Thursday, 10 July 2014

Culturing microorganisms

For over 100 years microbiologists have known, to their great consternation, that in any given environment, at least 90% of all microbes are unculturable. In the last few decades, we have developed molecular tools to see these microbes in the environment and determine their presences and number, but we still cannot bring them into culture. There are all sorts of ideas for why this might be true. Many of these microbes may be slow growing, and the fast growing microbes easily outpace them in laboratory medium. Maybe the media we use is "too rich" for many microbes and we need to create media with more dilute nutrients to capture these shy microbes. Maybe the metabolic demands of these microbes were too difficult to replicate in the laboratory and many of them will not grow. Whatever the reason, as of right now, we are nearly blind when it come to these unculturable microbes.

A landmark paper was published a year ago in Applied and Environmental Microbiology. Buerger et. al created a simple, elegant experiment to try to capture the slow-growing bacteria. Sediment from the ocean shore of Massachusetts Bay was diluted and then dispensed into wells of a microtiter plate such that each well received only 1 microbe per well. The growth medium used was 0.1x LB medium, a dilution of a common, rich medium. The plates was carefully wrapped and placed in a humidified chamber for 18 months. Yes, that is a 1.5 year experiment! Periodically the plates were checked for growth and any well showing growth was then characterized.

The expected result was that the slow-growing microbes would arrive later and the number of novel species isolated would increase as the length of the incubation increased. This would fit the hypothesis that the slow growers were being out competed. To their surprise, this was not the case. As the experiment went on, the percentage of novel species did not increase, but stayed remarkably similar. In addition, bacteria isolated by this technique did not grown much slower than microbes that have been cultivated for many years. Buerger et al. hypothesize that cells in the environment may be in a dormant state, and only randomly emerge from this state and grow. This "Scout Hypothesis" would explain the results that they were seeing. They also did a similar experiment on a soil sample and observed similar results

The implication of this experiment is that it should be possible to isolate and characterize many more unculturable species using very simple, and inexpensive techniques. Given enough experimentation, scientists may be able to culture a much larger proportion of the species present in an environment.
Posted by Paustian