The History of the Development of the Easygel ® and Coliscan ® Methods

In the late 1970s, Dr. Jonathan N. Roth began work on a temperature-independent gelling agent to be used in place of agar-agar, the "vegetable gelatin" upon which the growth of microorganisms on solid media depended. 

Dr. Roth was one of many scientists who found agar difficult to work with. The growth of microorganisms on solid media had depended on agar — a polysaccharide derived from marine red algae — for roughly 150 years. Yet because agar is derived from marine algae, the availability of agar is dependent upon marine environmental conditions. Additionally, since numerous algal species may be used as sources, agar is also subject to variations which can affect research results. 

An even more significant limitation of agar relates to the heating and cooling of the material. To be used, the dried material must be dissolved in 90-100° C water and dispensed at a temperature higher than 45° C as it solidifies below that temperature. This characteristic makes agar difficult to use, particularly when it is desirable to mix a test sample with the still-liquid medium before it solidifies in a container, such as a petri plate. The high temperature required for maintaining the liquid state harms many microorganisms. Scientists have observed that using this pour plate technique with agar-based media results in questionable accuracy in the determination of microbial populations of test samples. 

Dr. Roth's goal was to develop a temperature-independent gelling agent that would enable scientists to avoid the disadvantages of agar. His years of work and evaluation paid off, and Dr. Roth's work with low methoxyl pectins as agar substitutes resulted in the patented product, Easygel ® .

The Easygel Method Dr. Roth created utilizes liquid nutrient formulas containing the gelling agents. These media are poured into containers — most often petri dishes — that are covered with a thin coating of material containing calcium ions. When the liquid medium is poured into the coated dish, the ions diffuse up through the medium and complex with the gelling agents, causing a solid gel to form within 30 - 40 minutes. 

Significantly, the Easygel approach eliminates the temperature effect of hot agar on samples, and results in the first widely-used agar substitute in more than a century. The revolutionary technology is widely used today in water-quality work, and by the food and beverage industries. Many articles have been published on its applications, and Easygel has been approved by different federal and state agencies for use in various forms.

Yet the story of Dr. Roth's scientific developments doesn't end there. He continued to work on increasing the scope of possible applications of his Easygel methodology and technology, soon realizing that there was a need for alternative approaches to the isolation and identification of particular organisms in sample materials. 

Dr. Roth was interested in early work with indigo dye substrates and the identification of both general coliforms and E. coli (the primary fecal coliform) in liquid media. One early method detected whether general coliforms or E. coli were present in the liquid, and marked the presence by coloring the medium -- yellow for general coliforms and blue for E. coli. This method was based on the fact that one could identify a microbe specific to an enzyme, and that virtually all coliform members of the family Enterobacteriaceae produced galactosidase, but only E. coli produced glucuronidase. However, this approach was not quantitative, and the compounds which detected these agents in the medium did not work in solid media (where actual colonies might be counted) due to their solubility.

At that time, the only non-water soluble chromogenic enzyme subtrates that could potentially be incorporated in solid media and result in colored colonies of the desired microbes, contained a chromogen configuration which resulted in a blue colored product. This made it impossible to combine more than one water insoluble chromogenic substrate.

Dr. Roth believed that the availability of different chromogenic substrates able to produce differently colored insoluble dyes would be invaluable in developing a new approach to microbial diagnostics. He began work with dye chemist Dr. Wilfred J. Ferguson to develop a substrate with a color to contrast with the blue. 

The result of their work was Red-gal. Dr. Roth combined the Red-gal with X-gluc in a nutrient medium and confirmed that coliforms grew as pink/red colonies and E. coli grew as blue/purple colonies that were easily distinguishable from each other. The end product resulted in a 1993 patent and is available from Micrology Laboratories as their Coliscan ® media.

The publication of the new method incited a flurry of activity in the chemical industry. Many wanted to expand the current applications and develop new chromogenic compounds. Likewise, many corporations have tried to develop variations on the theme for commercial applications. 

Micrology Laboratories does not limit itself to past accomplishments, but continues to work on the development of new approaches and technologies. Geoffrey N. Roth, Research Chemist, has developed a new method for the detection of E. coli, coliforms and Aeromonas. This method — called ECA Check ® — not only isolates and identifies E. coli and coliforms, but also species of the genus Aeromonas. Aeromonas is a potentially dangerous pathogen commonly found in the intestinal tract of animals. Canada and some European countries closely monitor and control its presence in water, recognizing its importance not only as a pathogen, but as an indicator organism. ECA Check is ready to fill the need in the United States for a rapid, accurate method for the determination of these three organisms in one simple step!