Maintenance of Intracellular pH and Acid Tolerance in Rhizobium meliloti

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Abstract
The development and function of the Rhizobium meliloti-Medicago sp. symbiosis are sensitive to soil acidity. Physiological criteria that can be measured in culture which serve to predict acid tolerance in soil would be valuable. The intracellular pH of R. meliloti was measured using either radioactively labeled weak acids (5,5-dimethyloxazolidine-2,4-dione and butyric acid) or pH-sensitive fluorescent compounds; both methods gave similar values. Six acid-tolerant strains (WSM419, WSM533, WSM539, WSM540, WSM852, and WSM870) maintained an alkaline intracellular pH when the external pH was between 5.6 and 7.2. In contrast, two Australian commercial inoculant strains (CC169 and U45) and four acid-sensitive strains from alkaline soils in Iraq (WSM244, WSM301, WSM365, and WSM367) maintained an alkaline intracellular pH when the external pH was ≥6.5, but had intracellular pH values of ≤6.8 when the external pH was ≤6.0. Four transposon Tn5-induced mutants of acid-tolerant strain WSM419, impaired in their ability to grow at pH 5.6, showed limited control over the intracellular pH. The ability to generate a large pH gradient under acid conditions may be a better indicator of acid tolerance in R. meliloti under field conditions than is growth on acidic agar plates.