12 September 2016

Expansion of the bacterial front in the arena with antibiotics

Spectacular experiment of Harvard Medical School

Anatoly Alizar, Geektimes, based on Harvard Medical School: Bugs on Screen

Usually, in laboratory experiments, bacteria are bred in a homogeneous medium. Scientists from Harvard Medical School went further. They organized an unusual experiment on the evolution of bacteria in a mixed environment – on a huge "dish" measuring 120 × 60 cm. 

The space was divided into zones with different concentrations of the antibiotic, so that only generations of bacteria with suitable mutations pass into the next zone. As a result, "super-microbes" make their way to the final round in the center of the arena, that is, those who have maximally changed their genotype and phenotype as a result of evolutionary selection (see about the isolation of adaptants at the end of the article).

The arena of the battle of microbes with antibiotics is called MEGA (microbial evolution and growth arena). Such an environment is specially created so that, with an exponential increase in the number of bacteria, they do not compete with each other for limited resources, as is customary in most scientific experiments. Here, resources are practically unlimited, and bacteria are only required to capture new territories and adapt to new living conditions, multiplying their numbers almost without restrictions. In this sense, the movement of the bacterial front resembles the expansion of the human species on planet Earth in the Middle Ages with the colonization of new territories (presumably, the expansive development of the human race will continue beyond the home planet with the colonization of all habitable territories within reach).

The experiment has not only scientific, but also educational value. The large space of the MEGA arena allows you to visually observe mutations and natural selection during the spread of the bacterial population front. An impressive sight.

Previous studies have shown that structured microenvironments of this type increase the rate of evolution in small populations of bacteria with a change in genotype (Q. Zhang et al., Science 333, 1764-1767 (2011)). But until now, the question of how exactly this happens in large populations has remained unexplored.

For such an experiment, a rectangular Petri dish measuring 120× 60 cm was constructed, structured according to zones with an exponential increase in the concentration of the antibiotic trimethoprim from the periphery to the center, as well as with nutrients for bacterial reproduction. The large area of the arena did not allow E. coli bacteria to mix with each other in order to more clearly observe the mutations occurring.


The design of the MEGA arena and the result of the bacterial front spreading over 12 days are shown in Figure B. Circles of different colors indicate 182 species of mutated bacteria, the color indicates the concentration of bacteria. The lines between the species correspond to the direction of mutation, based on the video data.

With the constant increase in resistance from the antibiotic, numerous parallel lines of evolution appear in the bacterial population, which differ in phenotype and genotype.


Having studied the bacteria in the vanguard and behind the front of the bacterial population, scientists have found out several interesting things. It turned out that evolution is not always driven forward by the bacteria that are most resistant to antibiotics. Oddly enough, sometimes the most stable hereditary lines are locked behind the more sensitive bacteria. Apparently, this is due to "premature" mutations, when some bacteria are ready to survive in a higher concentration of the antibiotic, which will appear in the future, but has not yet appeared. In such a situation, potentially more adapted bacteria give way at the front to their relatives, who are adapted precisely to the current concentration that exists at the moment. 

To test this theory, scientists took samples of isolated bacterial colonies with "premature" mutations and forcibly placed them in front of the front. As expected, they survived in conditions in which the main bacterial front cannot survive.


The spatial trap of compensatory mutations is bacteria that are so ahead of their time that even after the onset of suitable conditions, they are already locked behind the advancing front. Illustration: Harvard Medical School

Scientists have carefully studied the genotype of the most mutated bacterial species that managed to survive in a solution with a maximum concentration of trimethoprim. It turned out that in these species, the folA gene, which encodes dihydrofolate reductase (DHFR) and is the target of trimethoprim, was most often mutated. The greater the resistance of bactria to the antibiotic, the more mutations there were in this gene. In addition, mutations were found in several other genes that are not related to the action of a particular antibiotic. Among them were the operons mar and sox, which are responsible for the stress response. It was previously known that these "stress" genes play an important role for successful antibiotic resistance.

Scientists have also found that better adaptation to the weak effects of an antibiotic accelerates adaptation to higher concentrations later (in the illustration below). It's like people who are better able to adapt to deteriorating living conditions if changes occur gradually and imperceptibly.


The experiment is described in a scientific article published on September 9, 2016 in the journal Science (Baym et al., Spatiotemporal microbial evolution on antibiotic landscapes).

Portal "Eternal youth" http://vechnayamolodost.ru  12.09.2016

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