Mesorhizobium ciceri is a pivotal bacterium in the nitrogen fixation process of chickpea (Cicer arietinum), playing a crucial role in enhancing soil fertility and crop productivity. This comprehensive review delves into the biochemical profiling and characterization of M. ciceri isolates from chickpea root nodules. It details the methodologies employed for isolating and identifying these bacterial isolates, encompassing morphological, biochemical, and molecular techniques. The review highlights various biochemical assays, including carbon source utilization, enzyme activity, and antibiotic resistance profiles, alongside genetic and phenotypic characterizations through 16S rRNA sequencing, multilocus sequence typing, and genomic fingerprinting and explores the implications of these findings for sustainable agriculture, focusing on biofertilizer development, soil health, and improved crop management. By providing an in-depth understanding of M. ciceri‘s role in chickpea symbiosis, this review aims to enhance strategies for sustainable agricultural practices and legume crop productivity.