[16] Phenotype characterization of genetically defined microorganisms and growth of bacteriophage in biofilms

This chapter discusses biofilms as a dominant mode of growth of bacteria and other microorganisms in nature. The Centers for Disease Control (Atlanta, GA) now estimates that at least 62% of all bacterial infectious diseases, particularly chronic infections, involve biofilms. In contrast to their planktonic counterparts, biofilm organisms are quite resistant to antibiotics and disinfectants. Biofilms are also important in other areas including wastewater treatment, biofouling of pipes, and microbially induced corrosion. The early studies of biofilms employed conventional scanning (SEM) and transmission electron microscopy (TEM) to observe biofilms with a high degree of resolution. Biofilms are highly hydrated, typically >90% H20 in most environments. As a result, the dehydration employed for conventional SEM and TEM causes hydrated structures within biofilms to collapse, giving a mistaken impression that individual cells were simply thrown together. Other drawbacks to conventional SEM and TEM are that the chemicals and high vacuum cause a loss of cell viability. Environmental SEM permits the examination of hydrated biofilms, however its major drawback is that organic molecules cannot be resolved unless stained with heavy metals. Scanning confocal laser microscopy (SCLM) lacks the resolving power of SEM and TEM; however, it permits the observation of fully hydrated and living biofilms. This chapter discusses two major issues: (1) identifying and testing genes for their importance in biofilm physiology, and (2) studying the interaction of bacteriophage with bacteria in their natural environment (that is, biofilms).