People often recognize social behaviors in complex organisms such as insects, nonhuman primates, and humans. But Megan Frederickson, an ecologist and evolutionary biologist at the University of Toronto, is interested in a different, microscopic social community: bacteria. “Cooperation is everywhere,” she said. “Cells cooperate in multicellular organisms; individuals cooperate in societies; and different species cooperate… Why would it not be the case that microbes cooperate with each other?”
Researchers have known for more than 20 years that bacteria participate in collective behaviors such as forming biofilms and acquiring nutrients from the environment. But being part of a cooperative group does not necessarily mean that every individual bacterium plays by the rules. Occasionally, cheaters arise.
Conflict in bacterial communities
For cheating to happen, it must arise from an adaptive strategy, said Frederickson. “[The individual] should have a fitness benefit from not cooperating or not cooperating as much as others in its environment.” Additionally, for a bacterium to be considered a cheater, it has to evolve from an ancestrally cooperative state. “We should reserve the word cheating to mean a breakdown in cooperation,” she added.
Cheaters exploit the work of others by consuming shareable resources that are costly to produce for an individual but benefit the entire population.1 These public goods can take many forms in a bacterial community: exopolysaccharides to build biofilms, enzymes that degrade antibiotics, or scavenging molecules to cope with nutrient scarcity in the environment.2
We should reserve the word cheating to mean a breakdown in cooperation.
—Megan Frederickson, University of Toronto
But how does a bacterium become a cheater? Unlike humans, bacteria do not make an intentional decision to cheat. “[These individuals] lack a gene or have a protein that is no longer functional. That is why they stop making a social trait and become cheaters,” said Rolf Kümmerli, an evolutionary biologist at the University of Zurich who studies how bacteria share metabolites such as enzymes and iron-scavenging molecules.
Armed with this knowledge, researchers tweak bacterial genetic material to better understand cheating. For instance, they can knock out genes responsible for the production of shared metabolites to generate cheating mutants in the lab. Then they can culture these nonproducers along with producers and measure their relative fitness, that is, the ratio of the growth rates of the two strains. If a resource is essential for growth, nonproducers should outgrow producers in a medium containing both strains as they consume the shared resource without paying the metabolic cost to produce it.
This bacterial arms race appears for many communal compounds. For example, by using the opportunistic pathogen Pseudomonas aeruginosa, scientists showed that cheaters can exploit iron-scavenging compounds called siderophores,3 as well as proteases used to digest nutrients.4, 5 Another example of an exploitable compound is a bacterial enzyme used to degrade antibiotics. Researchers conducted experiments using E. coli, which resist β-lactam antibiotics by producing β-lactamase enzymes. Mutants that do not produce those enzymes tolerate higher doses of ampicillin, a β-lactam antibiotic, for a longer time when sharing a biofilm with enzyme producers.6
An intriguing question for many scientists is what makes bacterial cooperation persist when cheaters exist in several natural and disease settings.2 “It is really fascinating why we do not see [cheating] happen more often. We see lots of examples of cooperation in bacteria and, by and large, they are not undermined all that often by the evolution of [cheaters],” said Frederickson.
Cheaters benefit from cooperators in bacterial communities
In cooperative bacterial communities, cheaters may arise. These bacteria do not pay the cost of producing public goods but still consume the shareable resources produced by other cooperative bacteria.
© Ashleigh Campsall
Keeping cheaters at bay
One idea that partially explains cooperation is frequency-dependent selection. According to this idea, a cheater genotype appears when there are few cheaters among many cooperators. If the cheater genotype becomes more frequent, the proportion of producers might decrease to a point that being a cheater is no longer an adaptive strategy, causing this genotype to be selected against.
Additionally, cooperators use different strategies to optimize public goods usage and reduce cheaters’ access to them. One such approach is spatial structuring, that is, the 3D arrangement of a bacterial community.
“Spatial structure is the easiest answer for why we don’t have cheaters running wild,” said Steve Diggle, a molecular microbiologist at the Georgia Institute of Technology who investigates microbial interactions and social behaviors. In the real world, bacteria are commonly organized in densely packed biofilms—slimy, multicellular structures—that favor cooperation. Biofilms help cooperators by keeping related individuals physically close to each other and limiting the diffusion of shared compounds due to its viscous composition. Kümmerli’s group replicated a biofilm environment in laboratory experiments to confirm this notion. The team cultured P. aeruginosa siderophore producers and nonproducer mutants in a viscous growth medium and found that limited dispersal of the iron-loaded scavenging molecules resulted in increased fitness of the producer population.7
Spatial structuring also helps bacteria keep their kin closer, and relatedness favors cooperation. “Relatedness is obviously a really key driver of social interaction. So, generally, the higher the relatedness between interacting partners, the more cooperation will be selected for because those cooperative genes are shared between the cooperator and the recipient,” said Siobhán O’Brien, an evolutionary ecologist at Trinity College, Dublin.
This idea, called kin selection, explains why a trait that does not directly benefit an individual might be favored in a bacterial group where a bacterium is most likely surrounded by cells sharing a similar genetic background. By helping a neighboring cell do well, a bacterium ensures that its genes pass on to the next generation.
Strategies used by cooperators to curb the cheater population in a bacterial community
© Ashleigh Campsall
Spatial structuring and kin selection
As cooperators build biofilms, they keep their kin close and limit cheaters’ access to communal resources.
Cooperators discriminate kin from non-kin and share resources only with highly related cells.
Cooperating bacteria can sanction cheaters by producing diffusible toxins together with a resource.
Another approach by cooperators to minimize cheater populations is to direct communal resources only to their kin. “Some public goods…are quite specific. They can only be taken up by cells with the correct receptor, and normally that means they are kind of strain specific. If [there is] another strain of the same species that is quite distantly related, they won’t possess the correct receptor to take up the public good,” O’Brien said. The iron-scavenging molecule pyoverdine is a good example of this. In a natural community, researchers showed that some pyoverdine producers secrete incompatible pyoverdines that cannot be used by nonproducers, suggesting that these cheaters lack the receptors needed to take up these molecules.8
Discriminating kin from non-kin also enables cooperators to directly sanction cheaters by policing them. To police communities, cooperators may produce an energetically costly compound, often a toxin, to control the number of cheaters within the group. For instance, P. aeruginosa produces a shareable protease along with a toxin, cyanide. Cooperators produce an immunity factor that protects them from the cyanide, thus selectively killing cheaters.9
As social creatures, bacteria perform a variety of collective behaviors that enable them to thrive in different environments. Like other social animals, they must also deal with individuals that do not cooperate but still want to take a share of the goods produced by others. Although cheaters threaten bacterial cooperation, they do not undermine it as both cooperators and natural selection weigh in to control their spread.
“The interest in the social lives of microbes is because they make a good experimental system for our larger ideas about how cooperation works, including in animals or in plants,” Frederickson said.
Additionally, a better understanding of the social dynamics of bacteria might help us either come up with new therapies or offer ideas that we could use for biological control, added Frederickson. “We think of [bacteria] as simple organisms, but they are not. They have been evolving on this planet for a very, very long time. And they are capable of some very sophisticated things. I think we don’t give them quite enough credit.”
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