A new study with implications for environmental and medical research found that the ability of bacterial communities to work together to form biofilms, the thin layers of microbes found throughout nature and in the human body, is highly dependent on temperature.

The study, published May 21 in Frontiers in Microbiology, fills a known gap in knowledge regarding the role of non-biological factors in biofilm formation. It also explores the complexity of interactions between different microbial species, which often behave and grow differently in communities than they would on their own.

"Microbial model communities are essential instruments to learn about metabolic interactions, genetic mechanisms and ecological principles governing and structuring communities," said senior author Johan Bengtsson-Palme, an assistant professor of infectious diseases at Göteborgs universitet in Sweden. "Our best shot at understanding this complexity is likely to reduce it into components we can more easily tease apart and then figure out the larger picture piece by piece."

Microbial communities that form biofilms are found almost everywhere in the environment, and they are also found in and on every larger organism, including people. Over 75% of infectious diseases are somehow associated with biofilms. These films can promote antibiotic resistance, contribute to chronic illnesses such as cystic fibrosis and can also contaminate medical devices, particularly those that are implanted into the body.

However, these microbial communities are not universally harmful. Human microbiota have been used to treat obesity, and Americans are more interested than ever in using probiotics as a wellness strategy to boost microbiome health.

In agriculture, plants have been shown to benefit from their microbial communities, and the symbiotic relationship between plants and biofilm-forming soil microbes has been implicated in optimal crop production. It was soil microbes that the current study focused on.

The researchers started off hoping to learn more about ecological invasion of soil microbes using THOR, which stands for The Hitchhikers Of the Rhizosphere, a microbial community consisting of three bacterial species that were isolated from the soil around soybean roots. This community was first developed by researchers at the University of Wisconsin-Madison, where Bengtsson-Palme did his postdoctoral research.

However, when he and his graduate student, study first author Emil Burman, attempted to establish their own populations of THOR, they encountered a problem.

"Emil could not [consistently] replicate one of the most basal properties of the community — the pattern of increasing biofilm formation with addition of community members that do not form biofilms on their own," Bengtsson-Palme said. "Sometimes the experiments showed the expected results, and sometimes it just would not work, seemingly at random."

As they started investigating what could be causing this disparity in results, the researchers discovered that, while they assumed they were culturing their bacteria at a consistent 20 degrees Celsius (68 degrees Fahrenheit), the ambient temperature of the lab varied significantly, particularly in the summer.

"The discovery that temperature seemed to be so important for community interactions led us to more systematically investigate how temperature influenced how the microbes in the THOR model community interacted with each other, using the biofilm formation pattern as a reporter for these interactions," Bengtsson-Palme explained.

To complete this systematic investigation, the team grew the THOR microbial community at different temperatures and measured how much light could pass through the resulting biofilms. The duo also compared this to the growth of each of the three species independently.

"We found that the interactions between the community members were highly dependent on temperature, and that this could not only be explained by changing growth rates of the individual members," Bengtsson-Palme said.

In other words, the effect of temperature goes beyond a simple cause-and-effect scenario. Rather, something happens at suboptimal temperatures that interferes with the ability of the microbes to work together, even if they are growing successfully on their own.

While the researchers determined that 18 degrees Celsius (64.4 degrees Fahrenheit) yielded the best biofilms, even a small drop to 15 degrees Celsius (59 degrees Fahrenheit) led to a large decrease in the level of biofilm formation. When they raised the temperature to 25 degrees Celsius (77 degrees Fahrenheit), just above room temperature, there was almost no detectable biofilm.

"This may be hugely important as an impact of climate change," Bengtsson-Palme said, "but what the exact effect would be is extremely hard to predict with the knowledge we have today."

Now that they've established the best conditions to grow THOR, the researchers are returning to their original research objective — using this community as a model to study soil microbe invasion — though they hope that their research will also inspire other labs to undertake similar projects.

"Ultimately, we would like to see more research on the stability of community behaviors in the face of changes in environmental conditions, such as temperature, pH, nutrient availability, et cetera," Bengtsson-Palme said.

More broadly, they hope that their findings will help other researchers culture these microbial communities more consistently, which is essential for studying real-world problems related to biofilms.

"Our findings likely extend to other microbial communities and other environmental parameters," Bengtsson-Palme said. "Temperature could affect community stability and may thereby indirectly influence things like soil productivity and disease suppression, the efficiency of wastewater treatment, as well as bioprocessing in industrial applications."

The study, "Microbial community interactions are sensitive to small changes in temperature," published May 21 in Frontiers in Microbiology, was authored by Emil Burman and Johan Bengtsson-Palme, Göteborgs universitet.