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Calorie restriction (CR) extends lifespan and retards age-related chronic diseases in most species. There is growing evidence that the gut microbiota has a pivotal role in host health and age-related pathological conditions. Yet, it is still unclear how CR and the gut microbiota are related to healthy aging. Here, we report findings from a small longitudinal study of male C57BL/6 mice maintained on either <i>ad libitum</i> or mild (15%) CR diets from 21 months of age and tracked until natural death. We demonstrate that CR results in a significantly reduced rate of increase in the frailty index (FI), a well-established indicator of aging. We observed significant alterations in diversity, as well as compositional patterns of the mouse gut microbiota during the aging process. Interrogating the FI-related microbial features using machine learning techniques, we show that gut microbial signatures from 21-month-old mice can predict the healthy aging of 30-month-old mice with reasonable accuracy. This study deepens our understanding of the links between CR, gut microbiota, and frailty in the aging process of mice.