Spiritual walking and stress resilience integrating psychological endocrine and genetic evidence from a longitudinal Iraqi cohort

Abstract

Background

Physical activity benefits stress and anxiety, yet spiritually motivated walking may confer distinct psychobiological advantages compared with competitive sports. While prior studies have been largely cross-sectional, the longitudinal dynamics of psychological, endocrine, and genetic interactions remain underexplored.

Methods

We conducted a mixed cross-sectional and longitudinal study including three hundred adults in Iraq (n = 100 athletes, n = 100 spiritual walkers, n = 100 sedentary controls). Subjects were tracked across four waves: baseline (T0), immediate post-activity (T1), 1-month follow-up (T2), and 3-month follow-up (T3). The results revealed perceived stress (PSS-10), state anxiety (STAI-S), salivary cortisol and DHEA-S (ELISA), and genotyping of BDNF Val66Met (rs6265) and 5-HTTLPR. Analyses used ANOVA/ANCOVA, linear mixed models (LMM), and regression models with FDR correction.

Results

Across the waves studied, spiritual walkers consistently showed lower levels of stress and anxiety compared to both athletes and control groups (all p ≤ 0.01). Longitudinal linear mixed models corroborated enduring disparities at T3, wherein the spiritual walkers exhibited the most advantageous psychological profile. Endocrine assessments indicated diminished cortisol levels and increased DHEA-S concentrations in walkers, resulting in a persistently favorable cortisol/DHEA-S ratio relative to the other groups. Genetic analyses indicated that BDNF Met/Met and 5-HTTLPR S/S carriers reported heightened distress; however, these effects were less pronounced among walkers. An integrated model elucidated 51.8% of the variance in anxiety, identifying cortisol (β = 0.42) and DHEA-S (β=−0.35) as the most significant predictors.

Conclusions

Spiritually motivated walking correlated with persistent decreases in stress and anxiety, along with improved regulation of the HPA-axis and a reduction in genetic vulnerability, effects that persisted for up to 3 months following the intervention. These results underscore the practical applicability of culturally relevant, affordable interventions that incorporate spiritual walking into community mental health initiatives, considering biological factors.

Introduction

Physical activity is widely recognized for its numerous benefits on mental health, including the lowering of stress and anxiety and depressive symptoms along with improvements in cognition and quality of life [1–3]. owever, most academic research has been primarily focused on structured competitive sport. Performance pressure, the negative outcomes of indivisible or team failure, overtraining and evaluative competition are some psychological stressors involved in competitive sports that will on occasion blunt mental health benefits. In contrast, spiritually driven practices—like long-distance walking at religious ceremonies—are under-studied despite their capacity to generate unique psychobiological adaptations [4, 5].

