At deep-sea hydrothermal vents, sulfur oxidation and iron oxidation are of the highest importance to microbial metabolisms, which are thought to contribute mainly in chemolithoautotrophic groups. In this study, 17 mixotrophic neutrophilic thiosulfate- and iron-oxidizing bacteria were isolated from hydrothermal fields on the Carlsberg Ridge in the Indian Ocean, nine to the γ-proteobacteria (Halomonas (4), Pseudomonas (2), Marinobacter (2), and Rheinheimera (1)), seven to the α-proteobacteria (Thalassospira, Qipengyuania, Salipiger, Seohaeicola, Martelella, Citromicrobium, and Aurantimonas), and one to the Actinobacteria (Agromyces), as determined by their 16S rRNA and genome sequences. The physiological characterization of these isolates revealed wide versatility in electron donors (Fe(II) and Mn(II), or thiosulfate) and a variety of lifestyles as lithotrophic or heterotrophic, microaerobic, or anaerobic. As a representative strain, Pseudomonas sp. IOP_13 showed its autotrophic gowth from 105 cells/ml to 107 cells/ml;carbon dioxide fixation capacity with the δ13CVPDB in the biomass increased from −27.42‰ to 3460.06‰; the thiosulfate-oxidizing ability with produced SO42− increased from 60 mg/L to 287 mg/L; and the iron-oxidizing ability with Fe(II) decreased from 10 mM to 5.2 mM. In addition, iron-oxide crust formed outside the cells. Gene coding for energy metabolism involved in possible iron, manganese, and sulfur oxidation, and denitrification was identified by their genome analysis. This study sheds light on the function of the mixotrophic microbial community in the iron/manganese/sulfur cycles and the carbon fixation of the hydrothermal fields.