Protein-Ligand Binding and Structural Modelling Studies of Pheromone-Binding Protein-like Sol g 2.1 from Solenopsis geminata Fire Ant Venom.

Sol g 2 is the major protein in Solenopsis geminata fire ant venom. It shares the highest sequence identity with Sol i 2 (S. invicta) and shares high structural homology with LmaPBP (pheromone-binding protein (PBP) from the cockroach Leucophaea maderae). We examined the specific Sol g 2 protein ligands from fire ant venom. The results revealed that the protein naturally formed complexes with hydrocarbons, including decane, undecane, dodecane, and tridecane, in aqueous venom solutions. Decane showed the highest affinity binding (Kd) with the recombinant Sol g 2.1 protein (rSol g 2.1). Surprisingly, the mixture of alkanes exhibited a higher binding affinity with the rSol g 2.1 protein compared to a single one, which is related to molecular docking simulations, revealing allosteric binding sites in the Sol g 2.1 protein model. In the trail-following bioassay, we observed that a mixture of the protein sol g 2.1 and hydrocarbons elicited S. geminata worker ants to follow trails for a longer time and distance compared to a mixture containing only hydrocarbons. This suggests that Sol g 2.1 protein may delay the evaporation of the hydrocarbons. Interestingly, the piperidine alkaloids extracted have the highest attraction to the ants. Therefore, the mixture of hydrocarbons and piperidines had a synergistic effect on the trail-following of ants when both were added to the protein.

A novel strategy for partial purification of alkane hydroxylase from P. chrysogenum SNP5 through reconstituting its native membrane into liposome.

Integral proteins or enzymes are still challenging to purify into their native state because of their need for an amphipathic environment and cofactors. Alkane hydroxylase (AlkB) is a membrane-bound enzyme that catalyzes the hydroxylation of a range of alkanes that have a broad spectrum of applications. In the current study, a novel approach has been explored for partial purification of alkane hydroxylase (AlkB) in its native state through restructuring the lipid bilayer of Penicillium chrysogenum SNP5 into a liposome to extend the native and protective environment to AlkB enzyme. Three different methods i.e., reverse-phase evaporation method (RPEM), detergent-based method (DBM), and ethanol injection method (EIM) have been used for reconstituting its native membrane into liposome. On characterizing liposomes through fluorescence imaging, AFM, and particle size analysis, the reverse-phase evaporation method gave the best results based on the size distribution (i.e., 100-300 nm), the morphology of liposomes, and maximum AlkB specific activity (i.e., 140.68 U/mg). The maximum reconstitution efficiency of 29.48% was observed in RPEM followed by 17.3% in DBM and 12.3% in EIM. On the characterization of the purified AlkB, the molecular weight was measured of 44.6 KDa and the thermostability of liposomes synthesized with the RPEM method was obtained maximum at 55 °C. This approach may open a new strategy for the purification of integral enzymes/proteins in their native state in the field of protein purification and its applications in diversified industries.

Anaerobic hexadecane degradation by a thermophilic Hadarchaeon from Guaymas Basin.

Hadarchaeota inhabit subsurface and hydrothermally heated environments, but previous to this study, they had not been cultured. Based on metagenome-assembled genomes, most Hadarchaeota are heterotrophs that grow on sugars and amino acids, or oxidize carbon monoxide or reduce nitrite to ammonium. A few other metagenome-assembled genomes encode alkyl-coenzyme M reductases (Acrs), ß-oxidation, and Wood-Ljungdahl pathways, pointing toward multicarbon alkane metabolism. To identify the organisms involved in thermophilic oil degradation, we established anaerobic sulfate-reducing hexadecane-degrading cultures from hydrothermally heated sediments of the Guaymas Basin. Cultures at 70°C were enriched in one Hadarchaeon that we propose as Candidatus Cerberiarchaeum oleivorans. Genomic and chemical analyses indicate that Ca. C. oleivorans uses an Acr to activate hexadecane to hexadecyl-coenzyme M. A ß-oxidation pathway and a tetrahydromethanopterin methyl branch Wood-Ljungdahl (mWL) pathway allow the complete oxidation of hexadecane to CO2. Our results suggest a syntrophic lifestyle with sulfate reducers, as Ca. C. oleivorans lacks a sulfate respiration pathway. Comparative genomics show that Acr, mWL, and ß-oxidation are restricted to one family of Hadarchaeota, which we propose as Ca. Cerberiarchaeaceae. Phylogenetic analyses further indicate that the mWL pathway is basal to all Hadarchaeota. By contrast, the carbon monoxide dehydrogenase/acetyl-coenzyme A synthase complex in Ca. Cerberiarchaeaceae was horizontally acquired from Bathyarchaeia. The Acr and ß-oxidation genes of Ca. Cerberiarchaeaceae are highly similar to those of other alkane-oxidizing archaea such as Ca. Methanoliparia and Ca. Helarchaeales. Our results support the use of Acrs in the degradation of petroleum alkanes and suggest a role of Hadarchaeota in oil-rich environments.

