the mass balance and percentage equations, respectively29.v (L) is the volume of solution, m (g) is the adsorbent weight, Co and Ce in mg L−1 are the initial and residual CEB concentrations, respectively.Exactly 0.1 g of CEB was dissolved in 1 L volumetric flask to obtain a stock solution of concentration 100 mg L−1 CEB solution at pH 7.0, a contact time of 180 min and temperature of 300 K. Thereafter, the solution was centrifuged and the amount of CEB remaining was determined by the UV spectrophotometer. The CEB-loaded material was oven-dried at 70 °C for 30 min, after which it was mixed with 10 mL of 0.2 M HCl eluent, agitated for 5 min, and left for a contact time of 25 min at 300 K. The concentration of CEB in the desorbed acidic solution was analyzed after centrifugation. The desorption of CEB was evaluated from equation30:D (L) is the volume of eluent used and CD (mg L−1) represents the concentration of CEB in the acidic desorbed solution. The parameter, qe (mg g−1) denotes the adsorption capacity of the material for CEB, m (g) represents the adsorbent’s mass. The regenerated adsorbents after oven drying at 80 °C for 30 min, was reused for CEB adsorption. Three cycles of adsorption–desorption experiments were carried out.The regeneration and reusability of the materials were evaluated by contacting 0.25 g of the adsorbent with 10 mL of 50 mg L2), calculated from the statistical function of Origin 2019b software. The closer the R2 value to one the best the model fit. The experimental responses were evaluated using descriptive statistics. Each experimental run was carried out in triplicate and the mean values computed. The figures were plotted using the origin 2019b software and the error bars in the figures represent the standard deviations where applicable.The best fit kinetic or isotherm model describing adequately the adsorption data was obtained from the coefficient of determination  of the adsorbents , pore volume, and pore diameter of ACP was 39.816 m2 g−1, 0.0245 cm3 g−1, and 1.184 nm, respectively. As shown in Fig. BET of 10.564 m2 g−1, a pore-volume of 0.0068 m2 g−1, and pore diameter of 1.761 nm. Despite the decrease in SBET after the impregnation of ZnONPs, the increase in pore diameter could be beneficial for efficient dye uptake32. The scanning electron microscopy (SEM) of ACP  was slightly improved with ZnO@ACP (6.52 mg g−1). The Langmuir RL value of 0.171–0.508 for ACP and 0.110–0.383 for ZnO@ACP as well as the Freundlich n values indicated an efficient affinity for CEB44. The maximum adsorption capacity of ACP and ZnO@ACP for CEB was compared with other low costs, pineapple, and ZnONPs based adsorbents used for cationic dye adsorption as shown in Table The mechanism of CEB uptake onto ACP and ZnO@ACP was evaluated from the kinetics, isotherm, and thermodynamic. The models’ descriptions are presented in the supplementary material. The isotherm of CEB adsorption on the materials was evaluated by Freundlich and Langmuir models and the plots are shown in Fig. O@ACP 6.5 mg g−1. 2 values, the pseudo-second-order kinetics presented the best fit in the overall adsorption of CEB on both ACP and ZnO@ACP than the pseudo-first-order model. This suggests the involvement of electrostatic interactions between CEB molecules in solution and the surfaces of ACP and ZnO@ACP56. The involvement of the intraparticle-diffusion mechanism was found to be the dominant diffusion mechanism for CEB uptake. However, we observed the presence of intercept values, showing that the plot did not pass through the origin. The implication is that boundary layer diffusion was also involved in the overall removal process of CEB on the materials and that the uptake was not restricted solely to the intraparticle diffusion mechanism39. The thermodynamic analysis showed that the uptake of CEB on ACP is non-spontaneous based on positive ∆Go values. Therefore, the initial agitation and high CEB concentration created a driving force for efficient interaction between CEB molecules and ACP. However, after the impregnation of ZnONPs on ACP, the uptake became spontaneous as indicated by negative Gibbs free energy changes at all temperatures, which is desirable  to form a hybrid (ZnO@ACP) was found to enhance the adsorption of celestine blue (CEB) at variations of pH, temperature, time, adsorbent dosage, and dye concentration. The SEM, EDX, and XRD analysis showed successful impregnation of 35.7 nm-sized ZnONPs on the hybrid. The presence of ZnONPS increased the thermal stability as well as the pH point of zero charge of the biosorbent. The dye uptake on both adsorbents was well explained by the Langmuir isotherm and pseudo-second-order kinetic models. The intraparticle diffusion mechanism was found to be the major mechanism of dye removal, while thermodynamics revealed that the presence of ZnONPS enhanced the spontaneity for CEB adsorption. We found that ZnO@ACP exhibited a faster uptake of CEB when compared to the pristine ACP. The hybrid also exhibited higher dye desorption from its loaded surface as well as higher adsorption over three cycles of regeneration and reuse. The adsorption behavior of CEB was similar to other cationic dyes reported in the literature. These results showed the viability of ZnO@ACP for CEB decontamination from wastewater. In addition, we recommend that more studies should be conducted on the adsorption of CEB on various materials to provide sufficient insights into the adsorption behavior.Supplementary file1.


