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The marine bacterium Alcanivorax borkumensis produces a surface-active glycine-glucolipid during growth with long-chain alkanes. A high-performance liquid chromatography (HPLC) method was developed for absolute quantification. This method is based on the conversion of the glycine-glucolipid to phenacyl esters with subsequent measurement by HPLC with diode array detection (HPLC-DAD). Different molecular species were separated by HPLC and identified as glucosyl-tetra(3-hydroxy-acyl)-glycine with varying numbers of 3-hydroxy-decanoic acid or 3-hydroxy-octanoic acid groups via mass spectrometry. The growth rate of A. borkumensis cells with pyruvate as the sole carbon source was elevated compared to hexadecane as recorded by the increase in cell density as well as oxygen/carbon dioxide transfer rates. The amount of the glycine-glucolipid produced per cell during growth on hexadecane was higher compared with growth on pyruvate. The glycine-glucolipid from pyruvate-grown cells contained considerable amounts of 3-hydroxy-octanoic acid, in contrast to hexadecane-grown cells, which almost exclusively incorporated 3-hydroxy-decanoic acid into the glycine-glucolipid. The predominant proportion of the glycine-glucolipid was found in the cell pellet, while only minute amounts were present in the cell-free supernatant. The glycine-glucolipid isolated from the bacterial cell broth, cell pellet, or cell-free supernatant showed the same structure containing a glycine residue, in contrast to previous reports, which suggested that a glycine-free form of the glucolipid exists which is secreted into the supernatant. In conclusion, the glycine-glucolipid of A. borkumensis is resident to the cell wall and enables the bacterium to bind and solubilize alkanes at the lipid-water interface. IMPORTANCE Alcanivorax borkumensis is one of the most abundant marine bacteria found in areas of oil spills, where it degrades alkanes. The production of a glycine-glucolipid is considered an essential element for alkane degradation. We developed a quantitative method and determined the structure of the A. borkumensis glycine-glucolipid in different fractions of the cultures after growth in various media. Our results show that the amount of the glycine-glucolipid in the cells by far exceeds the amount measured in the supernatant, confirming the proposed cell wall localization. These results support the scenario that the surface hydrophobicity of A. borkumensis cells increases by producing the glycine-glucolipid, allowing the cells to attach to the alkane-water interface and form a biofilm. We found no evidence for a glycine-free form of the glucolipid.

To this date, the extent of microbial biodegradation of plastics is still an open question. The presence of hydrocarbon degraders as part of the plastisphere identified from marine plastic debris (MPD) samples has prompted different ideas about the role of such a group of microorganisms in the degradation of these synthetic polymers. In this investigation, we have tested the potential of the hydrocarbonoclastic bacteria Alcanivorax sp. 24 (bacterial isolate obtained from MPD) to degrade polyethelene (PE) –one of the most recalcitrant carbon-based synthetic materials produced and, currently, the most ubiquitous plastic pollutant found in nature. Evidence indicates that an array of abiotic and biotic processes eventually breaks down PE. However, the biological impact and mechanistic understanding of the process are unclear. Here, using high-throughput proteomics, we investigated the molecular processes that take place in the hydrocarbon-degrading marine bacterium Alcanivorax sp. 24 when grown in the presence of low-density PE (LDPE). As well as efficiently utilising and assimilating the leachate of weathered LDPE, the bacterium was able to reduce the molecular weight distribution and overall mass of pristine LDPE films (0.9 % after 34 days of incubation). Most interestingly, Alcanivorax acquired the isotopic signature of the pristine plastic and induced an extensive array of metabolic pathways for aliphatic compound degradation. Presumably, the primary biodegradation of LDPE by Alcanivorax sp. 24 is possible via the production of extracellular reactive oxygen species as observed both by the material's surface oxidation and the measurement of superoxide in the culture with LDPE. Our findings confirm that hydrocarbon-biodegrading bacteria within the plastisphere may in fact have a role in degrading PE. Also see: https://micro2022.sciencesconf.org/427350/document, In MICRO 2022, Online Atlas Edition: Plastic Pollution from MACRO to nano

