Abstract

Background/Objectives: Rearrangement of the gut microbiota toward a symbiotic profile may be influenced by physical activity and diet in both healthy and obese individuals. This study aimed to characterize gut microbiota using next-generation sequencing (NGS) and to evaluate the effect of a 6-week Nordic Walking (NW) program combined with Time-Restricted Eating (TRE; 10 h eating window) in individuals with obesity. Methods: The study included healthy controls (C; n = 10; 64.7 ± 6.7 years) and individuals with obesity (A; n = 10; 60.0 ± 4.5 years). The intervention consisted of three moderate-intensity NW sessions per week, individually adapted to participants’ capacity. Gut microbiota was analyzed using nanopore 16S rRNA sequencing (V3–V9 regions). Results: Baseline microbial composition differed significantly between obese and control groups, with Bray–Curtis dissimilarity ranging from 49.98% (phylum) to 65.97% (species) (p ≤ 0.02). After intervention, within-group dissimilarity in the obese cohort (A vs. B) decreased to 25.24–51.86% but was not significant (p = 0.88–1.00). Post-intervention comparisons (B vs. C) still showed significant differences at higher taxonomic levels, including class (44.10%, p = 0.015), order (31.62%, p = 0.022), family (50.48%, p = 0.011), genus (58.09%, p = 0.005), and species (63.47%, p = 0.0003). Alpha diversity showed no significant differences at species and genus levels, but significant group effects were observed at higher ranks, including order (Shannon p = 0.01; Simpson p = 0.01), class (Simpson p = 0.02), and phylum (Shannon p = 0.02). Conclusions: The NW + TRE intervention was associated with partial normalization of gut microbiota structure and reduced microbial dissimilarity, suggesting a shift toward a more symbiotic profile; however, differences compared with controls persisted, indicating incomplete convergence after 6 weeks.

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