There are several types of tenodesis reconstruction designed for subtalar instability. However, no comprehensive comparison has been conducted among these procedures in terms of their correcting power so far. The objective of this study is to evaluate the biomechanical behaviors of 5 representative procedures through finite element analysis.Finite element models were established and validated based on one of our previous studies. The Pisani interosseous talocalcaneal ligament (ITCL) reconstruction, Schon cervical ligament (CL) reconstruction and Choisne calcaneofibular ligament (CFL) reconstruction were compared on the model with the CFL, ITCL and CL sectioned. The Schon triligamentous reconstruction and Mann triligamentous reconstruction were compared on the model with the CFL, ITCL and CL, as well as the ATFL sectioned. The inversion and external/internal rotation were quantified at different ankle positions based on the rotational moment. Then, the stress in ligaments and reconstructed grafts and the contact characteristics of the subtalar joint under inversional stress test were calculated and compared accordingly.For single ligament reconstruction, the Choisne CFL reconstruction provided the greatest degree of correction for subtalar instability, followed by the Schon CL reconstruction and then the Pisani ITCL reconstruction. For triligamentous reconstruction, the Mann procedure outperformed the Schon procedure in alleviating the subtalar instability.The finite element analysis showed that the Choisne CFL reconstruction and Mann triligamentous reconstruction provided the greatest degree of immediate postoperative subtalar stability. However, both procedures could not restore the biomechanical behaviors of the subtalar joint to normal. The long-term efficacy of these procedures warrants further investigation using a substantially larger sample of clinical cases. Subtalar instability is a common functional talocalcaneal instability always coupled with lateral ankle instability. It was reported that about 10–25% of patients with lateral ankle instability suffered from subtalar instability .The exact aetiology of subtalar instability is still unknown. Several cadaveric studies reported that the main ligaments which stabilize the subtalar joint were the calcaneofibular ligament (CFL), the interosseous talocalcaneal ligament (ITCL) and the cervical ligament (CL) –7. For cTherefore, new procedures including the Pisani ITCL reconstruction , Schon CAny procedures for joint stabilization should be evaluated for biomechanical effectiveness. The goal of these tenodesis reconstructions is generally to mimic the functions of ligaments as far as possible. To the best of our knowledge, for these newly developed tenodesis procedures, their effects in stabilizing the subtalar joint have not been compared before.Some cadaver studies have evaluated the effects of different ligaments in stabilizing the subtalar joint –7. HowevAn FE model of subtalar instability was developed based on one of our previous studies  Fig. 1)1). CT scbε-1)). The tibionavicular ligament was assigned with a linear stiffness k, which was provided [2 [2 [The ligaments of the tibia and fibula were assigned with non-linear force-displacement equations . The matprovided  Table 1bε-1). Thvided [2 , and thevided [2  and Shinvided [2  (Table 2vided [2 , 23 (Tabvided [2 . The coned [2 [2 . The hamed [2 [2 .Table 13,11 mm3,1.5 mm3 and cartilage meshes with element volume of 1.5 mm3,0.2 mm3,0.01 mm3 were generated for intact model. Meshes were considered converged when the average change in inversional angle between subsequent meshes was less than 5%. Based on the results of the mesh convergence study, all further analyses were completed with the bones meshes with volume of 1.5mm3 and cartilage meshes with volume of 0.2mm3. The final FE model consisted of 178,370 elements and 69,517 nodes.The rotational angle of the subtalar joint in inversional test were used for mesh convergence study. In the test, the fibula and tibia were flexed and a rotational moment of 4 Nm was applied on calcaneus about the medial-lateral axis. For verification, the element size was varied and the relative change of inversional angle was determined. The trial-error approach was employed in mesh convergence study . Bone meThe subtalar instability was simulated by removing the CFL, ITCL and CL, as suggested by a previous cadaveric study . Due to In previous cadaveric studies, the subtalar instability was usually simulated by sectioning the CFL, CL and ITCL , 13. ThiFive different types of tenodesis reconstruction were simulated in our FE model. The procedures of Pisani ITCL reconstruction , Schon CThe attachment point of the graft on the bone was determined by clinical approaches . One halThis procedure was simulated based on clinical approaches ,by usingThis procedure was simulated based on clinical approaches ,by usingThis procedure was simulated based on clinical approaches , by usinThis procedure was simulated based on clinical approaches , by usinTo simulate different ankle positions in numerical analysis, the loading condition was applied in two steps. First, the fibula and tibia were flexed at 10°dorsiflexion/plantarflexion or in the neutral position, while the 6 degrees of freedom of the foot were fixed. Then, the 6 degrees of freedom of the tibia and fibula were fixed too, and a rotational moment of 4 Nm was applied on the bottom of calcaneus about the medial-lateral axis (Fig. The