Physical Activity and Gut Microbiota in Children (PA-Microbiome Kids) (PA-Microbiome)

Effects of a 16-week Moderate-to-Vigorous Physical Activity Program on Physical Fitness and Gut Microbiota in Children: A Randomized Controlled Trial

This study evaluates the effect of a 16-week moderate-to-vigorous physical activity (MVPA) program on physical fitness and gut microbiota in healthy school-aged children. Participants will be randomly assigned to either an MVPA intervention group or an attention-control group. The primary outcome is the change in a composite physical fitness score. Secondary outcomes include changes in gut microbiota diversity and composition.

Study Overview

• Status: Completed
• Conditions: Healthy
• Intervention / Treatment: Behavioral: 16-week Structured MVPA Program; Behavioral: Attention Control Sedentary Activities

Detailed Description

This is a two-arm, parallel-group, randomized controlled trial with 1:1 allocation, conducted as an intervention extension of a previously published cross-sectional cohort study (Pan et al., Sci Rep 2025;15:21351; GSA-Human HRA010530). Healthy children aged 7-11 years at the time of intervention baseline (December 2025), who were 6-9 years old when they participated in the 2023 cross-sectional study in Weifang, Shandong, will be included.

In the 2023 baseline study, 120 children were sampled across four PA×PF quadrant groups defined by quartiles of physical activity (PA) and physical fitness (PF): P1 (low PA-low PF), P2 (high PA-low PF), P3 (low PA-high PF), and P4 (high PA-high PF). Of these, 113 provided valid baseline fecal samples and physical fitness data. These 113 participants were re-contacted and re-randomized 1:1 to MVPA or attention-control arms in December 2025. Randomization is stratified by baseline PA×PF quadrant (P1-P4) to ensure balanced allocation of baseline gut microbiome phenotypes across intervention and control arms. The reported enrollment of 113 refers to the number re-randomized at the intervention baseline (T1), not new recruits.

The intervention group will receive a 16-week structured MVPA program (3 sessions/week, 45-60 min/session). The control group will receive attention-matched sedentary activities (e.g., reading, crafts) and will be offered the MVPA program after the trial (waitlist). This is a PROBE-design trial: participants and intervention instructors are not blinded to group assignment, while outcome assessors (physical fitness tests) and laboratory/bioinformatics personnel (microbiota analysis) are blinded to allocation. The database uses codes (e.g., Group A/B) until primary analysis is complete. The primary analysis will follow the intention-to-treat (ITT) principle.

The study aims to (1) determine whether the MVPA intervention improves physical fitness and modulates the gut microbiota, and (2) determine whether baseline PA×PF quadrant membership and baseline abundance of key microbial taxa (Akkermansia, Faecalibacterium, Subdoligranulum, Bacteroides coprocola) modify the intervention response, thereby informing precision exercise prescription strategies.

Ethics and Oversight: This study operates under a collaborative framework in which the China Institute of Sport Science (CISS) serves as the primary institution responsible for ethical oversight and scientific integrity, while Aiyoudong Children and Youth Sports Health Research Institute serves as the implementing institution. Ethical approval was granted by the Ethics Committee of CISS (approval number: CISSLA20230110), covering both the 2023 baseline cross-sectional study and the current intervention extension. Aiyoudong Institute, which does not maintain an independent ethics committee, conducts the intervention under CISS oversight through a formal inter-institutional data use and oversight agreement. Written informed consent was obtained from legal guardians and assent from child participants at both baseline (2023) and intervention enrollment (December 2025).

Exploratory aim: to characterise the co-variation between intervention-induced changes in physical activity, physical fitness, gut microbiota, and psychosocial and behavioural measures, with the gut microbiota as the central node. These analyses are exploratory and hypothesis-generating, and no specific direction of change in the psychosocial or behavioural measures is presumed.

