Are they alive? Detection of viable organisms and functional gene expression using molecular techniques

Abstract

The ultimate measure of microbial viability and biological activity is growth in some form of culture system. Unfortunately, due to many limitations, growth is usually not the most sensitive or rapid detection method. This chapter describes many of the molecular-based tools for assessing viability and functional gene expression, and their applications for specific microbes in environmental samples. Methods include fluorescent nucleic acid binding dyes, enzymatic conversion of substrates to fluorescent compounds (often in conjunction with nucleic acid-based methods), various techniques based on amplification and detection of nucleic acids, nucleic acid amplification linked to biosensors and microarray detection platforms, detection and characterization of proteins, and molecular detection coupled with culturing. Principles supporting each of these techniques are discussed along with applications to bacteria, protozoa, and viruses, focusing primarily on microbes of concern to the drinking water and wastewater industries.