section of CFL, ITCL and CL led to obvious instability in the subtalar joint. The inversion, external rotation and internal rotation of the joint all increased at different ankle positions Fig. . The eveThe subtalar stress test was very important for diagnosis of subtalar instability. The force in ATFL in inversional stress test at neutral position was compared and shown in Table The stress of reconstructed grafts in inversional stress test was shown in Table The additional section of ATFL only led to a slight increase of motion in the subtalar joint compared to group 1. Although the Schon and Mann triligamentous reconstructions did not recreate the ITCL, these two procedures improved the joint stability very effectively Fig. . The ManThe triligamentous reconstruction procedures recreated the ATFL, CFL and CL. The stress in the recreated ATFL, CFL and CL were shown in Table Firstly, we applied 600 N vertical force on the tibia and calculated the contact pressure and area in the intact model. The pressure nephogram of the subtalar joint revealed 3 key stress area, corresponding to anterior, middle and posterior part of subtalar joint, respectively Fig. . It suggThe subtalar joint plays an important role in hindfoot stability. Specifically, the ligamentous contents in the subtalar joint are main structures contributing to its stability. Taillard et al. found that the CFL ruptured first with an inversional force exerted on the cadaveric model, followed by the CL and then the ITCL . SeveralPrevious cadaveric studies have compared the stabilizing effect of different subtalar ligaments. However, which ligament contributes the most remains controversial. The objective of the present study was not to examine the effect of injury in one of the mentioned ligaments. Thus, the ITCL, CFL and CL were removed simultaneously to simulate subtalar instability. Nevertheless, the results of different tenodesis reconstructions can reflect the contribution of these ligaments.Pisani stressed the importance of ITCL and developed a procedure to recreate the ITCL in1996 . A one-hThe Schon CL reconstruction was developed in 1991 . One-halChoisne described a tenodesis reconstruction procedure to recreate the CFL in 2016 . The entAfter a comparison of these three procedures, we found that the stress in reconstructed grafts of the 3 single ligament reconstruction all increased compared to the corresponding ligament of intact model in inversional test. On the contrary, the force of ATFL in reconstructed models all decreased. It suggest the reconstructed grafts withstood more stress than the normal ligament. In the long term, these grafts may not be able to withstand the increased internal stress.2, respectively [2 [For subtalar instability usually coexists with lateral ankle instability , 2, the ectively , 37. Theively [2 . Obviousively [2 . The incively [2 . TherefoIn general, although several experiments have evaluated the biomechanical function of subtalar ligaments, the conclusions remain controversial. Therefore, there is no consensus on which ligament should be recreated first by the tenodesis procedure. At the same time, the injury mechanism and clinical manifestation of subtalar instability are similar to that of ankle instability, and therefore, its diagnosis is often misinterpreted. As a result, the publications of surgical reconstruction regarding subtalar instability are very limited. In addition, most of these studies are retrospective case series with limited numbers of patients –13, 29. There are several limitations in the present study. First, some simplifications were introduced in our model. The material properties of the ligaments and grafts were determined in accordance with the literature rather than actual measurements. Different material models were used for soft tissues, including both linear hyperelastic and non-linear hyperelastic models, and the viscoelastic properties of the soft tissues were not considered. These data from published literatures introduced uncertainty to this study. And, some parts of the grafts that pass through the bone tunnels were also simplified. This may also affect the forces within the ligaments. Despite these simplifications, the validation test showed that our simulation results were very close to the experimental measurements of previous cadaveric studies. Second, our FE model was only simulated based on an young male volunteer. However, the validation comparator used were cadaver specimens from old male and female person. Therefore, when we use the conclusion of this article, was must take note of its applicability. Moreover, only one model was simulated in this study. However, the contact mechanics and contact patterns would vary from person to person. Therefore, subject-specific finite element models were needed to improve the accuracy of our analysis in future.In the present study, our numerical analysis showed that the Choisne CFL reconstruction and Mann triligamentous reconstruction were most effective in restoring the kinematics and contact stress distribution in the subtalar joint. However, both procedures could not restore the biomechanical behaviors of the subtalar joint to normal. This study provided a greater understanding of the biomechanical response of ligament reconstruction in subtalar instability. The efficacy of these procedures will be investigated in a substantial number of patients with a long-term follow-up period and optimal procedures for the patients will be developed.


