Forage legumes such as Leucaena leucocephala, besides being a rich source of protein for animal feed, its inclusion in silvopastoral systems provides fundamental components to improve soil properties. These plants have the ability to establish symbiotic association with the genus Rhizobium sp. and fix biologically atmospheric nitrogen favoring growth and development, being reflected in production increases. The objective of this research was to evaluate the capacity of native isolates of Rhizobium sp. and a commercial strain in biological nitrogen fixation (BNF) in L. leucocephala plants. Two separate bioassays to evaluate the efficiency of the FBN of the native isolates and the commercial strain under greenhouse conditions, followed by a randomized complete block design with 7 x 2 factorial arrangement, seven treatments, two factors: native isolates and commercial strains and bacterial concentrations (106 and 108 cells.ml-1), with three repetitions and five experimental units. The first bioassay was carried out with seeds, determining the percentage of germination, the length and thickness of the stem, number of leaves, dry weight of the aerial part, number of nodule/plant and the percentage of nitrogen accumulated in the area part of the plant. In the second bioassay with 30-day-old seedlings, the same variables of the first bioassay were determined, with the exception of the percentage of germination. In the first bioassay, a greater stimulation was found in the germination of L. leucocephala seeds at a concentration of 108 cells.mL-1. Regarding stem length and thickness, accumulated dry matter, leaf  development and nitrogen accumulation, better results were found in the treatments with the native isolates L27, L36 and L38 in a concentration of 106 cells.mL-1 above the commercial strain in concentration of 108 cells.mL-1. The native isolates of Rhizobium sp. exert a positive effect on the FBN and the germination of plants of L. leucocephala, which will allow to conduct future field studies that allow to potentiate the culture of L. leucocephala and silvopastoral systems for bovine feeding.