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The process of biofilm formation has knowingly, and even unsuspectingly, baffled scientists for almost as long as the field of microbiology itself has existed. This Special Issue of the International Journal of Molecular Sciences (IJMS) specifically addresses an important component of the biofilm, the extracellular matrix. This matrix forms the protective secretions that surround biofilm cells and afford a “built environment” to contain biofilm processes. During the earlier days of microbiology, it was intriguing to Claude ZoBell that attached bacteria sometimes were able to proliferate when their planktonic counterparts were unable to grow [1]. During the 1970s, this attached state was beginning to be explored [2], and it was realized to be anchored in a matrix of slime-like molecules. The slime-like matrix together with cells was to be called the “biofilm”, a term developed by the late Bill Costerton, Bill Characklis and colleagues. The scientific revelation that attached bacteria were different from free (i.e., planktonic) cells in their physiological behavior and adaptability, launched an era of focused exploration in this area of microbiology. It was initially surprising, though not unexpected in retrospect, that interest in biofilms has grown and now infiltrates virtually all aspects of our scientific study. Since that time there has been a near-exponential growth in the numbers of scientific publications addressing biofilms owing to their immediate relevance to ecology, biotechnology, health and industry.