In Iraq, a multitude of individuals engage annually in spiritual walking rituals, most notably the Arbaeen pilgrimage, traversing distances of up to 80 km under conditions that are both physically taxing and socially enriching while possessing significant spiritual importance [6]. In contrast to training focused solely on performance, these rituals integrate prolonged physical exertion with deep religious significance and communal solidarity, thereby providing a unique natural setting to examine stress-related processes that are distinct from those observed in traditional athletic endeavors [7]. The role of genetic polymorphisms including 5-HTTLPR and BDNF Val66Met in determining stress responsivity and resilience is supported by evidence from psychoneuroendocrinology and psychogenetics [7–9]. Hormonal markers specifically cortisol DHEA-S and their ratio are reliable indicators of hypothalamic–pituitary–adrenal (HPA) axis activity and stress adaptation [10, 11] Although athletes frequently show elevated cortisol and lower DHEA-S/cortisol ratios under competitive demands [12, 13]. Spiritually enriched environments seem to reduce HPA activity and buffer psychological distress [14, 15]. Recent data also suggests that neuroinflammatory mechanisms and epigenetic control are important mediators of the relationship between stress and mental health consequences. Alterations in neuroplasticity and mood dysregulation can result from long-term activation of the hypothalamic–pituitary–adrenal (HPA) axis which can increase pro-inflammatory cytokines like IL-6 and C-reactive protein (CRP) [16]. In parallel genes controlling synaptic plasticity and emotion regulation such as Brain-derived neurotrophic factor (BDNF) and serotonin transporter pathways have been demonstrated to undergo DNA methylation and histone modifications in response to stress [17, 18]. These results imply that immune-inflammatory and epigenetic mechanisms are also part of the biological embedding of stress which goes beyond endocrine responses. Testing integrative models of resilience that cover psychological endocrine genetic and molecular dimensions is made possible by examining such pathways within culturally embedded physical-spiritual practices. Despite these realizations no research has yet to directly compare the psychological effects endocrine markers and genetic susceptibility of competitive athletes and spiritual walkers in a single cohesive framework. In order to close this gap, the current study examines how biological predispositions and the motivational context of physical activity interact to affect stress and anxiety. Rather than focusing on general recreational exercise this study examined two specific activity contexts: competitive sport and walking driven by spiritual motivation. The diverse range of intensity purpose and structure among regular exercisers may make it difficult to interpret the psychological and biological effects of their activities. Thus, the study main goal was to find out how many advantages people who engage in spiritually inspired walking rituals experience. Compared to other types of exercise spiritual walking may have a greater impact on stress management because it blends extended physical activity with social interaction and personal significance. Research indicates that religiosity and spirituality may be protective for mental health systematic reviews have linked lower rates of anxiety and depression in youth to spiritual engagement [19]. Additionally, social support has been linked theoretically and empirically to reduced perceived stress and better psychological outcomes, consistent with stress and coping frameworks [20]. These findings provide a broader theoretical basis for Hypotheses 1 and 2.

We hypothesized that:

H1

Spiritual walkers would report lower stress and anxiety compared with athletes and sedentary controls.

H2

Spiritual walkers would exhibit lower cortisol, higher DHEA-S, and a more favorable cortisol/DHEA-S ratio than other groups.

H3

Carriers of BDNF Met and 5-HTTLPR S alleles would show higher distress, but these genetic risks would be buffered in spiritual walkers due to enriched social–spiritual context.

Materials and methods

Study design

The study employed a mixed observational design, including a cross-sectional comparison between groups and a longitudinal follow-up across four assessment waves [21].

Participants

Participants were classified into three groups. Athletes had a minimum of 3 years of continuous professional or semi-professional training. Spiritual walkers had completed at least 40 km during the Arbaeen pilgrimage. Sedentary controls reported less than 1 h per week of structured physical activity. Exclusion criteria: acute illness, major psychiatric or endocrine disorders, and current glucocorticoid use. Written informed consent was obtained from all participants. We obtained written informed consent from each participant. In compliance with the 2013 revision of the Declaration of Helsinki Al-Furat Al-Awsat Technical University Institutional Review Board granted ethical approval.

Table 1.

panel H1,H2,H3 illustrated group differences in salivary cortisol,DHEA-S and cortisol /DHEA-S ratio between waves

Participant recruitment

Participants were enrolled in the Iraqi cities of Najaf and Karbala using group-specific procedures. Athletes were recruited from local sports clubs in Najaf and Karbala. Coaches and club administrative staff notified eligible athletes of the study, and they contacted interested individuals to determine eligibility. Collaboration with medical centers offering healthcare services to pilgrims along the pilgrimage route was essential for recruiting participants in this study of the Arba’een pilgrimage between Najaf and Karbala. Research assistants stationed at these medical centers provided information about the study to potential participants and invited eligible individuals who were interested in participating.

Participants in the control (inactive) group were recruited from the general population of Najaf and Karbala through community outreach and university networks. Individuals who reported engaging in less than 1 h of structured physical activity per week were considered eligible for the control group.

Study setting

Data collection was conducted in the cities of Najaf and Karbala, Iraq.

