Polycyclic Aromatic Hydrocarbon (PAH) Degradation Pathways of the Obligate Marine PAH DegraderCycloclasticussp. Strain P1

ABSTRACTBacteria play an important role in the removal of polycyclic aromatic hydrocarbons (PAHs) from polluted environments. In marine environments,Cycloclasticusis one of the most prevalent PAH-degrading bacterial genera. However, little is known regarding the degradation mechanisms for multiple PAHs byCycloclasticus.Cycloclasticussp. strain P1 was isolated from deep-sea sediments and is known to degrade naphthalene, phenanthrene, pyrene, and other aromatic hydrocarbons. Here, six ring-hydroxylating dioxygenases (RHDs) were identified in the complete genome ofCycloclasticussp. P1 and were confirmed to be involved in PAH degradation by enzymatic assays. Further, five gene clusters in its genome were identified to be responsible for PAH degradation. Degradation pathways for naphthalene, phenanthrene, and pyrene were elucidated inCycloclasticussp. P1 based on genomic and transcriptomic analysis and characterization of an interconnected metabolic network. The metabolic pathway overlaps in many steps in the degradation of pyrene, phenanthrene, and naphthalene, which were validated by the detection of metabolic intermediates in cultures. This study describes a pyrene degradation pathway forCycloclasticus.Moreover, the study represents the integration of a PAH metabolic network that comprises pyrene, phenanthrene, and naphthalene degradation pathways. Taken together, these results provide a comprehensive investigation of PAH metabolism inCycloclasticus.IMPORTANCEPAHs are ubiquitous in the environment and are carcinogenic compounds and tend to accumulate in food chains due to their low bioavailability and poor biodegradability.Cycloclasticusis an obligate marine PAH degrader and is widespread in marine environments, while the PAH degradation pathways remain unclear. In this report, the degradation pathways for naphthalene, phenanthrene, and pyrene were revealed, and an integrated PAH metabolic network covering pyrene, phenanthrene, and naphthalene was constructed inCycloclasticus. This overlapping network provides streamlined processing of PAHs to intermediates and ultimately to complete mineralization. Furthermore, these results provide an additional context for the prevalence ofCycloclasticusin oil-polluted marine environments and pelagic settings. In conclusion, these analyses provide a useful framework for understanding the cellular processes involved in PAH metabolism in an ecologically important marine bacterium.

Draft Genome Sequence of Alcaligenes faecalis BDB4, a Polyaromatic Hydrocarbon-Degrading Bacterium Isolated from Crude Oil-Contaminated Soil

ABSTRACT Alcaligenes fecalis BDB4 was isolated from crude oil-contaminated soil in India. The genome sequence of A. faecalis BDB4 revealed the presence of important genes required for polyaromatic hydrocarbon (PAH) metabolism and other associated functions, such as chemotaxis, membrane transport, and biofilm formation, giving insight into the complete PAH mineralization potential of this bacterium.

Heterologous Expression of Polycyclic Aromatic Hydrocarbon Ring-Hydroxylating Dioxygenase Genes from a Novel Pyrene-Degrading Betaproteobacterium

ABSTRACTA betaproteobacterium within the familyRhodocyclaceaepreviously identified as a pyrene degrader via stable-isotope probing (SIP) of contaminated soil (designated pyrene group 1 or PG1) was cultivated as the dominant member of a mixed bacterial culture. A metagenomic library was constructed, and the largest contigs were analyzed for genes associated with polycyclic aromatic hydrocarbon (PAH) metabolism. Eight pairs of genes with similarity to the α- and β-subunits of ring-hydroxylating dioxygenases (RHDs) associated with aerobic bacterial PAH degradation were identified and linked to PG1 through PCR analyses of a simplified enrichment culture. In tandem with a ferredoxin and reductase found in close proximity to one pair of RHD genes, six of the RHDs were cloned and expressed inEscherichia coli. Each cloned RHD was tested for activity against nine PAHs ranging in size from two to five rings. Despite differences in their predicted protein sequences, each of the six RHDs was capable of transforming phenanthrene and pyrene. Three RHDs could additionally transform naphthalene and fluorene, and these genotypes were also associated with the ability of theE. coliconstructs to convert indole to indigo. Only one of the six cloned RHDs was capable of transforming anthracene and benz[a]anthracene. None of the tested RHDs were capable of significantly transforming fluoranthene, chrysene, or benzo[a]pyrene.

Langerhans Cells Facilitate Epithelial DNA Damage and Squamous Cell Carcinoma

Polyaromatic hydrocarbons (PAHs) are prevalent, potent carcinogens, and 7,12-dimethylbenz[a]anthracene (DMBA) is a model PAH widely used to study tumorigenesis. Mice lacking Langerhans cells (LCs), a signatory epidermal dendritic cell (DC), are protected from cutaneous chemical carcinogenesis, independent of T cell immunity. Investigation of the underlying mechanism revealed that LC-deficient skin was relatively resistant to DMBA-induced DNA damage. LCs efficiently metabolized DMBA to DMBA-trans-3,4-diol, an intermediate proximal to oncogenic Hras mutation, and DMBA-treated LC-deficient skin contained significantly fewer Hras mutations. Moreover, DMBA-trans-3,4-diol application bypassed tumor resistance in LC-deficient mice. Additionally, the genotoxic impact of DMBA on human keratinocytes was significantly increased by prior incubation with human-derived LC. Thus, tissue-associated DC can enhance chemical carcinogenesis via PAH metabolism, highlighting the complex relation between immune cells and carcinogenesis.

Polycyclic Aromatic Hydrocarbon Degradation by Biosurfactant-Producing Pseudomonas sp. IR1

We characterized a newly isolated bacterium, designated as IR1, with respect to its ability to degrade polycyclic aromatic hydrocarbons (PAHs) and to produce biosurfactants. Isolated IR1 was identified as Pseudomonas putida by analysis of 16S rRNA sequences (99.6% homology). It was capable of utilizing two-, three- and four-ring PAHs but not hexadecane and octadecane as a sole carbon and energy source. PCR and DNA hybridization studies showed that enzymes involved in PAH metabolism were related to the naphthalene dioxygenase pathway. Observation of both tensio-active and emulsifying activities indicated that biosurfactants were produced by IR1 during growth on both water miscible and immiscible substrates. The biosurfactants lowered the surface tension of medium from 54.9 dN cm−1 to 35.4 dN cm−1 and formed a stable and compact emulsion with an emulsifying activity of 74% with diesel oil, when grown on dextrose. These findings indicate that this isolate may be useful for bioremediation of sites contaminated with aromatic hydrocarbons.