• Study Type: Interventional
• Enrollment (Actual): 113
• Phase: Not Applicable

Contacts and Locations

Study Locations

• China
  • Shandong
    • Weifang, Shandong, China, 261000
      • Aiyoudong Children and Youth Sports Health Research Institute

Participation Criteria

Eligibility Criteria

• Ages Eligible for Study: Child
• Accepts Healthy Volunteers: Yes

Description

Inclusion Criteria:

• Participants from a previous cross-sectional study cohort conducted in 2023 (Pan et al., Sci Rep 2025;15:21351), at which time they were 6-9 years old.
• Able to complete the 16-week follow-up.
• Guardian provides written informed consent, and the child provides assent.
• Medically cleared by a physician or school nurse to participate in moderate-intensity physical activity (no contraindications).
• No antibiotic use in the 4 weeks prior to baseline.
• No fever or acute gastroenteritis in the 2 weeks prior to baseline.
• Able to provide stool samples as required.

Exclusion Criteria:

• Diagnosed with cardiopulmonary diseases, epilepsy, severe asthma, severe musculoskeletal disorders, or congenital heart disease.

  • Recent acute exacerbation of chronic gastrointestinal diseases.
  • Planning to follow a special diet (e.g., ketogenic, vegetarian) or requiring long-term prescription of antibiotics/probiotics.
  • Planning to be absent for >20% of the intervention period due to travel or other commitments.
  • Enrolled in other high-intensity sports training programs that cannot be cancelled.
  • Developmental or behavioral issues that would impede cooperation, as assessed by teachers and parents.

Study Plan

How is the study designed?

Design Details

• Primary Purpose: Prevention
• Allocation: Randomized
• Interventional Model: Parallel Assignment
• Masking: Single

Number of Arms

2

Arms and Interventions

Participant Group / Arm	Intervention / Treatment
Experimental: MVPA Program; Participants will engage in a 16-week structured moderate-to-vigorous physical activity (MVPA) program, 3 times per week, for 45-60 minutes per session. Sessions include warm-up, aerobic games, bodyweight strength circuits, and cool-down, supervised by trained instructors.	Behavioral: 16-week Structured MVPA Program; A 16-week structured moderate-to-vigorous physical activity program.
Other: Attention Control + Waitlist; Participants engage in time-matched sedentary activities such as reading, science experiments, or crafts, three times per week, and are instructed to maintain their usual physical activity levels. After the 16-week trial period, they are offered a comparable MVPA program (waitlist).	Behavioral: Attention Control Sedentary Activities; Time-matched sedentary activities such as reading, science experiments, or crafts.

What is the study measuring?

Primary Outcome Measures

Outcome Measure	Measure Description	Time Frame
Change in Composite Physical Fitness Z-score	Change from Baseline to 16 weeks in a composite physical fitness z-score. The score is calculated based on the age-appropriate items and weighting from the "National Student Physical Health Standard (2014 Revision)". The scale is a standardized z-score with a mean of 0 and a standard deviation of 1, based on a reference population. There are no theoretical upper or lower limits, but values typically range from -3 to +3. A higher score indicates better overall physical fitness.	Baseline and 16 weeks

