Studies by a Northwestern University-led team suggest that a common environmental bacterium, Comamonas testosteroni, may someday be harnessed as nature’s plastic recycling center. Most bacteria prefer to eat sugars, but C. testosteroni has a natural appetite for complex waste from plants and plastics. The new study has, for the first time, deciphered the metabolic mechanisms that enable C. testosteroni to digest these otherwise seemingly undigestible. The findings could potentially lead to novel biotechnology platforms that harness the bacteria to help recycle plastic waste.

Research lead Ludmilla Aristilde, PhD, an associate professor of civil and environmental engineering at Northwestern’s McCormick School of Engineering, explained to GEN, “In this research, we unravel the multi-level complex mechanisms of a bacterial species with metabolic capabilities for plastic waste degradation products … the methodological approaches used in this research could be explored to probe the mechanisms of other microbial species of biotechnological relevance.”

Aristilde and colleagues reported on their work in Nature Chemical Biology, in a paper titled, “Complex regulation in a Comamonas platform for diverse aromatic carbon metabolism.”

Comamonas species are found nearly everywhere—including in soils and sewage sludge. C. testosteroni first caught researchers’ attention with its natural ability to digest synthetic laundry detergents. “C. testosteroni KF-1 was isolated from sewage sludge for its capacity to degrade aromatic synthetic laundry surfactants,” the team explained. Scientists then discovered that this bacterium also breaks down compounds from plastics and the fibrous, woody plant material lignin.

“Soil bacteria provide an untapped, underexplored, naturally occurring resource of biochemical reactions that could be exploited to help us deal with the accumulating waste on our planet,” explained Aristilde, who is also a member of the Institute for Sustainability and Energy at Northwestern’s program on plastics, ecosystems, and public health.

Most research that aims to engineer bacteria has used Escherichia coli bacteria, which represents the most well-studied bacterial model organism. But E. coli, in its natural state, readily consumes various forms of sugar. As long as sugar is available, E. coli will consume that—and leave plastic chemicals behind.

In contrast, Comamonas bacteria are incapable of using sugars. “Strains of C. testosteroni lack the genes required for carbohydrate utilization and have an innate preference for gluconeogenic substances such as aromatic compounds,” the authors noted. Aristilde added, “Engineering bacteria for different purposes is a laborious process. It is important to note that C. testosteroni cannot use sugars, period. It has natural genetic limitations that prevent competition with sugars, making this bacterium an attractive platform.”

C. testosteroni instead utilizes others sources of carbon that contain compounds with a ring of carbon atoms. Aristilde further explained to GEN, that this inability of the bacterium to use carbohydrates (i.e., sugars) means that it can be used to metabolize specifically the aromatic carbons (i.e., non-sugar chemicals) found in plastics and plant waste. “In contrast, almost all microbial platforms currently being explored for plastics of plant waste can suffer from competition between carbohydrate substrates and the metabolism of aromatic substrates,” she said.

But while researchers have known that C. testosteroni can digest these ring compounds, the mechanisms involved haven’t been well understood. “Metabolic regulation in C. testosteroni strains during assimilation of aromatic compounds remains to be elucidated, thus severely limiting the potential exploitation of these wastewater isolates to metabolize xenobiotic aromatic compounds,” the investigators noted. Aristilde pointed out, “These are carbon compounds with complex bond chemistry,” she said. “Many bacteria have great difficulty breaking them apart.”

Aristilde further explained to GEN: “To leverage microbial metabolism and engineer a target objective using a microbial platform, it is important to figure out first how metabolism is naturally regulated in the microbe. The reason for this is because one can engineer one pathway for a specific target but the effort could be unsuccessful without overcoming the regulation controls of that pathways, resulting in a failure to achieve the target objective. The researchers wanted to investigate how the Comamonas bacteria can degrade and assimilate these complex forms of carbon. To do this, the researchers, including first author Rebecca Wilkes, and collaborators at the University of Chicago, Oak Ridge National Laboratory, and Technical University of Denmark, combined multiple forms of omics-based analyses.

