Formatted Title
Deep Defluorination of n:3 Acids via One-Carbon Removal Pathway by a Novel Enrichment Consortium
Background/Objectives
Historical discharges of aqueous film-forming foams (AFFFs), containing per-and polyfluoroalkyl substances (PFAS), have led to elevated PFAS concentrations in environmental settings. Phasing out perfluorooctane sulfonate (PFOS) resulted in a transition to wide uses of fluorotelomer-based AFFFs. As a result, dozens of fluorotelomers, such as 6:2 fluorotelomer sulfonate and 6:2 fluorotelomer sulfonamide alkyl betaine, and their semi-transformed byproducts are detected ubiquitously in the environment. Unlike PFOS or perfluorooctanoic acid (PFOA), n:2 fluorotelomers exhibit the potential for biological defluorination, forming perfluoroalkyl carboxylates (PFCA) and fluorotelomer carboxylic acids (FTCAs). FTCA may exhibit higher toxicity than PFCA of comparable chain length but is more biodegradable owing to its alkyl moieties. A "one carbon removal pathway", which was proposed more than 10 years ago for the extensive biotransformation observed for 6:2 fluorotelomer alcohol, resulted in sequential ─CF2─ removal from fluorotelomers, causing partial defluorination under aerobic conditions and producing 5:3, 4:3, 3:3 fluorotelomer acids (n:3 acids). However, the microorganisms implicated in this process remain under-researched. A thorough understanding of the microbes, genetics, and biochemistry behind these processes could lead to future cost-effective biological treatments.
Approach/Activities
In this study, we calculated the thermodynamics along the one-carbon removal pathway and scrutinized the aerobic defluorination of n:3 acids using activated sludge and soil microbial consortia under eight distinct co-metabolic laboratory conditions. A dual-stage enrichment strategy was employed: the initial stage utilized highly diluted culture media to support slow-growing microbes, while the subsequent stage used rich media to expedite microbial growth and degradation. The consortium was then optimized for medium, dilution rate and transfer interval. Its viability after reactivation from -80 ⁰C was also tested for long-term preservation. For the degradation experiment, PFAS biotransformation products, fluoride and microbial community structure were monitored and analyzed.
Results/Lessons Learned
After a year of enrichment with 5:3 acid, a degrading consortium was obtained under one specific condition, which was then maintained and optimized. This consortium demonstrated the ability to degrade 5:3 acid into 4:3 acid, 3:3 acid and 2:3 acid within 47 days. Its defluorination potential for n:3 acids (n = 2-5) and 6:2 fluorotelomer unsaturated acid was evaluated and defluorination rates of 24%, 85%, 33%, 55%, and 66%, respectively, were observed. Twenty-three bacterial strains were isolated from the consortium via dilution plate technique and identified using 16S rRNA gene sequencing. Interestingly, none of these isolates could perform defluorination individually. The bacterial community structure and evolution during the incubation were also characterized using 16S rRNA-based metagenomic analysis. This study represents the first successful enrichment and phylogenetic identification of microbes responsible for the one-carbon removal pathway in fluorotelomer degradation. The availability of the unique culture establishes the groundwork for future engineered biological treatment processes to achieve extensive or complete defluorination of fluorotelomers.