Genome of Alcanivorax sp. IO_7, an alkane degrading deep-sea bacteria isolated from hydrothermally-influenced Southwest Indian Ridge was sequenced and analysed. Genomic data mining revealed gene clusters for degrading n-alkane and cycloalkanes, including biosurfactant production. The strain was shown to grow on hexadecane as its sole carbon source, supporting the findings of genomic analysis. Presence of cyclohexanone monooxygenase among genomic islands suggest that this strain may have used gene transfer to enhance its hydrocarbon degradation ability. Genes encoding for heavy metal resistance, multidrug resistance and multiple natural product biosynthesis crucial for survival in the hydrothermally influenced deep sea environment were detected. In our comparative genome analysis, it was evident that marine Alcanivorax strains contain a suite of enzymes for n-alkane and haloalkanoate degradation. Comparative genome and genomic synteny analysis provided insights into the physiological features and adaptation strategies of Alcanivorax sp. IO_7 in the deep-sea hydrothermal environment.

Petroleum degrading enzymes can be used as an alternative way to improve petroleum bioremediation approaches. Alcanivorax borkumensis is an alkane-degrading bacteria that can produce petroleum degrading enzymes such as alkane hydroxylase and lipase. In this study, pilot-scale Alcanivorax borkumensis fermentation was developed for producing large volumes of petroleum degrading enzymes cocktail (∼900 L). Different process conditions, such as inoculum age 72 h and size 4% v/v, temperature 30 ± 1 °C, agitation speed at 150 rpm and, fermentation period 3 days were determined as the optimum for producing alkane hydroxylase and lipase activity. The oxygen transfer capacity was studied for obtaining better bacterial growth and higher enzyme activities in bioreactor process optimization as well as scale-up. Results showed that the maximum values of oxygen mass transfer coefficient (k

Understanding microbial responses to hydrocarbon and plastic pollution are crucial for limiting the detrimental impacts of environmental contaminants on marine ecosystems. Herein, we reported a new Alcanivorax species isolated from the North Atlantic Ocean capable of degrading alkanes and polyhydroxybutyrate (PHB) plastic (one of the emerging bioplastics that may capture the future plastic market). The whole-genome sequencing showed that the species harbors three types of alkane 1-monooxygenases (AlkB) and one PHB depolymerase (PhaZ) to initiate the degradation of alkanes and plastics. Growth profiling demonstrated that n-pentadecane (C15, the main alkane in the marine environment due to cyanobacterial production other than oil spills) and PHB could serve as preferential carbon sources. However, the cell membrane composition, PhaZ activity, and expression of three alkB genes were utterly different when grown on C15 and PHB. Further, Alcanivorax was a well-recognized alkane-degrader that participated in the ocean hydrocarbon cycles linking with hydrocarbon production and removal. Our discovery supported that the existing biogeochemical processes may add to the marine ecosystem's resilience to the impacts of plastics.

Marine microbial communities play an important role in biodegradation of subsurface plumes of oil that form after oil is accidentally released from a seafloor wellhead. The response of these mesopelagic microbial communities to the application of chemical dispersants following oil spills remains a debated topic. While there is evidence that contrasting results in some previous work may be due to differences in dosage between studies, the impacts of these differences on mesopelagic microbial community composition remains unconstrained. To answer this open question, we exposed a mesopelagic microbial community from the Gulf of Mexico to oil alone, three concentrations of oil dispersed with Corexit 9500, and three concentrations of Corexit 9500 alone over long periods of time. We analyzed changes in hydrocarbon chemistry, cell abundance, and microbial community composition at zero, three and six weeks. The lowest concentration of dispersed oil yielded hydrocarbon concentrations lower than oil alone and microbial community composition more similar to control seawater than any other treatments with oil or dispersant. Higher concentrations of dispersed oil resulted in higher concentrations of microbe-oil microaggregates and similar microbial composition to the oil alone treatment. The genus Colwellia was more abundant when exposed to multiple concentrations of dispersed oil, but not when exposed to dispersant alone. Conversely, the most abundant Marinobacter amplicon sequence variant (ASV) was not influenced by dispersant when oil was present and showed an inverse relationship to the summed abundance of Alcanivorax ASVs. As a whole, the data presented here show that the concentration of oil strongly impacts microbial community response, more so than the presence of dispersant, confirming the importance of the concentrations of both oil and dispersant in considering the design and interpretation of results for oil spill simulation experiments.