Sample size and power

The cross-sectional ANOVAs a priori power analysis (f = 0.20 α = 0.05 power = 0.90) showed that N = 252 it was exceeded by the final N = 300. N = 90/group attained ≈ 0.85 power for longitudinal Group×Time interactions in linear mixed-effects models (LMMs) assuming f = 0.12–0.15 and within-person correlation ρ ≈ 0.50 across four waves. To account for attrition we set the target at N = 100/group.

Measures

Psychological outcomes

Perceived stress was measured using the 10-item Perceived Stress Scale (PSS-10) [22], which assays the degree to which situations in one’s life are appraised as stressful. The assessment of the anxiety level was using the State subscale of the State–Trait Anxiety Inventory (STAI-S) [23], measuring transient anxiety related to current situational stress. Spiritual well-being was evaluate using the Functional Assessment of Chronic Illness Therapy–Spiritual Well-Being scale (FACIT-Sp12), a 12-item instrument assessing meaning, peace, and faith-related aspects of spirituality [24]. Social support was evaluated with the Medical Outcomes Study (MOS) Social Support Survey, which estimate perceived availability of emotional, informational, and tangible support [25]. Sleep quality was evaluated using the Pittsburgh Sleep Quality Index (PSQI) [26], a widely used 19-item measure that captures subjective sleep quality, latency, duration, and disturbances over the past month. Physical activity levels were assessed using the short form of the International Physical Activity Questionnaire (IPAQ-SF) [27], which estimates weekly energy expenditure across walking, moderate, and vigorous activities. Validated Arabic versions of all instruments were employed, each demonstrating strong psychometric reliability and validity in prior studies [28].

Neuroendocrine

Salivary cortisol and dehydroepiandrosterone sulfate (DHEA-S) were determined in morning saliva samples obtained between 08:00 and 09:00 a.m. after an overnight fast. Salivette® cotton swab collection devices (Sarstedt, Germany) were used to collect the samples. Samples were immediately centrifuged and all supernatants were stored at − 80 °C until analysis. Concentrations of hormones were determined in duplicate by commercially available enzyme-linked immunosorbent assay (ELISA) kits according to the manufacturer’s instructions. All laboratory analyses were conducted in the research laboratory of Al-Furat Al-Awsat Technical University. The intra-assay and inter-assay coefficients of variation (CV) were both below 8%, indicating acceptable assay precision and reliability [29, 30].

Genetic outcomes

Genetic analyses focused on the serotonin transporter-linked polymorphic region (5-HTTLPR) and brain-derived neurotrophic factor (BDNF) Val66Met polymorphism which are two noted candidate polymorphisms associated with stress responsiveness and affective regulation [31, 32]. These variations were chosen because other studies have shown that they are linked to things, like regulation and stress sensitivity and also resilience. The researchers picked these variations because they are connected to regulation and they are also connected to stress sensitivity and emotional regulation and resilience. Exploratory polygenic techniques were used in addition to single-variant analyses in order to capture cumulative genetic liability [33]. Using an additive model polygenic risk scores (PRS) were calculated across roughly 10–20 single nucleotide polymorphisms (SNPs) that have been linked to affective regulation and stress responsiveness in the past. Before being included in regression models risk alleles were standardized to a mean of 0 and a standard deviation of 1 and coded additively (0 1 or 2). This method offered a more thorough depiction of genetic susceptibility and made it possible to integrate several small-effect loci beyond single candidate genes [34].

Procedures

For comparability assessment, methods were standardized for each group. To prevent severe competition-related stress, athletes were evaluated during their competitive season but outside of important competitions. Spiritual walkers were evaluated 24–48 h after the pilgrimage was over. During their regular daily lives, sedentary control participants were evaluated without being exposed to any particular organized physical or spiritual activities. To reduce inter-assessor variability, the same skilled research team performed all of the assessments.

Survey administration

Psychological questionnaires were administered using paper-based forms. Participants completed the questionnaires on site under the supervision of trained research assistants who were available to clarify instructions when necessary. Completed questionnaires were checked for completeness at the time of collection to minimize missing responses.

Biospecimen standardization

Spiritual walkers were assessed within 24–48 h after completing the Najaf–Karbala pilgrimage route.