Genomic and transcriptomic analyses provide new insights into the allelochemical degradation preference of a novel Acinetobacter strain.

A novel n-alkane- and phenolic acid-degrading Acinetobacter strain (designated C16S1T) was isolated from rhizosphere soil. The strain was identified as a novel species named Acinetobacter suaedae sp. nov. using a polyphasic taxonomic approach. Strain C16S1T showed preferential degradation of three compounds: p-hydroxybenzoate (PHBA) > ferulic acid (FA) > n-hexadecane. In a medium containing two or three of these allelochemicals, coexisting n-hexadecane and PHBA accelerated each other's degradation and that of FA. FA typically hindered the degradation of n-hexadecane but accelerated PHBA degradation. The upregulated expression of n-hexadecane- and PHBA-degrading genes induced, by their related substrates, was mutually enhanced by coexisting PHBA or n-hexadecane; in contrast, expression of both gene types was reduced by FA. Coexisting PHBA or n-hexadecane enhanced the upregulation of FA-degrading genes induced by FA. The expressions of degrading genes affected by coexisting chemicals coincided with the observed degradation efficiencies. Iron shortage limited the degradation efficiency of all three compounds and changed the degradation preference of Acinetobacter. The present study demonstrated that the biodegradability of the chemicals, the effects of coexisting chemicals on the expression of degrading genes and the strain's growth, the shortage of essential elements, and the toxicity of the chemicals were the four major factors affecting the removal rates of the coexisting allelochemicals.

Broad Chain-Length Specificity of the Alkane-Forming Enzymes NoCER1A and NoCER3A/B in Nymphaea odorata.

Many terrestrial plants produce large quantities of alkanes for use in epicuticular wax and the pollen coat. However, their carbon chains must be long to be useful as fuel or as a petrochemical feedstock. Here, we focus on Nymphaea odorata, which produces relatively short alkanes in its anthers. We identified orthologs of the Arabidopsis alkane biosynthesis genes AtCER1 and AtCER3 in N. odorata and designated them NoCER1A, NoCER3A and NoCER3B. Expression analysis of NoCER1A and NoCER3A/B in Arabidopsis cer mutants revealed that the N. odorata enzymes cooperated with the Arabidopsis enzymes and that the NoCER1A produced shorter alkanes than AtCER1, regardless of which CER3 protein it interacted with. These results indicate that AtCER1 frequently uses a C30 substrate, whereas NoCER1A, NoCER3A/B and AtCER3 react with a broad range of substrate chain lengths. The incorporation of shorter alkanes disturbed the formation of wax crystals required for water-repellent activity in stems, suggesting that chain-length specificity is important for surface cleaning. Moreover, cultured tobacco cells expressing NoCER1A and NoCER3A/B effectively produced C19-C23 alkanes, indicating that the introduction of the two enzymes is sufficient to produce alkanes. Taken together, our findings suggest that these N. odorata enzymes may be useful for the biological production of alkanes of specific lengths. 3D modeling revealed that CER1s and CER3s share a similar structure that consists of N- and C-terminal domains, in which their predicted active sites are respectively located. We predicted the complex structure of both enzymes and found a cavity that connects their active sites.

AlmA involved in the long-chain n-alkane degradation pathway in Acinetobacter baylyi ADP1 is a Baeyer-Villiger monooxygenase.