Water column and sediment samples were collected in the southern Gulf of Mexico (GoMex) during 3 oceanographic cruises: XIXIMI-04 (September 2015), XIXIMI-05 (June 2016), and XIXIMI-06 (August 2017). DNA that was extracted from the samples was analyzed by qPCR to detect and quantify bacterial groups that have been reported to metabolize alkanes (Alcanivorax) and aromatic hydrocarbons (Cycloclasticus) and are involved in methane production (Methanomicrobiales). The results were then analyzed with regard to the water masses that are currently detected in the GoMex. Generally, we observed a decrease in the proportion of Alcanivorax and a rise in those of Cycloclasticus and Methanomicrobiales in samples from the surface to deep waters and in sediment samples. Scatterplots of the results showed that the relative abundance of the 3 groups was higher primarily from the surface to 1000 m, but the levels of Cycloclasticus and Methanomicrobiales were high in certain water samples below 1000 m and in sediments. In conclusion, oil-degrading bacteria are distributed widely from the surface to deep waters and sediments throughout the southern GoMex, representing a potential inoculum of bacteria for various hydrocarbon fractions that are ready for proliferation and degradation in the event of an oil spill from the seafloor or along the water column.

A Gram-stain-negative, rod-shaped, non-motile, strictly aerobic strain, designated as MTEO17T, was isolated from a 1000 m deep seawater sample of the Mariana Trench. Growth was observed at 10–45 °C (optimum, 37 °C), in the presence of 0.0–12.0 % NaCl (w/v; optimum, 3.0 %) and at pH 6.0–10.0 (optimum, pH 7.0–8.0). Phylogenetic analysis, based on the 16S rRNA gene sequence, revealed that strain MTEO17T belonged to the genus Alcanivorax and showed the highest sequence similarity of 97.9 % to Alcanivorax nanhaiticus MCCC 1A05629T. The estimated average nucleotide identity and DNA–DNA hybridization values between strain MTEO17T and A. nanhaiticus MCCC 1A05629T were 78.98 and 23.80 %, respectively. The significant dominant fatty acids were C16 : 0, summed feature 8 (C18 : 1ω6c and/or C18 : 1ω7c) and summed feature 3 (C16 : 1ω6c and/or C16 : 1ω7c). The polar lipids comprised two phosphatidylethanolamines, one phosphatidylglycerol, one unidentified phospholipid and four unidentified polar lipids. The DNA G+C content of strain MTEO17T was 57.5 %. On the basis of the polyphasic evidence, strain MTEO17T is proposed to represent a novel species of the genus Alcanivorax , for which the name Alcanivorax profundi sp. nov. is proposed. The type strain is MTEO17T (=KCTC 52694T=MCCC 1K03252T).

Alcanivorax borkumensis is a ubiquitous marine bacterium that utilizes alkanes as a sole carbon source. We observed two phenotypes in the A. borkumensis SK2 type strain: rough (R) and smooth (S) types. The S type exhibited lower motility and higher polysaccharide production than the R type. Full genome sequencing revealed a mutation in the S type involved in cyclic-di-GMP production. The present results suggest that higher c-di-GMP levels in the S type control the biofilm forming behavior of this bacterium in a manner commensurate with other Gram-negative bacteria.

Though many studies pertaining to soil bioremediation have been performed to study the microbial kinetics in shake flasks, the process efficiency in column tests is seldom. In the present study, soil columns tests were carried out to study the biodegradation of soil contaminated with a high concentration of diesel (≈19.5 g/kg) petroleum hydrocarbons expressed as C

In Antarctic regions, the composition and metabolic activity of microbial assemblages associated with plastic debris (“plastisphere”) are almost unknown. A macroplastic item from land (MaL, 30 cm) and a mesoplastic from the sea (MeS, 4 mm) were collected in Maxwell Bay (King George Island, South Shetland) and analyzed by Fourier transform infrared spectroscopy in attenuated total reflectance geometry (FTIR-ATR), which confirmed a polystyrene foam and a composite high-density polyethylene composition for MaL and MeS, respectively. The structure and function of the two plastic-associated prokaryotic communities were studied by complementary 16S ribosomal RNA gene clone libraries, total bacterioplankton and culturable heterotrophic bacterial counts, enzymatic activities of the whole community and enzymatic profiles of bacterial isolates. Results showed that Gamma- and Betaproteobacteria (31% and 28%, respectively) dominated in MeS, while Beta- and Alphaproteobacteria (21% and 13%, respectively) in MaL. Sequences related to oil degrading bacteria (Alcanivorax,Marinobacter) confirmed the known anthropogenic pressure in King George Island. This investigation on plastic-associated prokaryotic structure and function represents the first attempt to characterize the ecological role of plastisphere in this Antarctic region and provides the necessary background for future research on the significance of polymer type, surface characteristics and environmental conditions in shaping the plastisphere.