Statistical analysis

Two stages of statistical analyses were carried out in succession. The primary group features were characterized in the first phase by analyzing cross-sectional differences between groups at baseline (T0) using analysis of variance (ANOVA) for unadjusted comparisons and analysis of covariance (ANCOVA) when adjusting for predefined covariates. Non-parametric Kruskal-Wallis tests were used when distributional assumptions were broken. The Bonferroni method was used to correct subsequent pairwise comparisons.

In the second phase intraindividual dependencies resulting from repeated measurements were taken into account by employing linear mixed-effects models (LMMs) to analyze longitudinal changes across four evaluation waves (T0–T3). The independence assumption is avoided by using this two-phase analysis approach which also makes it possible to distinguish clearly between baseline group differences and individual changes over time. For all analyses R (version 4.2) and SPSS (version 26) were used.

In order to model the outcome variables as a function of group time and their interaction (group × time) longitudinal mixed-effects models were employed. These models included random intersections for each participant and random slopes for time where the model fit was favorable. To prevent presumptions of linear change across phases time was considered a classifying factor. The main method used to check for variations in group trajectories was the group × time interaction.

Co-opting variables were regularly included in the outcome groups and were pre-selected based on prior research and biological plausibility. These factors included the participants age sex body mass index medication use smoking status caffeine intake menstrual cycle phase for females and training load for athletes. P-values were adjusted using the Benjamin-Hochberg false discovery rate (FDR) procedure to account for multiple hypothesis testing. To aid in interpretation and reproducibility effect sizes are presented with p-values such as standardized β coefficients for regression and integration models Cohens d coefficient for pairwise comparisons and η² for overall group effects. Conventional standards (small medium and large effects) were used to interpret effect sizes.

Mechanistic analyses were performed after the initial group comparisons to explore potential explanatory pathways. Moderation analyses were used to investigate whether genetic factors modified the association between group context and stress outcomes, whereas mediation analyses were used to test whether endocrine markers (cortisol, DHEA-S) explained associations between predictors and psychological outcomes. These models were implemented within a multilevel structural equation modeling framework to account for the nested structure of repeated observations.

The main causes of the missing data were some item-level data loss and wave-level non-response. Twenty assigned datasets were generated assuming random missing data (MAR) in order to handle the missing data using multi-sequence equation assignment (MICE) in order to reduce bias and preserve statistical power. The assignment model comprised concomitant variables, time group, and all primary outcomes. Rubin’s guidelines were followed in compiling the estimates. Further robustness checks included the use of robust estimators and the exclusion of hormonal abnormalities greater than three standard deviations. Sensitivity analyses compared results from assigned datasets with whole-case analyses.

Transparency and ethics

The study protocol and analysis plan were preregistered on the Open Science Framework (https://osf.io/j53fq/). De-identified data and analysis code will be shared upon publication. Reporting adhered to STROBE/STROBE-ME [35].

Results

There are two sections to the results presentation. Initially, analysis of variance (ANOVA) and analysis of covariance (ANCOVA) are used to report the differences between groups at baseline (T0). Next, using linear mixed-effects models (LMMs), which serve as the fundamental inferential framework for repeated-measures analyses, the longitudinal effects across four waves (T0–T3) are reported.

Psychological outcomes

Both state anxiety (STAI-S) and perceived stress (PSS-10) showed statistically significant differences between groups according to basic cross-sectional analyses at the starting point (T0). Group had a statistically significant main effect on both PSS-10 (F (2297) = 32.4 p0.001 η² = 0.179) and STAI-S (F (2297) = 27.8 p0.001 η² = 0.158) according to analysis of variance (ANOVA). Spiritual walkers reported significantly lower levels of stress and anxiety compared with both athletes and controls. according to corrected post-analytical tests using Bonferronis correction (all p-values ≤ 0.01).