Many Acinetobacter species can grow on n-alkanes of varying lengths (≤C40). AlmA, a unique flavoprotein in these Acinetobacter strains, is the only enzyme proven to be required for the degradation of long-chain (LC) n-alkanes, including C32 and C36 alkanes. Although it is commonly presumed to be a terminal hydroxylase, its role in n-alkane degradation remains elusive. In this study, we conducted physiological, biochemical, and bioinformatics analyses of AlmA to determine its role in n-alkane degradation by Acinetobacter baylyi ADP1. Consistent with previous reports, gene deletion analysis showed that almA was vital for the degradation of LC n-alkanes (C26-C36). Additionally, enzymatic analysis revealed that AlmA catalyzed the conversion of aliphatic 2-ketones (C10-C16) to their corresponding esters, but it did not conduct n-alkane hydroxylation under the same conditions, thus suggesting that AlmA in strain ADP1 possesses Baeyer-Villiger monooxygenase (BVMO) activity. These results were further confirmed by bioinformatics analysis, which revealed that AlmA was closer to functionally identified BVMOs than to hydroxylases. Altogether, the results of our study suggest that LC n-alkane degradation by strain ADP1 possibly follows a novel subterminal oxidation pathway that is distinct from the terminal oxidation pathway followed for short-chain n-alkane degradation. Furthermore, our findings suggest that AlmA catalyzes the third reaction in the LC n-alkane degradation pathway.IMPORTANCEMany microbial studies on n-alkane degradation are focused on the genes involved in short-chain n-alkane (≤C16) degradation; however, reports on the genes involved in long-chain (LC) n-alkane (>C20) degradation are limited. Thus far, only AlmA has been reported to be involved in LC n-alkane degradation by Acinetobacter spp.; however, its role in the n-alkane degradation pathway remains elusive. In this study, we conducted a detailed characterization of AlmA in A. baylyi ADP1 and found that AlmA exhibits Baeyer-Villiger monooxygenase activity, thus indicating the presence of a novel LC n-alkane biodegradation mechanism in strain ADP1.

Continuous Fatty Acid Decarboxylation using an Immobilized Photodecarboxylase in a Membrane Reactor.

The realm of photobiocatalytic alkane biofuel synthesis has burgeoned recently; however, the current dearth of well-established and scalable production methodologies in this domain remains conspicuous. In this investigation, we engineered a modified form of membrane-associated fatty acid photodecarboxylase sourced from Micractinium conductrix (McFAP). This endeavour resulted in creating an innovative assembled photoenzyme-membrane (protein load 5 mg cm-2 ), subsequently integrated into an illuminated flow apparatus to achieve uninterrupted generation of alkane biofuels. Through batch experiments, the photoenzyme-membrane exhibited its prowess in converting fatty acids spanning varying chain lengths (C6-C18). Following this, the membrane-flow mesoscale reactor attained a maximum space-time yield of 1.2 mmol L-1 h-1 (C8) and demonstrated commendable catalytic proficiency across eight consecutive cycles, culminating in a cumulative runtime of eight hours. These findings collectively underscored the photoenzyme-membrane's capability to facilitate the biotransformation of diverse fatty acids, furnishing valuable benchmarks for the conversion of biomass via photobiocatalysis.

Exploring the use of hexadecane by Yarrowia lipolytica: Effect of dissolved oxygen and medium supplementation.

This work aimed to evaluate the effect of medium composition and volumetric oxygen transfer coefficient (kLa) on Y. lipolytica growth and production of microbial lipids and enzymes from hexadecane. In the stirred tank bioreactor, increasing kLa from 11 h-1 to 132 h-1 improved the hexadecane assimilation rate, biomass concentration, and lipids synthesis (0.90 g·L-1). A cost-effective hexadecane-based medium supplemented with corn steep liquor and a low amount of ammonium sulfate boosted lipids production up to 2.1 g·L-1, composed of palmitic, palmitoleic, oleic, and linoleic acids. The unsaturated/saturated fraction was dependent on the C/N ratio. Lipids of Y. lipolytica CBS 2075 are promising feedstock for animal feed, food additives, or the biodiesel industry. Simultaneous synthesis of extracellular lipase and protease from hexadecane was observed, which is a new feature that was not previously reported. The highest enzyme activity was obtained at the highest C/N ratio conditions. These results open new perspectives on the application of Y. lipolytica-based cultures for the biotransformation of hexadecane-polluted streams into valuable compounds, fulfilling an interesting strategy towards the circular economy concept.