The paper describes the Antipsychotic Medication Management Fidelity Scale and its psychometric properties, including interrater reliability, frequency distribution, sensitivity to change and feasibility. Fidelity assessors conducted fidelity reviews four times over 18 months at eight sites receiving implementation support for evidence-based antipsychotic medication management. Data analyses shows good to fair interrater reliability, adequate sensitivity to change over time and good feasibility. At 18 months, item ratings varied from poor to full fidelity on most items. Use of the scale can assess fidelity to evidence-based guidelines for antipsychotic medication management and guide efforts to improve practice. Further research should improve and better calibrate some items, and improve the procedures for access to information.Trial registration: ClinicalTrials.gov Identifier: NCT03271242.The online version of this article (10.1007/s10488-020-01018-1) contains supplementary material, which is available to authorized users. Clinical trials and systematic reviews have established the core components of evidence-based antipsychotic medication management. Clinical guidelines make this research-based knowledge more available for clinicians. However, implementation of clinical guidelines remains fraught with difficulties . Eight sites from four health trusts in Norway, assigned by randomization, received support to implement evidence-based antipsychotic medication management. Prior to the study, all sites were providing antipsychotic medication management, but without support for following evidence-based guidelines. The Regional Committee for Medical and Health Research Ethics approved the study (REK 2015/2169), which followed the principles in the Declaration of Helsinki. Later papers on results of the study will analyze and report the relationships between implementation support and fidelity, and the relationship between fidelity and patient outcome and experiences.Below  and Prescriber Practices . Fidelity assessors rate the Policies items based on semi-structured interviews with leaders and key clinicians on policies and procedures, and on reviewing written policies or procedures. The assessors rate the Prescriber Practices items based on information in 10 randomly selected patient records, including progress notes and prescription orders over the previous three months for inpatients and the previous six months for outpatients. The items are rated according to the number of criteria met, using a summary sheet and following the calculation rules described for each item in the fidelity scale. The scoring for the scale and subscales is calculated as the unweighted sum of the item ratings divided by the total number of items. The fidelity scale with instructions is available as an Online Appendix. Table After the final assessments, the fidelity assessors completed an online survey on their experiences with the fidelity scale. Questions included if the scale was clearly set out and with good instructions, if it was easy to get information and if it was easy to rate. Additional questions addressed the usefulness of different sources of information and the instructions for using the scale.We had independent fidelity ratings (1–5) by the two fidelity assessors on the Policies items (Items 1–6) across 32 assessments (8 sites each rated 4 times). For these items and the subscale we calculated the intraclass correlation coefficient . Instead, in order to determine interrater reliability, the assessors independently rated a subset of two patient records at each fidelity site visit. Thus from the 32 fidelity assessments we obtained independent dichotomous judgments (passed/failed for each item) for 57 patient records  reviewed independently by both assessors. The two assessors divided the other 8 randomly selected patient records at the site between them to save time and still obtain ratings on 10 patient records. The assessors then jointly reached a consensus fidelity rating of Items 7–15 at each site. Based on the 57 pairs of independent ratings we calculated percentage exact agreement and Cohen’s Kappa for the 9 Prescriber Practices items. To rate each Prescriber Subscale item, assessors first made dichotomous ratings for each patient record on either 4 or 5 specific criteria. We also calculated interrater agreement on these criteria . For kappa we interpreted the degree of interrater reliability as suggested in the guidelines by Cicchetti  with theAfter assessing interrater agreement and reliability, we used consensus ratings in all subsequent analyses. To estimate internal consistency of the two subscales and the total scale, we used Cronbach’s alpha, calculating an alpha coefficient for each assessment. For alpha we interpreted the degree of internal consistency as suggested in the guidelines by Cicchetti  with theWe next examined the item distributions at 18 months, examining mean, standard deviation, and distribution of scores across sites for full (rating = 5), adequate (4), and poor (1–3) fidelity. We also examined the distribution of site scores at 18 months.z) for within-subjects design (Lakens z ≥ 0.50).Next, we examined the longitudinal pattern of fidelity graphically and statistically for the total scale and the two subscales. We examined the pattern in change over time using one-way ANOVA repeated measures with pairwise post hoc tests with Bonferroni corrections between baseline and 6 months, and between 6 and 18 months. We also analyzed sensitivity of change in fidelity from baseline to 18 months using paired t-tests for each item, the total scale and the two subscales, including reporting means and standard deviations at baseline and 18 months. Change over time was estimated by calculating the standardized mean difference effect size .From the feasibility survey we reported how much time the fidelity assessors on average spent on a fidelity visit, and their experiences with using the fidelity scale. We are not aware of any established measure for feasibility, but we interpreted feasibility to be good for a scale