Imaging of total calcium in urediospore germlings of Uromyces by ion microscopy

Calcium has been implicated in growth and appressorium formation of urediospore germlings of the bean rust fungus, Uromyces appendiculatus. Using ion microscopy, a mass spectrometry-based imaging technique, intracellular stores of calcium were analyzed by direct imaging of total calcium in frozen freeze-dried germlings. Calcium concentration was calculated by ratioing and spatially registering (40)Ca to (12)C signals. Intracellular distributions of total potassium, sodium, magnesium, and carbon were similarly imaged in the same germlings for a direct comparison of their localizations to total calcium. Calcium was remarkably heterogeneous with highest concentrations (2 to 10 mM) in the mid-region of the germling between the nuclei and the apex. A similar distribution of Ca(2+) (assessed using Fluo-3) was also noted sequestered in organelles in live germlings. Distributions of remaining elements (K, Na, Mg, and C) were mostly homogeneous throughout the cytoplasm and nuclei of the fungal cell. The K/Na ratio ranged from 17 to 31.