By examining the data resulting from these multi-omics analyses, Aristilde and her team were able to map the metabolic pathways that the C. testosteroni KF-1 bacteria use to degrade the lignin-related aromatic compounds 4-hydroxybenzoate (4HB), and vanillate (VN), and the plastics-related xenobiotic aromatic compound, terephthalate, (TER), into carbons for food. “As a requisite to exploiting C. testosteroni and other non-model species, comprehensive multi-omics studies of their cellular physiology are necessary,” they wrote. “Leveraging orthogonal methods to elucidate the metabolic controls underlying aromatic carbon fluxes in C. testosteroni KF-1, we combined transcriptomics and proteomics measurements with targeted metabolomics data, 13C-kinetic profiling, 13C-fluxomics analysis, and 13C-isotopomer fragmentation analysis during feeding on 4HB, VAN, and TER.”

Such far-reaching multi-omics studies are massive undertakings that require a variety of different techniques. Aristilde leads one of few labs that does carry out such comprehensive studies. Through their work, the team discovered that the Comamonas bacteria first break down the ring of carbons in each compound. After breaking open the ring into a linear structure, the bacteria then continue to degrade it into shorter fragments.

“We started with a plastic or lignin compound that has seven or eight carbons linked together through a core six-carbon circular shape forming the so-called benzene ring,” Aristilde explained. “Then, they break that apart into shorter chains that have three or four carbons. In the process, the bacteria feed those broken-down products into their natural metabolism, so they can make amino acids or DNA to help them grow.”

Aristilde commented to GEN, “… plastics are polymers, which consist of a chain of monomers. Upon the breakdown of the plastics, the bacterium is making use of the monomers (i.e., the breakdown products) derived from the plastic polymer. Our study is focused on the metabolism of the breakdown products from the plastics. As we highlighted in the paper, several subsequent products (i.e., metabolites) in the metabolic network are known valuable chemicals of commercial value.”

The team’s analyses also showed that C. testosteroni can direct carbon through different metabolic routes. “Transcription-level regulation controls initial catabolism and cleavage, but metabolite-level thermodynamic regulation governs fluxes in central carbon metabolism,” the investigators wrote. “We found that the metabolism of C. testosteroni is regulated on different levels, and those levels are integrated,” Aristilde said. “The power of microbiology is amazing and could play an important role in establishing a circular economy.”

These different metabolic routes could lead to useful byproducts that might feasibly be used for industrially relevant polymers such as plastics. So while other researchers have worked to engineer bacteria that can breakdown plastic waste, Aristilde believes bacteria with natural abilities to digest plastics hold more promise for large-scale recycling applications.

The team is currently working on a project investigating the metabolism that triggers polymer biosynthesis. “New insights were obtained on the regulatory mechanisms during aromatic carbon utilization that involve complex relationships between transcript expression, protein abundance, metabolite levels, and metabolic fluxes,” the scientists stated. “… These new multi-omics perspectives combined with targeted 13C mapping of specific metabolic nodes present a framework of guiding principles to exploit metabolic reactions for aromatic carbon catabolism in C. testosteroni and other bacterial specialists of biotechnological relevance.”

Aristilde further stated, “These Comamonas species have the potential to make several polymers relevant to biotechnology. This could lead to new platforms that generate plastic, decreasing our dependence on petroleum chemicals. One of my lab’s major goals is to use renewable resources, such as converting waste into plastic and recycling nutrients from wastes. Then, we won’t have to keep extracting petroleum chemicals to make plastics, for instance.”

Aristilde expanded on this to GEN: “This work is really at the beginning … In general, we are interested in how we can explore natural abilities of different bacteria to make polymers. Some of these polymers can be used as precursors to plastics (i.e., bioplastics, essentially using plant or plastics wastes instead of petroleum as the starting material), some of these polymers can be used as a way to recover nutrients for nutrient recycling relevant to agriculture, etc … In our study, we only investigate degradation products from PET-type plastics. Given the capabilities of this bacterium—C. testosteroni—we anticipate that it can utilize degradation products from other plastics but this was not investigated explicitly in this current study.”

While continuing to work with Comamonas species, the researchers are also exploring the mechanisms of other known waste-degrading enzymes, GEN learned. “My team also has expertise in Pseudomonas species, which are also of biotechnological importance due to their diverse metabolic capabilities,” Aristilde noted.


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