Secondary Outcome Measures

Outcome Measure	Measure Description	Time Frame
Change in Gut Microbiota Alpha Diversity	Change from Baseline to 16 weeks in alpha diversity indices (Shannon and Chao1) of the gut microbiota, assessed via 16S rRNA gene sequencing. The Shannon index has a minimum value of 0, with no theoretical maximum. The Chao1 index estimates species richness with a minimum equal to the number of observed species, and also has no theoretical maximum. For both indices, higher values indicate greater diversity and are generally considered to reflect a healthier gut microbiome.	Baseline and 16 weeks
Difference in Gut Microbiota Beta Diversity	Difference in gut microbiota community structure (beta diversity) between the MVPA and control groups. This will be assessed using a Permutational Multivariate Analysis of Variance (PERMANOVA) on Bray-Curtis and UniFrac distance matrices. The outcome will be reported as an R-squared value, representing the percentage of variance in community structure explained by group assignment, and a p-value. A larger R-squared value and a smaller p-value would indicate a more significant difference between the groups.	Baseline and 16 weeks
Change in Body Mass Index (BMI) Z-score	Change from Baseline to 16 weeks in age- and sex-specific BMI z-score. This is a standardized score with a mean of 0, based on a reference population, and has no theoretical upper or lower limits. A higher score indicates a higher BMI relative to peers. In the context of this study focusing on healthy weight, an increase in the z-score is generally considered a worse outcome.	Baseline and 16 weeks
Heterogeneity of Treatment Effect Across Baseline PA×PF Quadrants	Heterogeneity of treatment effect (HTE) of the MVPA intervention on primary and secondary outcomes across the four pre-specified baseline phenotype groups (P1: low PA-low PF; P2: high PA-low PF; P3: low PA-high PF; P4: high PA-high PF) defined in the 2023 cross-sectional study. HTE will be assessed by including a Treatment × Baseline Quadrant interaction term in mixed-effects models. A statistically significant interaction (p < 0.05) would indicate that the intervention effect differs across baseline microbiome phenotypes. Effect sizes will be reported separately for each quadrant with 95% confidence intervals.	Baseline and 16 weeks
Baseline Microbial Taxa as Effect Modifiers of Intervention Response	Pre-specified analysis of whether baseline relative abundance (continuous, log-transformed) of Akkermansia, Faecalibacterium, Subdoligranulum, and Bacteroides coprocola modifies the MVPA intervention effect on composite physical fitness z-score and alpha diversity. Interaction terms (Treatment × Baseline Taxon Abundance) will be tested in mixed-effects models.	Baseline and 16 weeks

Other Outcome Measures

Outcome Measure	Measure Description	Time Frame
Change in Body Fat Percentage	Change from baseline to 16 weeks in body fat percentage. Higher values indicate greater adiposity.	Baseline and 16 weeks
Change in Waist Circumference and Waist-to-Hip Ratio	Change from baseline to 16 weeks in waist circumference (cm) and waist-to-hip ratio. Higher values indicate greater central adiposity.	Baseline and 16 weeks
Change in Resting Heart Rate	Change from baseline to 16 weeks in resting heart rate (beats per minute) measured after seated rest. Lower values generally indicate better cardiovascular fitness.	Baseline and 16 weeks
Change in Blood Pressure	Change from baseline to 16 weeks in resting systolic and diastolic blood pressure (mmHg).	Baseline and 16 weeks
Change in Vital Capacity	Change from baseline to 16 weeks in vital capacity (mL) measured by spirometry. Higher values indicate greater lung function.	Baseline and 16 weeks
Change in Stool Form and Defecation Frequency	Change from baseline to 16 weeks in stool form assessed by the Bristol Stool Form Scale (types 1 to 7) and in self-reported weekly defecation frequency.	Baseline and 16 weeks
Change in Resilience (CD-RISC-10)	Change from baseline to 16 weeks in resilience measured by the 10-item Connor-Davidson Resilience Scale. Scores range from 0 to 40; higher scores indicate greater resilience.	Baseline and 16 weeks
Change in Well-Being (WHO-5)	Change from baseline to 16 weeks in subjective well-being measured by the WHO-5 Well-Being Index. Raw scores (0 to 25) are transformed to a 0 to 100 scale; higher scores indicate better well-being.	Baseline and 16 weeks
Change in Anxiety Symptoms (SCARED)	Change from baseline to 16 weeks in anxiety symptoms measured by the child self-report Screen for Child Anxiety Related Emotional Disorders. Higher scores indicate more anxiety symptoms.	Baseline and 16 weeks
Change in Depressive Symptoms (CES-D)	Change from baseline to 16 weeks in depressive symptoms measured by the Center for Epidemiologic Studies Depression Scale. Higher scores indicate more depressive symptoms.	Baseline and 16 weeks
Change in Strengths and Difficulties (SDQ)	Change from baseline to 16 weeks measured by the Strengths and Difficulties Questionnaire. Higher total difficulties scores indicate more difficulties; higher prosocial subscale scores indicate more prosocial behaviour.	Baseline and 16 weeks
Change in Coping Style (SCSQ)	Change from baseline to 16 weeks in coping style measured by the Simplified Coping Style Questionnaire, reported as positive and negative coping subscale scores.	Baseline and 16 weeks
Change in Self-Concept	Change from baseline to 16 weeks in self-concept measured by an age-appropriate self-concept questionnaire for younger children. Higher scores indicate more positive self-concept.	Baseline and 16 weeks
Change in Emotion Regulation (ERQ-CA)	Change from baseline to 16 weeks in emotion regulation measured by the Emotion Regulation Questionnaire for Children and Adolescents, reported as cognitive reappraisal and expressive suppression subscale scores.	Baseline and 16 weeks
Change in Objective Sleep	Change from baseline to 16 weeks in accelerometer-derived sleep, including total sleep time and sleep efficiency.	Baseline and 16 weeks
Change in Physical Literacy	Change from baseline to 16 weeks in physical literacy measured by a physical literacy questionnaire. Higher scores indicate greater physical literacy.	Baseline and 16 weeks
Change in Exercise Self-Efficacy	Change from baseline to 16 weeks in exercise self-efficacy measured by an 18-item exercise self-efficacy questionnaire. Higher scores indicate greater self-efficacy.	Baseline and 16 weeks
Change in Dietary Self-Efficacy	Change from baseline to 16 weeks in dietary self-efficacy measured by a 16-item dietary self-efficacy questionnaire. Higher scores indicate greater self-efficacy.	Baseline and 16 weeks
Change in Physical Activity Enjoyment (PACES)	Change from baseline to 16 weeks in physical activity enjoyment measured by the Physical Activity Enjoyment Scale. Higher scores indicate greater enjoyment.	Baseline and 16 weeks
Change in Eating Behaviour (CEBQ)	Change from baseline to 16 weeks in eating behaviour measured by the Children's Eating Behaviour Questionnaire, reported by subscale.	Baseline and 16 weeks
Change in Executive Function and Attention		Baseline and 16 weeks
Change in Objective Physical Activity and Sedentary Time	Change from baseline to 16 weeks in accelerometer-derived moderate-to-vigorous physical activity (minutes per day) and sedentary time (minutes per day), used to verify intervention-induced change in activity.	Baseline and 16 weeks

