Biofilm and Planktonic Bacterial and Fungal Communities Transforming High-Molecular-Weight Polycyclic Aromatic Hydrocarbons

Abstract

High-molecular-weight polycyclic aromatic hydrocarbons (HMW-PAHs) are natural components of fossil fuels that are carcinogenic and persistent in the environment, particularly in oil sands process-affected water (OSPW). Their hydrophobicity and tendency to adsorb to organic matter result in low bioavailability and high recalcitrance to degradation. Despite the importance of microbes for environmental remediation, little is known about those involved in HMW-PAH transformations. Here, we investigated the transformation of HMW-PAHs using samples of OSPW and compared the bacterial and fungal community compositions attached to hydrophobic filters and in suspension. It was anticipated that the hydrophobic filters with sorbed HMW-PAHs would select for microbes that specialize in adhesion. Over 33 days, more pyrene was removed (75% ± 11.7%) than the five-ring PAHs benzo[a]pyrene (44% ± 13.6%) and benzo[b]fluoranthene (41% ± 12.6%). For both bacteria and fungi, the addition of PAHs led to a shift in community composition, but thereafter the major factor determining the fungal community composition was whether it was in the planktonic phase or attached to filters. In contrast, the major determinant of the bacterial community composition was the nature of the PAH serving as the carbon source. The main bacteria enriched by HMW-PAHs were Pseudomonas, Bacillus, and Microbacterium species. This report demonstrates that OSPW harbors microbial communities with the capacity to transform HMW-PAHs. Furthermore, the provision of suitable surfaces that encourage PAH sorption and microbial adhesion select for different fungal and bacterial species with the potential for HMW-PAH degradation.

FIG 1

Biotransformation of HMW-PAHs by the microbial communities derived from samples TPW and 2m. Enrichments were grown on Pyr (A), BaP (B), and BbF (C). Error bars represent the standard error of the mean (n = 3). *, significant degradation compared to abiotic control (Mann-Whitney U test, P < 0.05).

FIG 2

Gas chromatograms showing the transformation of the three HMW-PAHs over a 33-day incubation period by the microbial community derived from sample TPW. Enrichments were grown on Pyr (A), BaP (B), and BbF (C). There is a decrease of the Pyr (retention time [RT], 27.60 min), BaP (RT, 38.63 min), and BbF (RT, 37.94 min) peaks and the appearance of secondary and tertiary peaks associated with tentative metabolites.

FIG 3

Production of metabolites tentatively identified during the transformation of Pyr (A), BaP (B), and BbF (C) by microbial communities derived from sample TPW. Error bars represent the standard error of the mean (n = 3).

FIG 4

Cluster analysis illustrating similarities between bacterial and fungal community structures on day 33 of the biotransformation experiment on HMW-PAHs. (A) TPW bacterial community; (B) TPW fungal community; (C) 2m bacterial community; (D) 2m fungal community. P, planktonic community; F, filter community; TPW, TPW community with no added PAH; 2m, 2m community with no added PAH.

FIG 5

Filter and planktonic bacterial communities from sample TPW enriched on BbF and analyzed by 454 pyrosequencing of the 16S rRNA genes. Bacterial genera compromising >3% of the total community from the BbF filter (black bars), BbF planktonic (gray bars), and no-PAH control (white bars) communities are shown. Others, sum of abundances for genera representing <3% of the community composition.

FIG 6

Phylogenetic analysis of selected 16S rRNA gene sequences from isolates, pyrosequencing, and DGGE from the HMW-PAH-degrading communities. For each OTU, the most closely related sequences from GenBank are also indicated. Sequence analysis was performed using the neighbor-joining method. Bootstrap values represent percentages from 100 replicates of the data, and those of >80% are shown by an asterisk. The scale bar indicates 0.01 substitution per nucleotide base. Pyr, pyrene enrichments; BaP, benzo[a]pyrene enrichments; BbF, benzo[b] fluoranthene enrichments; F, filter; P, planktonic.