Symbiotic and phenotypic characteristics of thirty rhiobial isolates obtained from root nodules of two cowpea (Vigna unguiculata L. Walp) cultivars that grow in different sites of Fezzan (Southern part of Libya) were studied. Cultural characteristics and cross-nodulation with Arachis hypogega and Faidherbia albida showed that they were slow-growing rhizobia. Each isolate was found to coexist with non-symbiotic bacteria similar in their cultural characteristics to fast-growing rhizobia. All isolates formed symbiosis with the test plants, but different in their nitrogen-fixation efficiency. Numerical analysis of phenotypic characteristics showed that at boundary level of 70% average similarity, the isolates formed four distinguished groups and two isolates remained separate. Most isolates exhibited wide tolerance to acidity, alkalinity and extreme temperatures. They also resistant to some heavy metals such as mercury, copper, zinc, lead, cadmium and aluminum at low concentrations and antibiotics like polymyxin, colistin, bacitracin and nalidixic acid. Isolates displayed different response to salinity ranging from sensitive, which unable to grow in 1% NaCl to resistant and grow at 2% NaCl or above. Urea was hydrolyzed by most of them and carbohydrates utilizations were different. Sucrose and maltose were metabolized by most of the test isolates, whereas, monosaccharide and sugar alcohols were poorly utilized.