Rhamnolipid-augmented medium enhances growth of Dehalobium chlorocoercia on chloroethenes and polychlorinated...

Polychlorinated biphenyls (PCBs) and chlorinated ethenes, such as perchloroethene (PCE), are persistent environmental contaminants that pose significant ecological and health risks. Dehalobium chlorocoercia strain DF-1 can reductively dechlorinate both PCEs and PCBs, offering a sustainable remediation strategy. However, its large-scale cultivation has been limited by dependence on an undefined growth-stimulatory factor produced by Pseudodesulfovibrio sp. (DSV). Filtrates of DSV-autoclaved cultures passed through a 10-kDa cutoff filter enhanced growth-linked dechlorination of DF-1, indicating that the active component is a small-molecular-weight, nonsecretory, thermostable compound. Acid precipitation of the autoclaved DSV extract yielded a glycolipid-enriched fraction that also supported DF-1 growth; in parallel, the model biosurfactant surfactin stimulated DF-1 activity, consistent with a biosurfactant-mediated effect. Liquid chromatography coupled to high-resolution tandem mass spectrometry analysis of the acid-precipitated fraction detected ions and fragmentation patterns consistent with multiple rhamnolipid congeners, including mono- and di-rhamnolipids with C8-C12 acyl chains. Both glycolipid-enriched extract and commercial rhamnolipids substituted for DSV extracts, stimulating DF-1 growth-linked reductive dechlorination of PCE, 2,3,4,5-tetrachlorobiphenyl (PCB 61), and 2,2',3,3',4,5,5',6-Octachlorobiphenyl (PCB 199) in a concentration-dependent manner. Maximal DF-1 growth-linked dechlorination rates were observed at submicellar rhamnolipid concentrations, whereas higher concentrations were inhibitory. Rhamnolipid supplementation reduced growth lag and increased DF-1 biomass and dechlorination rates by solubilizing hydrophobic electron acceptors, thereby increasing their bioavailability in the medium. These findings suggest that rhamnolipid-type biosurfactants in Pseudodesulfovibrio sp. extracts have a role in enhancing the growth of DF-1 and demonstrate that commercial rhamnolipid supplementation can replace undefined coculture extracts, enabling reproducible large-scale cultivation of DF-1 for bioaugmentation of PCB- and PCE-contaminated environments.IMPORTANCEPolychlorinated biphenyls (PCBs) and chlorinated ethenes remain widespread contaminants that are difficult to remove with conventional technologies, creating a need for scalable bioremediation strategies. Dehalobium chlorocoercia DF-1 is one of the few isolates available that can reductively dechlorinate both PCBs and PCE, but its application has been hindered by reliance on undefined extracts from a Pseudodesulfovibrio sp. This study shows that the key stimulatory activity is associated with rhamnolipid-type glycolipid biosurfactants released from Pseudodesulfovibrio sp. cells upon lysis. Glycolipid-enriched extracts and defined commercial rhamnolipids enhance DF-1 growth-linked dechlorination of PCE and PCB congeners in a dose-dependent manner, primarily by increasing the bioavailability of hydrophobic electron acceptors. By replacing complex coculture extracts with a defined biosurfactant supplement, these findings remove a major barrier to reproducible, large-scale cultivation of DF-1 and advance the feasibility of bioaugmentation for PCB- and PCE-contaminated environments.