Nondividing, Bacteroid-Like Rhizobium trifolii: In Vitro Induction Via Nutrient Enrichment

Rhizobium trifolii 0403 maintained in exponential phase via periodic dilution doubled in 210 min in mannitol-salts medium and doubled in 244 min in glycerolsalts. In both media, cell number and optical density increased in parallel. When exponentially growing cells in either medium were supplemented with a mixture of glucose, Casamino Acids, succinate, and yeast extract, optical density continued to increase but within less than the time required for one doubling, division ceased. The increase in optical density coupled with division cessation resulted in the formation of large, pleomorphic, nondividing cells. Large cells apparently increased in size as a result of swelling only at regions of most recent cell envelope synthesis. Greater than 95% of the cells in a population swelled, and commitment to swelling occurred within two doubling time equivalents. Swollen cells eventually reached a characteristic maximum size and exhibited osmotic fragility.