Potential biodegradation of polycyclic aromatic hydrocarbons (PAHs) and petroleum hydrocarbons by indigenous fungi recovered from crude oil-contaminated soil in Iran.

A total of 265 fungal individuals were isolated from soils exposed to heavy oil spills in the Yadavaran oil field in Iran to discover indigenous fungal species with a high potential to biodegrade petroleum hydrocarbon pollutants. Morphological and molecular identification of obtained fungal species led to their assignment into 16 genera and 25 species. Alternaria spp. (78%), Fusarium spp. (5%), and Cladosporium spp. (4%) were the most common genera, along with Penicillium spp., Neocamarosporium spp., Epicoccum sp., Kotlabaea sp., Aspergillus sp., Mortierella sp., and Pleurotus sp. A preliminary screening using the DCPIP indicator revealed that approximately 35% of isolates from Alternaria, Epicoccum, Neocamarosporium, Cladosporium, Fusarium, Stachybotrys, Penicillium, and Stemphylium demonstrated promising tolerance to crude oil. The best-performing isolates (12 fungal individuals) were further investigated for their capacity to mineralize a mixture of four polycyclic aromatic hydrocarbons (PAH) for 47 days, quantified by GC-MS. Eventually, two top-performing isolates, namely 5c-12 (Alternaria tenuissima) and 3b-1 (Epicoccum nigrum), were applied to petroleum-contaminated soil. The GC-MS analysis showed that 60 days after inoculation, these isolates successfully degraded more than 70% of the long-chain hydrocarbons in the soil, including C8-C16 n-alkanes, C36 n-alkane, and Pristane. This study introduces two fungal species (5c-12 and 3b-1) with high potential for biodegrading petroleum compounds and PAHs, offering promising prospects for the decontamination of oil-contaminated soil.

Tar patties are hotspots of hydrocarbon turnover and nitrogen fixation during a nearshore pollution event in the oligotrophic southeastern Mediterranean Sea.

Weathered oil, that is, tar, forms hotspots of hydrocarbon degradation by complex biota in marine environment. Here, we used marker gene sequencing and metagenomics to characterize the communities of bacteria, archaea and eukaryotes that colonized tar patties and control samples (wood, plastic), collected in the littoral following an offshore spill in the warm, oligotrophic southeastern Mediterranean Sea (SEMS). We show potential aerobic and anaerobic hydrocarbon catabolism niches on tar interior and exterior, linking carbon, sulfur and nitrogen cycles. Alongside aromatics and larger alkanes, short-chain alkanes appear to fuel dominant populations, both the aerobic clade UBA5335 (Macondimonas), anaerobic Syntropharchaeales, and facultative Mycobacteriales. Most key organisms, including the hydrocarbon degraders and cyanobacteria, have the potential to fix dinitrogen, potentially alleviating the nitrogen limitation of hydrocarbon degradation in the SEMS. We highlight the complexity of these tar-associated communities, where bacteria, archaea and eukaryotes co-exist, likely exchanging metabolites and competing for resources and space.

Structure and Function of Alkane Monooxygenase (AlkB).