quality  if more than 60% of the fidelity assessors rated agreed or agreed strongly to it in the feasibility survey. All data analyses were done using SPSS version 25  of the total fidelity scale at each of the four fidelity assessments ranged from unacceptable to fair (range 0.43 to 0.75). Alpha was unacceptable (range 0.13 to 0.38) for the Policies Subscale and fair to good (range 0.69 to 0.86) for the Prescriber Practices Subscale.z = 2.66). The increase was also significant for both subscales, and Cohen’s dz was 3.09 for the Policies Subscale and 1.79 for the Prescriber Practices Subscale. At no time period did the total mean fidelity exceed 4.00, the benchmark for adequate fidelity.Figure The Policies Subscale fidelity and the Prescriber Practices Subscale fidelity both showed a development parallel to the overall fidelity with the main increase during the first 6 months, but with the Policies Subscale fidelity below and the Prescriber Practices above the overall fidelity. The Pearson correlation between the Policies Subscale rating and the Prescriber Practices Subscale rating across all sites was moderate and negative (− 0.43 and − 0.45) at baseline and 6 months, and negligible (0.03 and 0.05) at 12 months and 18 months, respectively.The 19 fidelity assessors reported that they spent on average 4.3 h (SD 1.3) on a fidelity visit, including an average of 2.9 h (SD 0.8) on reading and rating patient records. Altogether 89% agreed that the scale was clearly set out, 26% that it was easy to get information, 47% that it was easy to rate, 72% that it had good instructions, and 61% that it had good instructions for preparations. Regarding data sources, assessors reported interviews with clinicians and reading patient records were useful sources of information, while interviews with leaders were less useful.The 15-item Antipsychotic Medication Management Scale operationalized evidence-based components from the research to assess the quality of antipsychotic medication management. The interrater reliability (ICC) was excellent for the Policies Subscale and good to excellent for the subscale items. The interrater reliability (kappa) for the Prescriber Practice Subscale items was on average fair and ranging from good to poor, but the percentage exact agreement was high, which may yield poor kappas because low frequency events result in a restriction of range in item ratings Supplementary file2 (DOCX 24 kb)Below is the link to the electronic supplementary material.


N = 746). Each of the seven primary studies presented participants with high- and/or low-quality evidence and measured implausible claim endorsement and evaluations of evidence persuasiveness . A linear mixed-effect model was used to predict persuasiveness from the interaction between implausible claim endorsement and evidence quality. Our results showed that endorsers were significantly more persuaded by the evidence than non-endorsers, but both groups were significantly more persuaded by high-quality than low-quality evidence. The interaction between endorsement and evidence quality was not significant. These results suggest that the formation and maintenance of implausible beliefs by endorsers may result from less critical evidence evaluations rather than a failure to analyse. This is consistent with a limited rather than a lazy approach and suggests that interventions to develop analytical skill may be useful for minimising the effects of implausible claims.Past research suggests that an uncritical or ‘lazy’ style of evaluating evidence may play a role in the development and maintenance of implausible beliefs. We examine this possibility by using a quasi-experimental design to compare how low- and high-quality evidence is evaluated by those who do and do not endorse implausible claims. Seven studies conducted during 2019–2020 provided the data for this analysis ( Information is more abundant and accessible than ever before. The constant stream of news contains true information, as well as errors, exaggeration, and lies. Consequently, some people come to believe highly implausible claims—for example, that the COVID-19 pandemic is a hoax. These beliefs can be costly for individuals and society, making it vital to understand who believes implausible claims and why. Research suggests that a ‘lazy’ uncritical style of evaluating evidence may be associated with the formation and maintenance of implausible beliefs. Our quasi-experimental study tests whether those who endorse implausible claims evaluate high-quality or low-quality evidence differently to those who do not. We argue that if those who believe implausible claims are generally ‘lazy’ uncritical thinkers, then they will find high- and low-quality evidence equally persuasive, while non-endorsers will not. Analysis of data from seven different studies shows that high-quality evidence was more persuasive overall than low-quality evidence for both endorsers and non-endorsers. However, endorsers were more persuaded by the presented evidence than non-endorsers were. These findings suggest that those who hold implausible beliefs are sensitive to evidence quality, but are more persuaded than those who do not hold implausible beliefs. Thus, implausible beliefs may result from limited evaluative skills, rather than a ‘lazy’ thinking style.Information is more accessible now than ever before. The constant stream of material from news and social networks contains true information as well as errors, exaggeration, and lies. However, our capacity to process and evaluate the reliability of this information is limited and can lead to errors in thinking and judgment Hills . For exaThere is evidence that those who more strongly believe one implausible claim are also more likely to strongly believe other unsubstantiated claims , cognitive style is associated with the formation and maintenance of various implausible beliefs  account is one alternative explanation, which suggests that deliberation may actually bias people to favour information that aligns with their ideology—irrespective of epistemic value  from 0 to 