Background: The main aim of this study was to examine the effect of chemical mutagens on enhancement of extreme acid and alkaline condition of Vicia faba nodulating rhizobial isolates from Hararghe highlands in Ethiopia. Methods: A total of 50 wild rhizobial isolates were isolated from different regions of Hararghe highlands soils by using pot experiment. All selected isolates showed significantly (P ≤ 0.05) higher nodule dry weight (NDW) than the positive control. Above the 50 isolates only 10 highly performed isolates were subjected to chemical mutation. After the mutagenic treatment we selected only eight survived isolates, three isolates from sodium azide and five isolates form hydroxyl amine hydrochloride. The selected mutagen was subjected to study their symbiotic effectiveness. Results: The sodium azide treated muatgens showed higher performance in terms of symbiotic effectiveness compared with hydroxyl amine hydrochloride. Chemical mutagen hydroxyl amine hydrochloride showed the acidic tolerance (pH 4) only three mutant isolates such as HUFBR12M4, HUFBR39M6 and HUFBR18M8. In case of sodium azide, it did not show any mutant isolates under acidic condition. In the meantime alkaline tolerance mutant isolates (HUFBR50M1, HUFBR31M2, HUFBR12M3, HUFBR12M4, HUFBR23M5, HUFBR39M6, HUFBR50M7, and HUFBR18M8) were observed in both the mutagenic chemicals (sodium azide and hydroxyl amine hydrochloride). All the isolates showed the pH range 10.5 to 12. Conclusion: Finally, it was concluded that mutagenic chemical sodium azide is suitable for inducing chemical mutation to enhance acidic tolerance of mutant rhizobium in Faba bean and both the mutagenic chemicals (sodium azide and hydroxyl amine hydrochloride) are suitable for alkaline tolerance of mutant rhizobium in Faba bean.