ConspectusEvery year, perhaps as much as 800 million tons of hydrocarbons enters the environment; alkanes make up a large percentage of it. Most are transformed by organisms that utilize these molecules as sources of energy and carbon. Both aerobic and anaerobic alkane transformation chemistries exist, capitalizing on the presence of alkanes in both oxic and anoxic environments. Over the past 40 years, tremendous progress has been made in understanding the structure and mechanism of enzymes that catalyze the transformation of methane. By contrast, progress involving enzymes that transform liquid alkanes has been slower with the first structures of AlkB, the predominant aerobic alkane hydroxylase in the environment, appearing in 2023. Because of the fundamental importance of C-H bond activation chemistries, interest in understanding how biology activates and transforms alkanes is high.In this Account, we focus on steps we have taken to understand the mechanism and structure of alkane monooxygenase (AlkB), the metalloenzyme that dominates the transformation of liquid alkanes in the environment (not to be confused with another AlkB that is an α-ketogluturate-dependent enzyme involved in DNA repair). First, we briefly describe what is known about the prevalence of AlkB in the environment and its role in the carbon cycle. Then we review the key findings from our recent high-resolution cryoEM structure of AlkB and highlight important similarities and differences in the structures of members of class III diiron enzymes. Functional studies, which we summarize, from a number of single residue variants enable us to say a great deal about how the structure of AlkB facilitates its function. Next, we overview work from our laboratories using mechanistically diagnostic radical clock substrates to characterize the mechanism of AlkB and contextualize the results we have obtained on AlkB with results we have obtained on other alkane-oxidizing enzymes and explain these results in light of the enzyme's structure. Finally, we integrate recent work in our laboratories with information from prior studies of AlkB, and relevant model systems, to create a holistic picture of the enzyme. We end by pointing to critical questions that still need to be answered, questions about the electronic structure of the active site of the enzyme throughout the reaction cycle and about whether and to what extent the enzyme plays functional roles in biology beyond simply initiating the degradation of alkanes.

HADEG: A curated hydrocarbon aerobic degradation enzymes and genes database.

Databases of genes and enzymes involved in hydrocarbon degradation have been previously reported. However, these databases specialize on only a specific group of hydrocarbons and/or are constructed partly based on enzyme sequences with putative functions indicated by in silico research, with no experimental evidence. Here, we present a curated database of Hydrocarbon Aerobic Degradation Enzymes and Genes (HADEG) containing proteins and genes involved in alkane, alkene, aromatic, and plastic aerobic degradation and biosurfactant production based solely on experimental evidence, which are present in bacteria, and fungi. HADEG includes 259 proteins for petroleum hydrocarbon degradation, 160 for plastic degradation, and 32 for biosurfactant production. This database will help identify and predict hydrocarbon degradation genes/pathways and biosurfactant production in genomes.

Polystyrene-colonizing bacteria are enriched for long-chain alkane degradation pathways.

One of the most promising strategies for the management of plastic waste is microbial biodegradation, but efficient degraders for many types of plastics are still lacking, including those for polystyrene (PS). Genomics has emerged as a powerful tool for mining environmental microbes that may have the ability to degrade different types of plastics. In this study, we use 16S rRNA sequencing to analyze the microbiomes for multiple PS samples collected from sites with different vegetation in Taiwan to reveal potential common properties between species that exhibit growth advantages on PS surfaces. Phylum enrichment analysis identified Cyanobacteria and Deinococcus-Thermus as being the most over-represented groups on PS, and both phyla include species known to reside in extreme environments and could encode unique enzymes that grant them properties suitable for colonization on PS surfaces. Investigation of functional enrichment using reference genomes of PS-enriched species highlighted carbon metabolic pathways, especially those related to hydrocarbon degradation. This is corroborated by the finding that genes encoding long-chain alkane hydroxylases such as AlmA are more prevalent in the genomes of PS-associated bacteria. Our analyses illustrate how plastic in the environment support the colonization by different microbes compared to surrounding soil. In addition, our results point to the possibility that alkane hydroxylases could confer growth advantages of microbes on PS.

Exploring novel alkane-degradation pathways in uncultured bacteria from the North Atlantic Ocean.

IMPORTANCE: Petroleum pollution in the ocean has increased because of rapid population growth and modernization, requiring urgent remediation. Our understanding of the metabolic response of native microbial communities to oil spills is not well understood. Here, we explored the baseline hydrocarbon-degrading communities of a subarctic Atlantic region to uncover the metabolic potential of the bacteria that inhabit the surface and subsurface water. We conducted enrichments with a 13C-labeled hydrocarbon to capture the fraction of the community actively using the hydrocarbon. We then combined this approach with metagenomics to identify the metabolic potential of this hydrocarbon-degrading community. This revealed previously undescribed uncultured bacteria with unique metabolic mechanisms involved in aerobic hydrocarbon degradation, indicating that temperature may be pivotal in structuring hydrocarbon-degrading baseline communities. Our findings highlight gaps in our understanding of the metabolic complexity of hydrocarbon degradation by native marine microbial communities.