Time-series longitudinal analyses with linear mixed-effects models validated these group differences. Different trajectories between the groups were indicated by significant group-time interactions (p0.01) and significant main group effects on the State Anxiety and Intensity Scale (STAI-S) and the Perceived Psychological Stress Scale (PSS-10) (both p0.001). Throughout all four phases (T0–T3) the spiritual practitioners group continuously displayed the lowest levels of stress and anxiety followed by the athletes with intermediate levels and the control group with the highest scores as shown in Fig. 1, perceived stress levels differed significantly between groups. Similar findings were obtained using adjusted models that controlled for body mass index age sex and other predetermined covariates.

Fig. 1.

(Panel P1P2). Psychological results between groups and time. A Methodological Synonyms (mean SEM) of State Anxiety (STAI, S, solid lines) and Perceived Stress (PSS, 10, dashed lines) between four assessment waves (T0-T3). Spiritual Walkers were the group with the lowest scores at all times, Athletes showed moderate levels, and controls had the highest (p < 0.01, Bonferroni, corrected). B Boxplots of STAI, S scores by group and wave, confirming significant group differences across timepoints. Athletes scored lower than Controls, while Walkers kept the most favorable profile. Asterisks now indicate the significant differences between groups at each assessment wave without brackets, ensuring consistency with the other panels. The explanation of the significance symbols is given in the figure caption. Asterisks indicate statistically significant between-group differences at each wave(* p < 0.05, ** p < 0.01, *** p < 0.001; Bonferroni-corrected)

Endocrine outcomes

The levels of salivary cortisol dehydroepiandrosterone sulfate (DHEA-S) and cortisol-to-DHEA-S ratio were statistically significant (all p 0.001) at baseline (T0) according to cross-sectional analyses. Significant differences were observed in cortisol levels between groups (F (2297) = 41.2 p 0.001 η² = 0.217) with the control group exhibiting the highest concentration athletes exhibiting intermediate levels and spiritual practitioners exhibiting the lowest. Conversely the control group had the lowest DHEA-S levels and spiritual practitioners had the highest (F ((2297) = 28.9 p 0.001 η² = 0.163).

The main group effects on cortisol DHEA-S and cortisol-to-DHEA-S ratio levels were statistically significant according to longitudinal linear mixed-effects models (all p 0.01). Hormonal pathways varied between groups over time as evidenced by statistically significant group-time interactions. Participants in the Spiritual March program maintained a consistent hormonal profile throughout all follow-up phases as illustrated in Fig. 2 with low cortisol high DHEA-S and a low cortisol-to-DHEA-S ratio. Following sensitivity analyses that ruled out atypical hormonal values and concomitant variable adjustments these results stayed the same.

Fig. 2.

(Panels H2,H3,H4). Salivary endocrine markers for different groups over time. A Cortisol concentrations across four waves (T0–T3) are displayed in boxplots, revealing that Controls consistently had the highest levels, Athletes had the levels in between, and Spiritual Walkers had the least levels. B The DHEA-S boxplots show that the spiritual walkers had the highest concentration, followed by Athletes, controls with the lowest levels. C The ratio cortisol to DHEA-S, which is a measure of HPA axis homeostasis, was much lower in Walkers and Athletes in comparison to Controls at all timepoints, thus indicating a better stress-resilience profile. Asterisks indicate significant between-group differences at each wave (Bonferroni-corrected). *p < 0.05, **p < 0.01, ***p < 0.001 (Bonferroni-corrected)

Genetic outcomes

All genotype distributions were consistent with Hardy-Weinberg equilibrium (χ² tests p 0.05 ) and there were no significant differences between groups for BDNF Val66Met and 5-HTTLPR. This supported the appropriateness of further analyses. For some genotype classes the genetic analyses were deemed exploratory due to the small sample sizes and the restricted number of candidate polymorphisms. According to regression analyses people who were homozygous for Val/Val had lower anxiety scores than those who carried the BDNF Met allele (β = 0.19 95% confidence interval [0.04–0.34] p 0.05). Groups were not entirely distinguished by individual polymorphisms. However compared to control groups spiritual and athletic people had significantly lower polygenic risk scores (PRS) (β=−0.28 p0.01) which is consistent with a more flexible genetic profile (Fig. 3).

Fig. 3.

