Hunter Stoddard ¹ Daniel Kulas ¹ Ali Zolghadr ¹ Sulihat Aloba ¹ Laura G. Schaerer ² Lindsay Putman ² Isabel Valencia ² Jeffrey A. Lacey ³ David R. Shonnard ¹ Stephen Techtmann ² Rebecca G. Ong ^1*

• ¹ Department of Chemical Engineering, Michigan Technological University, Houghton, United States
• ² Department of Biological Sciences, Michigan Technological University, Houghton, United States
• ³ Biological Processing Department, Idaho National Laboratory (DOE), Idaho Falls, Idaho, United States

Accumulation of plastic waste in the environment is a serious global issue. To deal with this, there is a need for improved and more efficient methods for plastic waste recycling. One approach is to depolymerize plastic using pyrolysis or chemical deconstruction followed by microbial-upcycling of the monomers into more valuable products. Microbial consortia may be able to increase stability in response to process perturbations and adapt to diverse carbon sources, but may be more likely to form biofilms that foul process equipment, increasing the challenge of harvesting the cell biomass. To better understand the relationship between bioprocess conditions, biofilm formation, and ecology within the bioreactor, in this study a previously-enriched microbial consortium (LS1_Calumet) was grown on (1) ammonium hydroxide-depolymerized polyethylene terephthalate (PET) monomers and (2) the pyrolysis products of polyethylene (PE) and polypropylene (PP). Bioreactor temperature, pH, agitation speed, and aeration were varied to determine the conditions that led to the highest production of planktonic biomass and minimal formation of biofilm. The community makeup and diversity in the planktonic and biofilm states were evaluated using 16S rRNA gene amplicon sequencing. Results showed that there was very little microbial growth on the liquid product from pyrolysis under all fermentation conditions. When grown on the chemically-deconstructed PET the highest cell density (0.69 g/L) with minimal biofilm formation was produced at 30 °C, pH 7, 100 rpm agitation, and 10 sL/hr airflow. Results from 16S rRNAsequencing showed that the planktonic phase had higher observed diversity than the biofilm, and that Rhodococcus, Paracoccus, and Chelatococcus were the most abundant genera for all process conditions. Biofilm formation by Rhodococcus sp. and Paracoccus sp. isolates was typically lower than the full microbial community and varied based on the carbon source. Ultimately, the results indicate that biofilm formation within the bioreactor can be significantly reduced by optimizing process conditions and using pure cultures or a less diverse community, while maintaining high biomass productivity. The results of this study provide insight into methods for upcycling plastic waste and how process conditions can be used to control the formation of biofilm in bioreactors.