Confocal Laser Scanning Microscopy of Environmental Samples

Enormous technical advances in imaging and data acquisition techniques, combined with a continuing increased scope for fluorescence labeling of specific constituents of living organisms, have brought about a revolution in approaches to biological problems. Whereas spatial organisation at an ultrastructural level would have seemed definitely the province of electron microscopy just a few years ago, now the new methods of light microscopy can offer not only a complementary approach, but can achieve more, especially in terms of noninvasive and real-time measurements. Confocal laser scanning microscopy (CLSM) is the most revolutionary development in optical microscopy since the early sevent eenth century. From the microbial ecologist’s point of view, the availability of this powerful new technique, which has been driven primarily by the needs of the biomedical sciences to define in situ in cells the interactions of ions, molecules and macromolecules, with membranes with a minimum of perturbation, could not be more timely. The past century has seen great advances in microbial physiology and biochemistry, mostly with organisms grown in suspension on rich media and at high growth rates. More recently there is a growing awareness that surface growth, nutrient deprivation, or pulsatile nutrient sufficiency are more realistic modes of laboratory cultivation with respect to the natural environment. But the study of heterogeneity in space and time brings a host of methodological problems not encountered in the shake flask or continuously stirred tank reactor.