Abstract
The gut microbiota forms early in life and undergoes dynamic changes that are essential for host health and development. Although body condition is a key fitness-related trait and predictor of viability in many animal species, its association with gut microbiota, especially during early life stages in wild populations, remains poorly understood. In this study, we collected fecal samples and used 16S rRNA gene sequencing to investigate temporal shifts in gut microbiota of Japanese tits (Parus minor) across nestling stages (days 3, 6, 10, and 14 post-hatching, denoted as D3, D6, D10, and D14, respectively; n = 70, repeatedly sampled) and in adults (n = 25), and examined their association with nestling body condition. The results showed that nestlings harbored distinct microbial communities compared to adults, with the latter exhibiting higher relative abundances of Bacteroidota and Verrucomicrobiota (LMMs, p < 0.001 for both). LEfSe analysis identified Actinobacteriota as a biomarker for D14 nestlings and Bacteroidota for adults. Alpha diversity decreased from D3 to D14, with adults showing higher diversity than late-stage nestlings (D10-D14), but comparable diversity to early-stage nestlings (D3-D6). Beta diversity revealed clear separation between nestlings and adults, and within nestlings, early stages (D3-D6) differed significantly from D14. Nest effects also contributed to microbial structure. Time-lagged analyses showed that Firmicutes abundance at D6 positively predicted scaled mass index (SMI) at D10 (p = 0.002), whereas Proteobacteria negatively predicted SMI (p = 0.006). Overall, these findings highlight dynamic, stage-specific shifts in the gut microbiota of Japanese tits and suggest that microbial succession may influence nestling growth and physiological adaptation.