CANT-HYD: A curated database of phylogeny-derived Hidden Markov Models for annotation of marker genes involved in hydrocarbon degradation

Discovery of microbial hydrocarbon degradation pathways has traditionally relied on laboratory isolation and characterization of microorganisms. Although many metabolic pathways for hydrocarbon degradation have been discovered, the absence of tools dedicated to their annotation makes it difficult to identify the relevant genes and predict the hydrocarbon degradation potential of microbial genomes and metagenomes. Furthermore, sequence homology between hydrocarbon degradation genes and genes with other functions often results in misannotation. A tool that systematically identifies hydrocarbon metabolic potential is therefore needed. We present the Calgary approach to ANnoTating HYDrocarbon degradation genes (CANT-HYD), a database containing HMMs of 37 marker genes involved in anaerobic and aerobic degradation pathways of aliphatic and aromatic hydrocarbons. Using this database, we show that hydrocarbon metabolic potential is widespread in the tree of life and identify understudied or overlooked hydrocarbon degradation potential in many phyla. We also demonstrate scalability by analyzing large metagenomic datasets for the prediction of hydrocarbon utilization in diverse environments. To the best of our knowledge, CANT-HYD is the first comprehensive tool for robust and accurate identification of marker genes associated with aerobic and anaerobic hydrocarbon degradation.

Marine sediments harbor diverse archaea and bacteria with the potential for anaerobic hydrocarbon degradation via fumarate addition

Marine sediments can contain large amounts of alkanes and methylated aromatic hydrocarbons that are introduced by natural processes or anthropogenic activities. These compounds can be biodegraded by anaerobic microorganisms via enzymatic addition of fumarate. However, the identity and ecological roles of a significant fraction of hydrocarbon degraders containing fumarate-adding enzymes (FAE) in various marine sediments remains unknown. By combining phylogenetic reconstructions, protein homolog modelling, and functional profiling of publicly available metagenomes and genomes, 61 draft bacterial and archaeal genomes encoding anaerobic hydrocarbon degradation via fumarate addition were obtained. Besides Desulfobacterota (previously known as Deltaproteobacteria) that are well-known to catalyze these reactions, Chloroflexi are dominant FAE-encoding bacteria in hydrocarbon-impacted sediments, potentially coupling sulfate reduction or fermentation to anaerobic hydrocarbon degradation. Among Archaea, besides Archaeoglobi previously shown to have this capability, genomes of Heimdallarchaeota, Lokiarchaeota, Thorarchaeota and Thermoplasmata also suggest fermentative hydrocarbon degradation using archaea-type FAE. These bacterial and archaeal hydrocarbon degraders occur in a wide range of marine sediments, including high abundances of FAE-encoding Asgard archaea associated with natural seeps and subseafloor ecosystems. Our results expand the knowledge of diverse archaeal and bacterial lineages engaged in anaerobic degradation of alkanes and methylated aromatic hydrocarbons.

Anaerobic benzene mineralization by natural microbial communities from Niger Delta

The Niger Delta is one of the most damaged ecosystems in the world, mainly due to petroleum contamination by oil exploration accidents. We investigated the natural attenuation potential of Niger Delta subsurface sediment samples for anaerobic hydrocarbon degradation using benzene as a model compound under iron-reducing, sulfate-reducing, and methanogenic conditions. Benzene was slowly mineralized under methanogenic and iron-reducing conditions using nitrilotriacetic acid (NTA)-Fe(III), or poorly crystalline Fe(III) oxyhydroxides as electron acceptors, analyzed by measurement of 13CO2 produced from added 13C-labelled benzene. Highest mineralization rates were observed in microcosms amended with Fe(III) oxyhydroxides. The microbial communities of benzene-mineralizing enrichment cultures were characterized by next-generation sequencing of the genes coding for 16S rRNA and methyl coenzyme M reductase A (mcrA). Abundant phylotypes were affiliated to Betaproteobacteriales, Ignavibacteriales, Desulfuromonadales, and Methanosarcinales of the genera Methanosarcina and Methanothrix, illustrating that the enriched benzene-mineralizing communities were diverse and may contain more than a single benzene degrader. The diversity of the microbial communities was furthermore confirmed by scanning helium-ion microscopy which revealed the presence of various rod-shaped as well as filamentous microbial morphotypes.