Genetic analyses: correlation between distribution, psychology, and polygenic risk. A The stacked bar charts shows the percentage composition of the BDNF Val66Met (Val / Val, Val / Met, Met / Met) and 5 HTTLPR (L / L, L / S, S / S) genotypes in Athletes, Spiritual Walkers, and Control individuals. B The STAI-S (State Anxiety) boxplots show significantly higher levels of anxiety for Met carriers than for Val / Val homozygous individuals (p < 0.05). C The Polygenic risk score boxplots indicate that Athletes and Spiritual Walkers exhibited significantly lower PRS compared to Control group (p < 0.01) therefore displaying more resilient profiles

Integrative analyses

Higher scores on the Situational Anxiety Scale (STAI-S) and the Perceived Psychological Stress Scale (PSS-10) were found to be negatively correlated with dehydroepiandrosterone sulfate (DHEA-S) levels social support and spiritual well-being and positively correlated with salivary cortisol levels according to bilateral correlation analyses (all |r| values ≥ 0.30 p 0.01 Fig. 4).

Fig. 4.

Relationship heatmap across psychological, endocrine, and social spiritual measures. The correlation matrix shows the relationships between psychological outcomes (STAI-S, PSS, 10), endocrine markers (Cortisol, DHEA-S), and social spiritual resources (Support, Spirituality). Higher levels of anxiety and perceived stress related positively to Cortisol and negatively to DHEA-S, social support, and spirituality. In contrast, DHEA-S had positive correlations with support and spirituality, indicating protective pathways. These connections highlight the link between psychological, biological, and social spiritual factors in stress resilience

Using longitudinal framework integrative pathway models were estimated to investigate multivariable pathways. DHEA-S and cortisol were found to be significant predictors of situational anxiety with DHEA-S exhibiting a negative correlation (β ≈ − 0.35) and cortisol exhibiting a positive correlation (β ≈ 0.42). Up to 51.8% of the variation in anxiety scores was explained by these models. In line with proposed biopsychosocial pathways the groups effects were indirectly combined through their associations with psychosocial resources and hormonal profiles. Figure 5 displays standardized coefficients to make comparing effect sizes across predictive variables easier.

Fig. 5.

State anxiety was assessed at four times (T0-T3) through the use of time-varying regression coefficients (TVC) (e.g., Cortisol vs. DHEA-S). At each time, standardized regression coefficients (β) for Cortisol and DHEA-S and their corresponding 95% confidence intervals were computed at the subject level. Cortisol was consistently a predictor of anxiety at all times; however, DHEA-S was a consistent predictor of decreased anxiety, thus providing a protective effect. The changing nature of the relationships between endocrine markers and psychological outcomes suggest that these relationships are stable over time, thus supporting the role of endocrine markers as mediators of resilience to stress

Discussion

This study examined psychological, endocrine, and genetic correlates of stress and anxiety among competitive athletes, spiritual walkers, and sedentary controls in a sufficiently powered multi-wave cohort (N = 300). Findings demonstrate that spiritually motivated walking, exemplified by the Arbaeen pilgrimage, is associated with reduced psychological distress, An important consideration is whether the observed group differences primarily reflect the acute effects of the pilgrimage or athletic season, or rather the broader lifestyle and psychosocial context of these groups. According to our longitudinal results, group differences were already noticeable at baseline and stayed mostly consistent throughout follow-up waves. This suggests that long-term lifestyle factors, like regular exercise, social integration, and meaningful engagement, might be more important than the actual acute physical events. favorable regulation of the HPA-axis and reduced genetic susceptibility to stress (Table 2).

Table 2.

