Rhizobia are important bacteria, playing a vital role in atmospheric nitrogen fixation. In addition, it increases soil fertility, enhancing crop production and preserving the soil from pollution resulting from inorganic fertilizers’ use. From 81 bacteria samples, nine isolates segregated from nine types of leguminous crops are economically valuable for cultivation within nine different locations in Nineveh Governorate, Iraq. The isolates underwent initial diagnosis by phenotypical and cultural characteristics and host specialization tests. Molecular characterization also ran on the selected bacterial isolates. Performing the 16S rRNA region amplification applied forward primer 27F and a reverse primer 1541R using the polymerase chain reaction (PCR) technique. The results of restriction fragment length polymorphism (RFLP-PCR) employing three endonuclease enzymes showed that EcoRI had similar sequences among the isolates under study but significantly different for MspI and HaeIII. Based on the analysis of results using the statistical program MVSP version 3.22, the degree of similarity divided the isolates into three groups. Results revealed that there were genetic differences between the isolates and the suppressing enzymes, providing more accurate results in recognizing the similarity among the bacterial isolates as a result of the difference in the location and environmental conditions from which the bacteria were isolated, in addition to the different agricultural processes used for those regions.

Keywords: Leguminous crops, rhizobium, culture and biochemical tests, endonuclease enzymes, RFLP-PCR, 16S Rrna

Key findings: The use of 16S rRNA-RFLP technology enables the division of isolated bacteria by studying their genetic polymorphisms treated with restriction enzymes. Detecting three distinct groups from three different sections in Mosul City, Iraq, was possible. The differences were due to the influence of the environments and plants from which the bacteria were isolated and were evident using RFLP. It was possible to determine the phylogenetic tree that shows the degree of genetic affinity between the bacterial isolates.