Co-culture of Acinetobacter sp. and Scedosporium sp. immobilized beads for optimized biosurfactant production and degradation of crude oil.

The widespread exploration and exploitation of crude oil has increased the prevalence of petroleum hydrocarbon pollution in the marine and coastal environment. Bioremediation of petroleum hydrocarbons using cell immobilization techniques is gaining increasing attention. In this study, the crude oil degradation performance of bacterial and fungal co-culture was optimized by entrapping both cells in sodium-alginate and polyvinyl alcohol composite beads. Results indicate that fungal cells remained active after entrapment and throughout the experiment, while bacterial cells were non-viable at the end of the experimental period in treatments with the bacterial-fungal ratio of 1:2. A remarkable decrease in surface tension from 72 mN/m to 36.51 mN/m was achieved in treatments with the bacterial-fungal ratio of 3:1. This resulted in a significant (P < 0.05) total petroleum hydrocarbon (TPH) removal rate of 89.4%, and the highest degradation of n-alkanes fractions (from 2129.01 mg/L to 118.53 mg/L), compared to the other treatments. Whereas PAHs removal was highest in treatments with the most fungal abundance (from 980.96 µg/L to 177.3 µg/L). Furthermore, enzymes analysis test revealed that catalase had the most effect on microbial degradation of the target substrate, while protease had no significant impact on the degradation process. High expression of almA and PAH-RHDa genes was achieved in the co-culture treatments, which correlated significantly (P < 0.05) with n-alkanes and PAHs removal, respectively. These results indicate that the application of immobilized bacterial and fungal cells in defined co-culture systems is an effective strategy for enhanced biodegradation of petroleum hydrocarbons in aqueous systems.

Alcanivorax borkumensis biofilms enhance oil degradation by interfacial tubulation.

During the consumption of alkanes, Alcanivorax borkumensis will form a biofilm around an oil droplet, but the role this plays during degradation remains unclear. We identified a shift in biofilm morphology that depends on adaptation to oil consumption: Longer exposure leads to the appearance of dendritic biofilms optimized for oil consumption effected through tubulation of the interface. In situ microfluidic tracking enabled us to correlate tubulation to localized defects in the interfacial cell ordering. We demonstrate control over droplet deformation by using confinement to position defects, inducing dimpling in the droplets. We developed a model that elucidates biofilm morphology, linking tubulation to decreased interfacial tension and increased cell hydrophobicity.

Distribution and abundance of oil-degrading bacteria in seawater of the Yellow Sea and Bohai Sea, China.

Petroleum hydrocarbons are widespread in seawater. As an important sea area in northern China, the content and distribution of petroleum hydrocarbons in seawater need our attention because of the high toxicity and lasting polluting effects on the ecological environment of the Yellow Sea and Bohai Sea. In addition, there are few reports comparing the diversity of oil-degrading bacteria before and after enrichment. Therefore, we collected surface seawater from 10 sites in the Yellow Sea and Bohai Sea in the autumn of 2020 to study the distribution characteristics of total petroleum hydrocarbons (TPH) and the diversity of oil-degrading bacteria. The concentration of TPH was 81.65 µg/L-139.55 µg/L at ten sites in the Bohai Sea and the Yellow Sea, which conformed to the China Grade II water quality standard (GB3097-1997). Moreover, the pristine/phytane (PR/PH) value of most sites was close to 1, indicating that the area was obviously polluted by exogenous petroleum hydrocarbons. We found that oil-degrading bacteria in the seawater of the Yellow Sea and the Bohai Sea had a good degradation effect on C11-C14 short chain alkanes (degradation rate of 59.19-73.22 %) and C1-C4 phenanthrene (degradation rate of 48.19-60.74 %). In terms of the diversity of oil-degrading bacteria, Gammaproteobacteria and Alphaproteobacteria dominated the enriched bacterial communities. Notably, the relative abundance of Alcanivorax changed significantly before and after enrichment. We proposed that surface seawater in the Bohai Sea and Yellow Sea could form oil-degrading bacteria mainly composed of Alcanivorax, which had great potential for oil pollution remediation.

Biodegradation of saturate fraction of crude oil and production of signature carboxylic acids.