Baseline demographic and anthropometric characteristics of the study groups

Characteristic	Athletes (n = 100)	Spiritual Walkers (n = 100)	Controls (n = 100)	p-value (ANOVA/χ2)
Age, years (mean ± SD)	26.4 ± 4.8	27.1 ± 5.2	26.9 ± 5.0	0.62
Sex, male (%)	58 (58%)	55 (55%)	57 (57%)	0.89
BMI, kg/m2 (mean ± SD)	23.7 ± 2.8	24.0 ± 3.1	24.2 ± 2.9	0.48
Education, ≥ college (%)	72 (72%)	70 (70%)	68 (68%)	0.77
Smoking status (%)	19 (19%)	17 (17%)	21 (21%)	0.84
Sleep duration, h/night	7.2 ± 0.9	7.3 ± 1.0	7.1 ± 0.8	0.55

Psychological well-being and spiritual practice

The lowest stress and anxiety scores were consistently reported by spiritual walkers. In contrast to athletes, whose mental states are frequently influenced by pressures related to performance, walkers benefited from ritual rhythm, strong social support, and intrinsic meaning. It appears that the processes of making sense of our lives have an important association with the psychological benefits associated with behaviors like walking spiritually. The structured narrative, which defines or characterises transcendence through an experience of divine purpose and a sense of belonging offered by many spiritual and/or religious practices, forms a key component to our ability to cope with difficult experiences through developing meaning in our lives. It is important to note that there may be advantages associated with walking spiritually through meaning-making processes, rather than through spirituality per se. Further investigations are needed to clarify the distinction. This supports earlier research showing that spirituality improves meaning-making and resilience [36–38]. The results extend the concept of “spiritual hormesis” [39], whereby moderate exertion in a meaningful context fosters adaptive stress tolerance.

Neuroendocrine biomarkers

In terms of their physiological stress regulation, walkers showed significantly reduced levels of cortisol and higher levels of DHEA-S resulting in a favorable ratio of cortisol/DHEA-S, which is an established marker of resilience and proper functioning of the hypothalamic-pituitary-adrenal (HPA) axis [40, 41]. The athletes were at their highest levels of cortisol, which is expected given the stress associated with the training they had been doing, while the control group was shown to have the least adaptive physiological stress regulation profile. These findings demonstrate how cultural and spiritual factors can affect one’s physiological stress regulation by creating different patterns than what is typically observed during competition.

Genetic moderators of stress sensitivity

Similar to previous studies, carriers of the BDNF Met/Met and 5-HTTLPR S/S genotypes experienced more distress than non-carriers. However, the pilgrimage environment seemed to mitigate some of the risk associated with these genotypes, and supports gene-environment interact [42, 43]. Polygenic risk exploratory analyses suggested that walkers and athletes had decreased genetic liability compared with control subjects, though further study using broader multi-omics approaches will be required to clarify mechanisms.

Integration with broader literature

In summary, these findings not only support a biopsychosocial perspective of how culture and spirituality shape psychological well-being through endocrine and genetic pathways; they also support a meaning-focused model of coping [44] and demonstrate the importance of incorporating spirituality into resilience theory. Collectively, these data indicate the importance of considering both the cultural/social context and biological factors (e.g., psychological processes like stress & anxiety, biological systems like genetics & HPA axis) while providing evidence that stress adaptation is due to the social/culturally contextual factors more than it is biological as seen from this integrated position.

Public health integration

These results have translational value in addition to theoretical implications. Spiritual walking is an inexpensive scalable and culturally relevant activity that can be incorporated into community-based mental health initiatives. From this angle it is conceivable that other meaningful group settings like military units educational cohorts or community service projects might display comparable psychobiological characteristics. The current study was not intended to test these comparisons directly though and future studies should specifically look into whether meaningful group activities outside of religious contexts have similar stress-reduction effects. In locations where there is a tradition of religious rituals, publicly funded health programs could use these pre-existing community cultural practices to improve their citizens’ ability to cope with stress and have good mental health. By incorporating these practices into public health policy, they can have greater access to them. They provide opportunities for resource-poor communities to offered holistic care, using a combination of biological, psychological, and cultural services; and augmenting traditional methods of providing care.