100.https://aspredicted.org/3rv9g.pdf).This design, including the data for in/exclusion, high-/low-quality conditions, non-/endorsement criteria, and analytic approach, was preregistered before formal or informal inspection of implausible claim items, computation of endorsement status, or examination of the effects of endorsement status and evidence quality on the dependent variables . Participants from Study 7 were students recruited from a large south-western university in the USA who received course credit for their participation (n = 37). All participants completed a reCAPTCHA to ensure respondents were human . Attractiveness  was varied using images of two male experts, one rated as high in attractiveness and the other rated as low in attractiveness. Expert quality was varied by constructing a forensic gait expert who was either ‘strong’ or ‘weak’ on each of the eight attributes in the Expert Persuasion Expectancy (ExPEx) framework . Expert quality was again varied from ‘strong’ to ‘weak’ using the ExPEx framework. The strong-ExPEx/attractiveness-absent condition served as high-quality evidence for the secondary analysis. Participants in this condition read about a valid technique, used by a practitioner with relevant qualifications and extensive specialist training, who employed bias mitigation strategies, used a valid form of expression, and whose work was independently verified and agreed with by two other experts. The weak-ExPEx/attractiveness-absent condition served as low-quality evidence for the secondary analysis. Participants in this condition read about an unvalidated technique, used by a practitioner who trained in an irrelevant field, who had limited specialist training or experience, who was ignorant of and displayed bias, who provided invalid opinions, and whose work was not independently reviewed or verified by relevant experts.Had the same primary aim and attractiveness manipulation as Study 1. However, in this study participants evaluated a lengthy (15-page) trial transcript adapted from the real testimony of an expert witness providing speech spectrography evidence . There were three types of judicial admissibility decision in this study: control, explicit-admit, explicit-exclude. These conditions were operationalised the same way as in Study 3. The experimental materials also included information about ‘high’ or ‘low’ expert ability. In the high-ability conditions, participants were told that the engineering professor providing evidence had scored 90% accuracy on relevant proficiency tests. In the low-ability conditions, participants were told that the engineering professor providing evidence had scored 50% accuracy on relevant proficiency tests. The high-ability/explicit-admit condition served as high-quality evidence for the secondary analysis. The low-ability/explicit-exclude condition served as low-quality evidence in the secondary analysis.Examined the effects of expert ability and judicial admissibility decisions on evidence persuasiveness (preregistered at https://tinyurl.com/yyjsvzad). Participants read a report either about a high-reliability  or low-reliability forensic discipline . The report provided either a detailed- or a sparse-disclosure of important information about the evidence and opinion. In the detailed disclosure conditions, the report was modelled on best-practice recommendations for expert reports submitted to police and courts . Participants in this study read and evaluated the same detailed expert footwear comparison report before completing one of three different measures to assess their reasoning. The report contained three important flaws that undermined the quality of the evidence. Specifically, the report contained information that the expert performs with 45–55% accuracy on relevant proficiency tests, a fallacy in the reporting of the results . Participants in this study read an opinion from a DNA analyst stating either the ‘biased’ (race-specific) or ‘unbiased’  assumptions associated with the analytic method. Analyses completed using race-specific rather than race-neutral DNA databases are often conducted to produce more conservative random match probability estimates that inflate the likelihood that the defendant was the source of DNA associated with a crime  How valuable was the evidence? From 0 ‘not at all’ to 100 ‘definitely valuable’; (3) How much weight do you give to the evidence? From 0 ‘none at all’ to 100 ‘the most possible’. Participants in Study 6 only answered question three.Participants in Studies 1–5 and 7 answered three questions about the specific type of evidence they were presented using on-screen sliders: (1) Sharks are mammals and A kilogram is heavier than a gram) from 0 ‘not at all’ to 100 ‘definitely true’. Two of the three implausible claims included in the battery were based on items used in past research: Vaccines are harmful, and this fact is covered up . The general knowledge battery containing the implausible claims was presented after all study-specific dependent measures and before the demographic questions . Finally, all participants were debriefed and thanked for their participation.lmer  by 85 participants (11.4%), the vaccine claim was endorsed by 44 participants (5.9%), and the flat earth claim was endorsed by 19 participants (2.5%). See “M = 80.3, SD = 20.4) than low-quality evidence  = 13.45, p < 0.001, 95% CI , Cohen’s d = 1.04; see Fig. M = 67.3, SD = 30.9) than non-endorsers  = 3.33, p < 0.001, 95% CI , d = 0.24). The interaction between endorsement and evidence quality was not significant  = 1.74, p = 0.083, 95% CI , d = 0.13), but it is important to note that this result does not constitute evidence against such an interaction. Endorsers’ ratings of low-quality  and high-quality evidence  did not significantly differ from non-endorsers ratings of low-quality  and high-quality evidence . See “N = 1,747) and different definitions of non-/endorsement status.Overall, participants were significantly more persuaded by high-quality  that levels of analytical reasoning did not significantly predict ability to discriminate between true and