Bacteria degrading large portion of saturated hydrocarbons are important for crude oil bioremediation. This study investigates Novosphingobium sp. S1, Gordonia amicalis S2 and Gordonia terrae S5 capability of degrading wide range of saturated hydrocarbons from Congo Bilondo crude oil and discusses the degradation pathway. A parallel analytical approach combining GC-MS and LC-HRMS enabled characterization of saturated hydrocarbons and comprehensive determination of carboxylic acid metabolites produced during biodegradation, respectively. Results showed that the three strains could efficiently degrade the n-alkanes (C10-C28) as well as methyl-substituted alkanes (C11-C26). The series of mono-, hydroxy- and dicarboxylic acids identified in this study confirmed the active biodegradation of the saturate fraction and suggest their degradation was via the bi-terminal oxidation pathway. This is the first study linking these bacterial species to bi-terminal oxidation of the saturated hydrocarbons. The study highlights the potential application of the bacterial strains in the bioremediation of crude oil contaminated sites. Additionally, while carboxylic acids is indicated as a suitable and valuable metabolic biomarker, its application is considered feasible and cost effective for rapid monitoring and evaluation of hydrocarbon biodegradation.

Genome analysis of a coral-associated bacterial consortium highlights complementary hydrocarbon degradation ability and other beneficial mechanisms for the host.

Here we report the oil degradation genetic potential of six oil-degrading bacteria (ODB), previously used as a bioremediation consortium, isolated from the hydrocoral Millepora alcicornis and seawater. The strains were identified as Halomonas sp. (LC_1), Cobetia sp. (LC_6), Pseudoalteromonas shioyasakiensis (LC_2), Halopseudomonas aestusnigri (LC_3), Shewanella algae (LC_4), and Brucella intermedia (LC_5). The taxonomic identification differed from that of the original paper when we used whole genome gene markers instead of just 16S rRNA gene. Genes responsible for the degradation of aromatic hydrocarbons and n-alkanes were found in all genomes, although different (and complementary) steps of the metabolic pathways were unique to each strain. Genes for naphthalene and toluene degradation were found in various strains. We annotated quinate degradation genes in LC_6, while LC_3 and LC_5 presented genes for biosurfactant and rhamnolipid biosynthesis. We also annotated genes related to beneficial mechanisms for corals, such as genes involved in nitrogen and DMSP metabolism, cobalamin biosynthesis and antimicrobial compounds production. Our findings reinforce the importance of using bacterial consortia for bioremediation approaches instead of single strains, due to their complementary genomic arsenals. We also propose a genome-based framework to select complementary ODB that can provide additional benefits to coral health.

Microaerobic degradation of crude oil and long chain alkanes by a new Rhodococcus strain from Gulf of Mexico.

Bacterial degradation of crude oil is a promising strategy for reducing the concentration of hydrocarbons in contaminated environments. In the first part of this study, we report the enrichment of two bacterial consortia from deep sediments of the Gulf of Mexico with crude oil as the sole carbon and energy source. We conducted a comparative analysis of the bacterial community in the original sediment, assessing its diversity, and compared it to the enrichment observed after exposure to crude oil in defined cultures. The consortium exhibiting the highest hydrocarbon degradation was predominantly enriched with Rhodococcus (75%). Bacterial community analysis revealed the presence of other hydrocarbonoclastic members in both consortia. In the second part, we report the isolation of the strain Rhodococcus sp. GOMB7 with crude oil as a unique carbon source under microaerobic conditions and its characterization. This strain demonstrated the ability to degrade long-chain alkanes, including eicosane, tetracosane, and octacosane. We named this new strain Rhodococcus qingshengii GOMB7. Genome analysis revealed the presence of several genes related to aromatic compound degradation, such as benA, benB, benC, catA, catB, and catC; and five alkB genes related to alkane degradation. Although members of the genus Rhodococcus are well known for their great metabolic versatility, including the aerobic degradation of recalcitrant organic compounds such as petroleum hydrocarbons, this is the first report of a novel strain of Rhodococcus capable of degrading long-chain alkanes under microaerobic conditions. The potential of R. qingshengii GOMB7 for applications in bioreactors or controlled systems with low oxygen levels offers an energy-efficient approach for treating crude oil-contaminated water and sediments.