Rhizobium japonicum derivatives differing in nitrogen-fixing efficiency and carbohydrate utilization

Area of Science:

• Microbiology
• Plant-Bacterial Interactions
• Biochemistry

Background:

• Rhizobium japonicum 110 derivatives were isolated exhibiting distinct colony morphologies.
• These derivatives nodulate Lee soybeans and display varying acetylene-reducing activity, indicating differences in nitrogen fixation efficiency.
• Standard culture techniques failed to isolate pure clones due to cell adherence, leading to mixed colonies.

Purpose of the Study:

• To characterize and differentiate four morphologically distinct derivatives of Rhizobium japonicum 110.
• To investigate the metabolic basis for observed differences in colony morphology and growth characteristics.
• To assess the genetic relatedness and nitrogen fixation efficiency of these Rhizobium derivatives.

Main Methods:

• Isolation of Rhizobium japonicum 110 derivatives based on colony morphology on yeast extract-mannitol-HM salts medium.
• Addition of Tween 40 detergent to facilitate the isolation of pure clones.
• Metabolic analysis including utilization of p-mannitol and D-arabitol for growth.
• Growth rate studies on various carbohydrates (glucose, fructose, gluconate, arabinose, glycerol, mannose).
• Antibiotic sensitivity testing, deoxyribonucleic acid (DNA) synthesis inhibitor response, and bacteriophage susceptibility assays.
• DNA analysis including buoyant density in CsCl gradients, thermal denaturation, and preliminary DNA/DNA hybridization.

Main Results:

• Two clones (L1-110 and L2-110) showed 5- to 10-fold lower acetylene-reducing activity compared to I-110 and S-110.
• L1-110 and L2-110 utilized p-mannitol and D-arabitol for growth, while I-110 and S-110 did not.
• Growth rate differences were observed on glucose and fructose between clones I-110 and L1-110.
• Clones I-110 and L1-110 exhibited identical responses to antibiotics and DNA synthesis inhibitors, and similar bacteriophage susceptibility.
• DNA analysis indicated identical base composition and high sequence homology between I-110 and L1-110.

Conclusions:

• Morphological differences in Rhizobium japonicum 110 derivatives are linked to distinct metabolic capabilities, particularly in carbohydrate utilization.
• Variations in p-mannitol and D-arabitol metabolism correlate with differences in nitrogen fixation efficiency.
• Despite metabolic and growth rate variations, clones I-110 and L1-110 are genetically highly similar, suggesting they represent distinct variants within the same strain.