fabricated stories. Both of these results are inconsistent with a generalised failure to think. Thus, it may be a mistake to infer that the more intuitive, reflexive cognitive style of endorsers shows that they are lazy and do not analyse (Pennycook and Rand Overall, endorsers were more persuaded by the presented evidence than non-endorsers. This general overvaluing could be because endorsers were relatively more optimistic about the strengths of evidence, and/or less pessimistic about the weaknesses of the evidence—although the former appears more likely given our data. Either way, the result suggests that endorsers differ from non-endorsers in their perceptions of what is or should be persuasive. Consequently, we may need to consider different strategies for reducing implausible belief formation and maintenance than those typically described in the literature.impact of various strengths and weaknesses on evidence quality. Thus, interventions focused on building analytical competence—for instance through education about research methods or threats to validity (McAuliff et al. Researchers examining implausible beliefs and cognitive style have tended to advocate for interventions that will shift people towards a more deliberative, reflective analytical strategy, for example, by ‘slowing down for a moment’ (Ward and Garety It is important to be aware of some limitations when considering our results. First, we did not explicitly measure the cognitive style of our participants using, for example, the CRT or the AOT. As a result, we do not know whether endorsers in our sample had a more or less reflective analytical style than non-endorsers. We can only say that endorsers engaged in a reflective form of evidence evaluation that resulted in high-quality evidence being rated as more persuasive than low-quality evidence. Future research could measure both analytical performance and cognitive style to examine whether aspects of cognitive style can help to explain the differences between endorsers and non-endorsers that we observed.It is also important to acknowledge that we used an ad hoc approach for assessing beliefs in implausible claims. We included three implausible claims in a general knowledge test battery and classified those who strongly believed any one of the claims as ‘endorsers’, and those who regarded all of them as more false than true as ‘non-endorsers’. This approach may have resulted in over- or under-inclusive definitions, which in turn could affect our results. However, the distribution of endorsement ratings suggests it is unlikely that the composition of endorsement groups would substantially change if we used more or less conservative definitions see “”. We alsOverall, our study suggests that it is not laziness that separates those who believe implausible claims from those who do not. Instead, limited analytical skills may play a role in the development and maintenance of a range of implausible beliefs. These limitations could be addressed through interventions targeting evaluative performance. However, further research examining the relative contributions of cognitive style and analytical skill is vital for developing the most effective interventions to minimise the harms caused by implausible beliefs.


The increasing prevalence of delayed and missed diagnoses for dementia constitutes major public concern. In this regard, inadequate knowledge and poor understanding of the condition may create a barrier to timely dementia screening. This cross-sectional study assessed dementia literacy, then identified the association between dementia literacy and willingness to undergo routine dementia screening among community-dwelling older adults in two urban areas of Japan. More specifically, structured questionnaires were distributed to a total of 854 individuals aged ≥ 65 years. A multivariate logistic regression was then used to explore the factors associated with dementia literacy and willingness to undergo routine dementia screening. Results showed that younger respondents and respondents who received dementia information from television/radio and/or paper-based sources were more likely to have high dementia literacy. While less than half of participants were willing to undergo routine dementia screening, those with higher dementia literacy were more willing to do so . Although there are pros and cons to routine dementia screening, it is necessary to implement such a system to detect dementia and cognitive impairment. Further, assessments should also attempt to gain information about individual beliefs and understandings related to dementia information. A recent systematic review of the global burden of diseases reported that 43.1 million people are living with dementia worldwide, with the number more than doubling from 1990 to 2016 [Health literacy is defined as the range of cognitive and social skills that enable people to obtain, understand, and use information for the purpose of enhancing their health and well-being and engage in healthcare-related decision-making . Health As mentioned above, there are both pros and cons to routine dementia screening. For example, the US Preventive Service Task Force concluded that there was a lacking of direct evidence for the benefits of early screening to detect cognitive impairments in older adults . DementiJapan has one of the highest rates of population aging in the world. In this context, current estimates indicate that more than 7 million people aged 65 years and older will have dementia by 2025, with the total costs of the disease expected to increase 1.6 times between 2014 and 2060 . For thoThis study was conducted in an area adjacent to the cities of Akashi and Kobe in Japan. Both initiated subsidized systems for dementia screening for older adults in 2019 fiscal year. In the same year, an initial survey was conducted among all community-dwelling persons aged 65 years and above who lived in this area (details are given elsewhere) . Among tThe outcomes of the variables were set as dementia literacy and willingness to undergo regular dementia screening. The dementia