In the past decade, the development of new tools to better visualize microbes at the cellular scale has spurred a
renaissance in the application of microscopy to the study of bacteria in their natural environment. This renewed interest
in microscopy may be largely attributable to the advent of the confocal laser scanning microscope (CLSM) and to the
discovery of the green fluorescent protein. This article provides information about the use of fluorescence microscopy
combined with fluorescent labels such as GFP, DsRed, and DNA stains, with immunofluorescence, and with digital
image analysis, to examine the behavior of bacteria and other microbes on plant surfaces. Some of the advantages and
pitfalls of these methods will be described using practical examples derived from studies of the ecology of foodborne
pathogens, namely Salmonella enterica and E. coli O157:H7, on fresh fruit and vegetables. Confocal microscopy has
been a powerful approach to uncover some of the factors involved in the association of produce with epidemics caused
by these human pathogens and their interaction with other microbes in their nonhost environment.
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