This study evaluated the activity of rhizobia isolates inoculated in large (18 mm) and small (11 mm) seeds on lima bean growth, nodulation and N fixation. Selected rhizobia isolates were compared with a reference strain CIAT899 and two controls without inoculation. Large seeds contributed for highest plant growth, nodulation and N fixation than small seeds. The isolates UFPI-59, UFPI-18 and UFPI-38 promoted the highest values of shoot and root dry weight, respectively. The isolates UFPI-32 promoted the highest values of nodule number, while UFPI-59 promoted the highest values of nodule dry weight. The isolates UFPI-38 and UFPI-59 promoted the highest accumulation of N. This study showed that seed size really influences lima bean growth, nodulation and BNF. Considering rhizobia isolates, UFPI-59, UFPI-38, and UFPI-18 contributed for plant growth, promoted better nodulation and effectiveness on biological N fixation.