literacy component was developed based on eight questions developed by Zhang et al.  specificAs relevant information sources are related to dementia literacy , particiDescriptive statistics were applied to calculate the proportions and numbers of each variable. Each question on dementia literacy was scored as either correct (1 point) or incorrect/do not know (0 points) . The average dementia literacy score was 4.2, thus categorized as “high dementia literacy” based on a threshold of ≥4 points, with “low dementia literacy” being determined at scores <4 points. Willingness to undergo dementia screening was scored as either “yes” (1 point) or “no/do not know” (0 points). A chi-squared test was then conducted to identify the factors associated with dementia literacy and willingness to undergo regular dementia screenings while a multiple logistic regression model was applied to control for confounding factors. All statistical analyses were performed using the STATA 16.0 software  and statistical significance was set to 5%.All participants received written explanations of the study protocol. We then obtained written, informed consent from those who agreed to participate. This study received ethical approval by the Ethics Committee of Kobe Gakuin University .The average participant age was 78.2 years (range of 65–100), with 53% being female. Among all participants, 14% were assessed as possibly having dementia (DASC-21 ≥ 31), while 28% had depressive symptoms (GDS-5 ≥ 2).Only 1.7% of the respondents answered all dementia literacy questions correctly. The question with the highest rate of correct answers was that on memory loss from dementia (88.2%), while the lowest rate was for the item on the prevalence of dementia in Japan (22.7%) . The majA multivariate logistic regression showed that younger respondents and those who obtained dementia information from television/radio and/or paper-based sources were more likely to have high levels of dementia literacy .Among all respondents, 41% were willing to undergo regular dementia screening, with more than half being unwilling or undecided. The main reasons for unwillingness were “bothersome to visit the clinic” (42%) and “do not know which doctors can be consulted” (32%) .A univariate logistic regression model showed that respondents with higher dementia literacy were more likely to accept regular dementia screenings than those with low dementia literacy (odds ratio (OR) = 1.40, 95% confidence interval (CI) = 1.02, 1.92). However, there were no statistically significant differences after controlling for other variables . Older adults with poor self-rated health were more likely to undergo regular dementia screening than those with better self-rated health  .This study assessed dementia literacy and willingness to undergo regular dementia screenings among community-dwelling older adults. Results showed that respondents of younger ages had higher dementia literacy after controlling for educational level, depression, and cognitive impairment. A systematic review reports that older age was strongly associated with limited health literacy , while aIn this study, we found that the majority of participants gathered dementia-related information from a variety of sources, and diversity of sources was significantly associated with higher dementia literacy. Among the variety of sources, respondents who obtained dementia information from television/radio and/or paper-based sources had higher overall levels of dementia literacy than those who did not. Similar findings were reported by a previous study targeting older adults in Vietnam, which showed that television and Internet usage were associated with higher health literacy . An empiOur initial survey found that 60% of community-dwelling older adults intended to undergo dementia screenings using the subsidized system for dementia diagnosis . AlthougThis study showed that higher dementia literacy was marginally associated with an increased preference for regular dementia screening. A previous literature review reported that low levels of awareness about screening led to misunderstandings about both the reason for dementia tests and the implication of test results . AnotherFurther, participants with poor self-rated health status were more willing to undergo regular dementia screenings than those with better self-rated health status. A study involving older adults in the USA also found that unhealthy individuals were more willing to undergo routine dementia screenings , while aThis study also had limitations. First, respondents agreed to participate in the follow-up survey after receiving information on the study purpose. In this regard, these may have generally been more interested in dementia and were, thus, more active in collecting dementia information. Sampling bias may have, therefore, led to an overestimation of dementia literacy. Second, content validity of the instrument for measuring dementia literacy was not high . Various instruments and aspects of dementia literacy have been used for dementia research ; thus, fMore than half of this study’s participants were unwilling to undergo routine dementia screenings, with many referring to the burdens associated with clinic visitation and the lack of sufficient information about screening. It is, therefore, important to increase access to adequate dementia information, which should raise general awareness about dementia and, thus, improve timely diagnosis of dementia. Further, sources such as television/radio and/or paper-based information may be effective for improving dementia literacy, which influences decisions to undergo dementia screening and receive advance care. Although there are pros and cons to routine dementia screening, a system of detecting dementia and cognitive impairment should be implemented to better understand individual beliefs and understandings related to the disease. Further research also needs to clarify which interventions are effective in increasing dementia literacy as well as changing perspectives on undergoing dementia screening among community-dwelling older adults.