Collaborators and Investigators

• Sponsor: Aiyoudong Children and Youth Sports Health Research Institute

Study record dates

Study Major Dates

• Study Start (Actual): December 31, 2025
• Primary Completion (Actual): April 22, 2026
• Study Completion (Actual): May 31, 2026

Study Registration Dates

• First Submitted: December 3, 2025
• First Submitted That Met QC Criteria: December 20, 2025
• First Posted (Actual): December 30, 2025

Study Record Updates

• Last Update Posted (Actual): June 3, 2026
• Last Update Submitted That Met QC Criteria: May 31, 2026
• Last Verified: May 1, 2026

More Information

Terms related to this study

• Keywords: Physical Activity; Microbiome; Children; Health Promotion; Gut Microbiota; Physical Fitness
• Additional Relevant MeSH Terms: Behavior; Motor Activity
• Other Study ID Numbers: PA-Microbiome-Kids-2025; 2244 (Other Grant/Funding Number: China Institute of Sport Science)

Plan for Individual participant data (IPD)

• Plan to Share Individual Participant Data (IPD)?: YES
• IPD Plan Description: De-identified participant-level data including 16S rRNA sequencing reads, alpha/beta diversity indices, taxonomic relative abundance tables, physical fitness test results, and physical activity assessment scores will be shared. Sequencing reads from the 2023 baseline have been deposited in the Genome Sequence Archive (GSA-Human: HRA010530). Intervention-phase sequencing data will be deposited in the same repository upon publication of the primary results manuscript.
• IPD Sharing Time Frame: Data will be available starting 12 months after publication of the primary outcomes manuscript, with no end date.
• IPD Sharing Access Criteria: Data will be publicly accessible via the Genome Sequence Archive (https://ngdc.cncb.ac.cn/gsa-human) for sequencing data. Phenotypic data will be available upon reasonable request to the corresponding author, subject to a data use agreement and approval by the CISS Ethics Committee.
• IPD Sharing Supporting Information Type: STUDY_PROTOCOL; SAP; ICF; ANALYTIC_CODE; CSR

Drug and device information, study documents

• Studies a U.S. FDA-regulated drug product: No
• Studies a U.S. FDA-regulated device product: No