Marine Sediments Harbor Diverse Archaea and Bacteria With Potentials for Anaerobic Hydrocarbon Degradation via Fumarate Addition

Background: Marine sediments can contain large amounts of alkanes and methylated aromatic hydrocarbons that are introduced by natural processes or anthropogenic activities. These compounds can be biodegraded by anaerobic microorganisms via enzymatic addition of fumarate. Previous gene- and genome-based surveys have detected ubiquitous and novel fumarate-adding enzymes (FAE), but these were neither confirmed as occurring within full degradation pathways nor affiliated with known organisms. The identity and ecological roles of a significant fraction of anaerobic hydrocarbon degraders in marine sediments therefore remains unknown.Results: By combining phylogenetic reconstructions, protein homolog modelling, and functional profiling of publicly available and newly sequenced metagenomes and genomes, 61 draft bacterial and archaeal genomes encoding anaerobic hydrocarbon degradation via fumarate addition were obtained. Besides Deltaproteobacteria that are well-known to catalyze these reactions, Chloroflexi are dominant FAE-encoding bacteria in hydrocarbon-impacted sediments, potentially coupling sulfate reduction or fermentation to anaerobic hydrocarbon degradation. Among Archaea, besides Archaeoglobi previously shown to have this capability, genomes of Heimdallarchaeota, Lokiarchaeota, Thorarchaeota and Thermoplasmata also suggest fermentative hydrocarbon degradation using archaea-type FAE. The biogeography survey reveals these bacterial and archaeal hydrocarbon degraders occur in a wide range of marine sediments, including high abundances of FAE-encoding Asgard archaea associated with natural seeps and subseafloor ecosystems.Conclusions: Our results expand the knowledge of novel microbial lineages engaged in anaerobic degradation of alkanes and methylated aromatic hydrocarbons, and shed new light on the importance of marine sedimentary archaea in hydrocarbon degradation.

Metagenomic analysis of an anaerobic alkane-degrading microbial culture: potential hydrocarbon-activating pathways and inferred roles of community members

A microbial community (short-chain alkane-degrading culture, SCADC) enriched from an oil sands tailings pond was shown to degrade C6–C10 alkanes under methanogenic conditions. Total genomic DNA from SCADC was subjected to 454 pyrosequencing, Illumina paired-end sequencing, and 16S rRNA amplicon pyrotag sequencing; the latter revealed 320 operational taxonomic units at 5% distance. Metagenomic sequences were subjected to in-house quality control and co-assembly, yielding 984 086 contigs, and annotation using MG-Rast and IMG. Substantial nucleotide and protein recruitment to Methanosaeta concilii, Syntrophus aciditrophicus, and Desulfobulbus propionicus reference genomes suggested the presence of closely related strains in SCADC; other genomes were not well mapped, reflecting the paucity of suitable reference sequences for such communities. Nonetheless, we detected numerous homologues of putative hydrocarbon succinate synthase genes (e.g., assA, bssA, and nmsA) implicated in anaerobic hydrocarbon degradation, suggesting the ability of the SCADC microbial community to initiate methanogenic alkane degradation by addition to fumarate. Annotation of a large contig revealed analogues of the ass operon 1 in the alkane-degrading sulphate-reducing bacterium Desulfatibacillum alkenivorans AK-01. Despite being enriched under methanogenic–fermentative conditions, additional metabolic functions inferred by COG profiling indicated multiple CO2 fixation pathways, organic acid utilization, hydrogenase activity, and sulphate reduction.