Acidimicrobiaceae sp. A6 [A6]) is a novel bacterium, capable of oxidizing NH₄⁺ while reducing ferric iron, as shown in the equation below.

3Fe₂O₃ ⋅0.5H₂O +10H⁺ + NH₄⁺ → 6Fe²⁺ + 8.5H₂O + NO₂^-

This process has been named Feammox. A6 has been isolated  and its genome has been sequenced. Through metagenomic analyses, it was revealed that the genome of A6 contains multiple dehalogenase genes, which were also detected in the A6 enrichment culture (GenBank accession numbers MK358459-MK358462). These include a gene for a reductive dehalogenase homolog (rdhA), a gene for a fluoroacetate dehalogenase homolog (FceA), and two putative haloacid dehalogenase genes (dhl_1 and dhl_2). Results of initial incubations with perfluorooctanoic acid (PFOA) and perfluorooctanesulfonic acid (PFOS) as well as other PFAS, revealed that A6 could degrade these PFAS and produce F- during the Feammox process as well as several shorter carbon chain perfluoroalkyl acides (PFAAs). Initial results also revealed that both, the rdhA and FceA genes are expressed when PFAS are being defluorinated.

The objective of this proof-of-concept study was to verify these initial findings, determine if other compounds of interest such, as perfluorohexane sulfonate (PFHxS), could also be degraded, and gain further insights into the mechanism by which A6 defluorinates these PFAS, including the role of the two dehalogenase genes expressed during the defluorination.

Incubations were conducted in vials containing either PFOA, PFOS, or PFHxS, with pure cultures of A6 and A6 enrichment cultures. Incubations were conducted with these cultures while oxidizing NH4+ and reducing Fe(III). The parent compound, potential degradation intermediates, as well as end products including F-, SO42- for PFOS and PFHxS, and acetate were tracked over time. A6 numbers in both cultures and change in the microbial community of the enrichment culture were tracked, as well as the expression of genes for key oxygenases and dehalogenases, that are thought to play a role in the degradation of these PFAS.

These incubations allowed for a careful monitoring of the expression of the rdhA (reductive dehalogenase homolog) and FceA (fluoroacetate dehalogenase homolog) genes. It was decided to use initial concentrations of ~ 10 mg/L, which is substantially lower than the 100 mg/L used previously, but still allowed for tracking of fluoride produced.

The key conclusions from this research include:

1. PFOA, PFOS, and PFHxS can be defluorinated by the pure A6 and A6 enrichment cultures during the oxidation of NH4 + by A6 while reducing Fe(III). Consistent with previous findings, PFOA is defluorinated at a higher rate than PFOS, especially by the pure A6 culture.
2. For all PFAS tested so far, a gene for a reductive dehalogenase homolog (rdhA) and a gene for a fluoroacetate dehalogenase homolog (FceA), are always expressed when PFAS are being defluorinated by A6.
3. There is a strong correlation between the expression of the rdhA gene and the production of F during the incubations. This correlation is much weaker for the FceA gene.
4. Consistent with the stronger correlations described above, gene knockout experiments indicate that the rdhA gene plays a key role in the defluorination of PFAS. Defluorination still proceeded when the FceA gene was knocked out.
5. The detection of H-PFOA during the incubations with PFOA, with the missing fluorine likely from the alpha carbon, indicates that the first step in the defluorination is the removal of the fluorine on that carbon. Since there is no detected defluorination in most of the incubations with the rdhA mutants, this first defluorination step is attributed to the rdhA.
6. The role of the FceA in the PFAS degradation process, if any, is not clear at this point.
7. So far, only degradation by A6 of 1,2,3 TCP as a chlorinated compound of interest was examined. The preliminary results indicate that A6 is capable of degrading TCP and produce 43 DCP in this process. While the rdhA gene was not expressed, or less than in the positive control, during the degradation of TCP, the two putative haloacid dehalogenase genes (dhl_1 and dhl_2) were expressed, indicating that defluorination and dechlorination by A6 is carried out by different enzymes.
8. Incubations of mixtures of 1,2,3 TCP and PFOA show that all genes discussed above are expressed, and that the presence of either compound does not affect the degradation of the other compound, confirming the conclusion that defluorination and dechlorination by A6 are separate processes.
9. To avoid a long lag time, A6 requires acclimation to PFAS. Acclimation results in PFAS carryover, including their degradation products, even after multiple washing of the cultures, which needs to be accounted for in these experiments for their proper design and interpretation. The high carryover made it impossible to detect the production of intermediates in the incubations with a much lower Feammox activity.
10. The defluorination kinetics of PFOA/PFOS are first order with respect to their concentration over the concentration range examined. The kinetics are proportional to the rate of NH4 + (electron donor) oxidation, and for the experimental conditions discussed here, are proportional to the A6 biomass to the 1/3 power.
11. Given the direct proportionality between the PFAS defluorination rate and the NH4+ oxidation rate, and the proportionality to the A6 biomass only to the 1/3 power, enhancing the Feammox process by optimizing the electron transfer to the ferric iron mineral surface via the use of electron shuttles or different Fe(III) phases/concentrations, might be required to significantly enhance the PFAS defluorination rate by A6, and should be a strategy examined in addition to increasing the A6 biomass.

Preliminary results indicate that PFAS, including PFOA and PFOS, can be mineralized by A6 while oxidizing NH4+ and reducing Fe(III) (Feammox process). Ultimately, the Feammox process may be able to be enhanced in constructed wetlands setting for the removal of NH4+ and in soil column experiments for the removal of TCE. Hence, demonstrating that this process can degrade and possibly mineralize PFOA, PFOS, and PFHxS will point to new in situ PFAS remediation technologies.