MICROSCOPICAL TECHNIQUES FOR THE STUDY OF PHOTOTROPHIC BIOFILMS ON STONE FACES

摘要

The application of light and electron microscopy methods to the study of microbial communities forming biofilms on the surface of different substrata has shown to offer valuable possibilities in the visualisation of microorganisms, providing the means to characterise structural interactions between species and in resolving ultrastructural details at a micro- and nanometer scale. However, examination of microorganisms in their natural habitat may be achieved most effectively through the application of a variety of microscopic techniques that are necessary to assess the microenvironment, the state of the organism and its taxonomic affiliation. Early studies on microbial films began with the use of scanning electron microscopy (SEM) mainly because of the minerals present in the samples. This method revealed the external diversity and the stratification of microorganisms as well as some of the processes involved in mineralisation. Transmission electron microscopy (TEM) was, however, essential for the identification of many species, especially of those possessing distinctive ultrastructural features like photosynthetic microorganisms. Depending on the presence of intracytoplasmic chromatophore membranes or chlorosomes, green and purple bacteria can be distinguished. Similarly, cyanobacteria and the various groups of eukaryotic microalgae and mosses can be identified by the arrangement of thylakoids and the presence or absence of phycobilisomes on the photosynthetic membranes. Intracellular inclusions like carboxysomes, polyphosphates cyanophycin, poly-β-hydroxybutyrate, polyhydroxy-alkanoates, glycogen and other C-storage compounds are also useful in the identification of different species of microorganisms and provide a good picture of their nutritional and/or metabolic conditions. In addition, cytochemistry and immunocytochemistry, the former based on the use of specific stains and the latter on gold conjugated secondary antibodies for different primary antisera, can be successfully applied for the localisation of sugars, proteins and key enzymes in metabolic processes.