Cultivation-independent assessment of bacterial viability is essential when (1) results are required fast and at high throughput, and/or (2) when the specific target or mode-of-action of a certain bactericidal process is of interest, and/or (3) when the organisms under investigation are regarded as "uncultivable". However, aside from cultivation, there exists no "silver bullet" method that demonstrates with absolute certainty whether an organism is alive or dead, and all currently available methods are prone to produce varying results with different organisms and in different environments. Here we discuss the fundamental concept of viability in bacteria, with specific focus on the main aspects that define it. It is argued that the presence of intact and functional nucleic acids, as well as an intact and polarized cytoplasmic membrane are essential components of cellular viability, while numerous other parameters and processes that are linked to viability are explored. Different methods/approaches are discussed with particular emphasis on the advantages and disadvantages of each approach, the applicability of the methods toward environmental samples, and the underlying link between the various viability parameters.