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Studies on gut microbiomes of different animals have provided many ecological and evolutionary information showing a strict link with health and disease conditions. Despite that, few studies concerned herbivorous reptiles. The aim of this study was to characterize the faecal microbial community of Aldabrachelys gigantea, herbivorous giant tortoises, living in two different environmental conditions: at Seychelles and at Parco Natura Viva Zoological Park (PNV) in Verona (Italy). Individual fecal samples were collected from: 17 giant tortoises located in Curieuse Island (CI), 5 in Botanical Garden (BG) at Mahé in the Seychelles and 11 in PNV. Samples were collected in RNA later for microbiota NGS analysis and in agar slant to obtain isolates. Bacterial DNA was extracted using FastDNA SPIN Kit for Soil (MP Biomedicals) and the V3-V4 regions of the 16S rRNA gene were sequenced using the Illumina MisSeq platform. Amplification of good quality of DNA have been obtained from 17 samples (6 (CI), 3 (BG) and 8 (PNV) subjects). Bioinformatic analysis was carried out using the DADA2 pipeline and the Silva database as reference for taxonomy assignment was used. Alpha (Shannon, InvSimpson and Chao1) and beta (Bray Curtis distance) diversity indices were calculated and analysed using ANOVA and PERMANOVA model respectively. A total of 25 Phyla (42 % Bacteroidetes, 32% Firmicutes), 167 Families (14% Ruminococcaceae, 14% Rikenellaceae) and 310 Genera (7% Treponema, 6% Rikenellaceae_RC9_gut_group) were identified among samples. A higher microbial diversity within a sample (alpha indices) was observed in the BG compared to the PNV (Shannon: 5.39 vs 4.43; InvSimpson: 80.7 vs 25; Chao1: 584 vs 377 p ≤ 0.05). For the beta diversity, the microbiological composition of the samples was different between the three locations (p =0.001). Overall, the present study suggest that different environmental conditions can drive a shift of microbial profile of Aldabrachelys gigantea, which can be attributed to different diet or environmental conditions. Further investigations are needed.