Aging is a time-dependent complex biological process of organisms with gradual deterioration of the anatomical and physiological functions. The role of gut microbiota is inevitable in the aging process. Probiotic interventions improve gut homeostasis and support healthy aging by enhancing beneficial species and microbial biodiversity in older adults. The present preliminary clinical trial delves into the impact of an 8-week Lactobacillus rhamnosus intervention (10 × 10⁹ CFU per day) on the glycaemic index, lipid profile, and microbiome of elderly subjects. Body weight, body fat, fasting blood glucose, total cholesterol, triglyceride, high-density lipoprotein, and low-density lipoprotein (LDL) are assessed at baseline (Week 0) and after treatment (Week 8) in placebo and probiotic groups. Gaussian regression analysis highlights a significant improvement in LDL cholesterol in the probiotic group (p = 0.045). Microbiome analysis reveals numeric changes in taxonomic abundance at various levels. At the phylum level, Proteobacteria increases its relative frequency (RF) from 14.79 ± 5.58 at baseline to 23.46 ± 8.02 at 8 weeks, though statistically insignificant (p = 0.100). Compared to the placebo group, probiotic supplementations significantly increased the proteobacteria abundance. Genus-level analysis indicates changes in the abundance of several microbes, including Escherichia-Shigella, Akkermansia, and Bacteroides, but only Butyricimonas showed a statistically significant level of reduction in its abundance. Probiotic supplementations significantly altered the Escherichia-Shigella and Sutterella abundance compared to the placebo group. At the species level, Bacteroides vulgatus substantially increases after probiotic treatment (p = 0.021). Alpha and beta diversity assessments depict subtle shifts in microbial composition. The study has limitations, including a small sample size, short study duration, single-strain probiotic use, and lack of long-term follow-up. Despite these constraints, the study provides valuable preliminary insights into the multifaceted impact of L. rhamnosus on elderly subjects. Further detailed studies are required to define the beneficial effect of L. rhamnosus on the health status of elderly subjects.