Limitations

It’s important to note that there are a number of limitations associated with this study. Firstly, the sample size (N = 300) is a combination of large samples compared to other studies of similar-sized cohorts; however, the small number of cells that equate to the combination of sex, age, and genotype has limited our ability to perform subgroup analysis, especially for such things as looking at differences in sex or looking at rarer genetic groupings, e.g., A/A, A/G, or G/G genotypes. Due to this, we cannot generalize findings to other sexes, age groups, or genetic groupings, and this limited our statistical power to detect gene-environmental interaction and lesser genes; therefore, future studies need to be larger and multicenter to verify and build on our results. The co-varying gender (sex) with the other covariates in analyses also means that we do not have sufficient power to conduct subgroup analyses using sex. Future investigations using a larger sample should continue to investigate possible biological sex differences in stress resilience, neuroendocrine regulation, and spirituality. The only candidate polymorphisms/ST genes included in the current analyses were BDNF Val66Met and 5-HTTLPR in addition to exploratory polygenic scores. While both exploratory polygenic scores and these two candidate genes have excellent theoretical support, the current findings underemphasize the complex nature of genetic contributions to stress and resilience because of the small number of genes examined. Additional transcriptomic research, epigenetic studies, or genome-wide association studies (GWAS) may help to better define the biological pathways related to the mechanisms of resilience in response to stressful events. With respect to genotyping, if functional studies to link genetic variations to neurobiologic processes had been conducted, the conclusions would have clearly been strengthened. Psychological outcomes were evaluated using only the self-report questionnaires, which are subject to bias sources, including social desirability, recall error, and the influence of the setting. While these outcomes were evaluated using validated Arabic versions of standardized measures, the lack of neuropsychological or clinician-rated measures limits the objectivity of the results. Future studies should utilize multimodal measures, such as electrical momentary assessment (EMA) behavioral tasks or neuroimaging measures of emotional regulation and stress response, to provide a broader understanding of psychological outcomes. In addition, this study’s findings must be regarded in light of its cultural and contextual specificity. In other cultural contexts, these characteristics might not apply to spiritual practices or secular walking interventions. Therefore, controlled intervention trials and comparative cross-cultural research are required to ascertain whether the noted psychobiological advantages apply to different environments and populations.

Conclusion

The present study indicates that walking for spiritual motives, such as a pilgrimage to the Arbaeen, is linked to lesser psychological stress, a more normal pattern of the HPA axis hormone activity, and a reduced genetic predisposition to stress-related disorders when compared with those who are physically active or even inactive.

By combining evidence from psychology, hormonal processes, and genetics within a culturally meaningful context, these results deepen the understanding of stress management in individuals. Besides, they point to the potential benefits of simple, spiritually guided methods for improving the mental health of a wider population.

Further research that applies cutting-edge methodologies, multi-level biological exploration, and brain imaging, etc., is required to determine the exact mechanisms of these effects and thus to deepen the understanding of the relationship.

Author contributions

H.M.K.A. conceived and designed the study. A.J.O. and D.K.A. coordinated data collection. H.S.A. processed biological samples. M.A.K. performed statistical analyses. All authors interpreted the results, revised the manuscript, and approved the final version.

Funding

This research received no specific grant from any funding agency, commercial or not-for-profit sectors.

Data availability

Upon publication, de, identified data, analysis scripts, and figure source files will be uploaded to the Open Science Framework (DOI will be provided). The corresponding author will consider reasonable requests for additional materials.

Code availability

All analysis code (R v4.2 scripts and SPSS syntax) will be uploaded to OSF together with the dataset.

Declarations

Ethics approval and consent to participate

This study was approved by the Institutional Review Board of Al-Furat Al-Awsat Technical University, Iraq (Approval No. ATU-ETH-2024-019). All research procedures were conducted in accordance with the ethical standards of the Institutional Review Board of Al-Furat Al-Awsat Technical University and adhered to the principles outlined in the Declaration of Helsinki (2013 revision), ensuring compliance throughout the study. Written informed consent was obtained from all participants prior to participation in the study.

Consent to publish

Not applicable. This study does not contain any individual person’s data in any form (including images, videos, or case details).

Reporting summary

Further information on research design is available in the Nature Research Reporting Summary linked to this article.

Competing interests

The authors declare no competing interests.