Death of root-nodule bacteria on drying

Abstract

A study has been made of the survival of Rhizobium trifolii, suspended in simple media and applied to glass beads and seeds of subterranean clover, and held in atmospheres of controlled relative humidity (R.H.) within the range 0–100%, at 26° and at atmospheric pressure. Under drying conditions, two distinct phases could be recognized: a period of approximately 24 hr in which the main loss of water occurred and in which there was rapid death, and a storage period in which the death rate was reduced. Cells of Rh. trifolii lost viability rapidly when suspended in glass-distilled water and spread on glass beads, under both drying (0 and 20% R.H.) and non-drying (100% R.H.) conditions. Suspension in 9% maltose, though without effect on rate of water loss, reduced the death rate considerably during drying and storage at low R.H., and permitted multiplication at 100% R.H. As a protectant during drying and storage, maltose was markedly superior to yeast extract, sorbitol, and five other sugars, including its ß-isomer, cellobiose. These materials were, however, superior to water alone, in contrast to sodium chloride and Ringer's solution (equiosmolar with maltose), which gave no protection. Low maltose concentrations (0.9 and 3.6%) were inferior to higher concentrations (9, 18, and 27%). Seeds of subterranean clover and a seed extract tested on beads increased the early death rate. This acceleration of death was not affected by maltose and was most marked at 100% R.H. and during the first few hours of drying (0% R.H.), when water was still abundant. Gum arabic gave some protection against this toxic action, as well as being slightly more protective than maltose on glass beads. The death rate during drying of cells in water appeared to be inversely related to the concentration of inoculum. Maltose removed any such dependence on inoculum size. Under conditions where survival during storage was measurable (maltose and glass beads), 60% R.H. was inferior to 0 and 20%.