The environmental hazards of oil spills cannot be underestimated. Bioremediation holds promise among various approaches to tackle oil spills in soils and sediments. In particular, using oil-degrading bacteria is an efficient and self-regulating way to remove oil spills. Using animals for oil spills remediation is in its infancy, mostly due to the lack of efficient oil-degrading capabilities in eukaryotes. Here we show that Caenorhabditis elegans nematodes survive for extended periods (up to 22 days) on pure crude oil diet. Moreover, we report for the first time the use of Alcanivorax borkumensis hydrocarbonoclastic bacteria for colonisation of C. elegans intestines, which allows for effective digestion of crude oil by the nematodes. The worms fed and colonised by A. borkumensis demonstrated the similar or even better longevity, resistance against oxidative and thermal stress and reproductivity as those animals fed with Escherichia coli bacteria (normal food). Importantly, A. borkumensis-carrying nematodes were able to accumulate oil droplet from oil-contaminated soils. Artificial colonisation of soil invertebrates with oil-degrading bacteria will be an efficient way to distribute microorganisms in polluted soil, thus opening new avenues for oil spills zooremediation.

Two novel Alcanivorax-related strains, designated ST75FaO-1T and 521-1, were isolated from the seawater of the South China Sea and the deep-sea sediment of the West Pacific Ocean, respectively. Both strains are Gram-stain-negative, rod-shaped, and non-motile, and grow at 10–40 °C, pH 5.0–10.0, in the presence of 1.0–15.0% (w/v) NaCl. Their 16S rRNA gene sequences showed 99.9% similarity. Phylogenetic analysis based on the 16S rRNA gene sequences indicated that both strains belong to the genus Alcanivorax, and share 92.9–98.1% sequence similarity with all valid type strains of this genus, with the highest similarity being to type strain Alcanivorax venustensis DSM 13974T (98.0–98.1%). Digital DNA–DNA hybridization (dDDH) and average nucleotide identity values between strains ST75FaO-1T and 521-1 were 75.7% and 97.1%, respectively, while the corresponding values with A. venustensis DSM 13974T were only 25.4–25.6% and 82.4–82.7%, respectively. The two strains contained similar major cellular fatty acids including C16:0, C18:1 ω7c/ω6c, C19:0 cyclo ω8c, C16:1 ω7c/ω6c, C12:0 3-OH, and C12:0. The genomic G + C content of strains ST75FaO-1T and 521-1 were 66.3% and 66.1%, respectively. Phosphatidylglycerol, phosphatidylethanolamine, two unidentified phospholipids, and one unidentified polar lipid were present in both strains. On the basis of phenotypic and genotypic characteristics, the two strains represent a novel species within the genus Alcanivorax, for which the name Alcanivorax profundimaris sp. nov. is proposed. The type strain is ST75FaO-1T (= MCCC 1A17714T = KCTC 82142T).

A new Alcanivorax sp. VBW004 was isolated from a shallow hydrothermal vent in Azores Island, Portugal. In this study, we determined VBW004 was resistant to copper. This strain showed maximum tolerance of copper concentrations up to 600 μg/mL. Based on 16S rRNA gene sequencing and phylogeny revealed that this strain was more closely related to Alcanivorax borkumensis SK2. We sequenced the genome of this strain that consist of 3.8 Mb size with a G + C content of 58.4 %. In addition, digital DNA-DNA hybridizations (dDDH) and the average nucleotide identities (ANI) analysis between Alcanivorax borkumensis SK2 and Alcanivorax jadensis T9 revealed that Alcanivorax sp. VBW004 belongs to new species. Functional annotation revealed that the genome acquired multiple copper resistance encoding genes that could assist VBW004 to respond to high Cu toxicity. Our results from biosorption analysis presumed that the VBW004 is an ecologically important bacterium that could be useful for copper bioremediation.

The environmental accumulation of plastics worldwide is a consequence of the durability of the material. Alternative polymers, marketed as biodegradable, present a potential solution to mitigate their ecological damage. However, understanding of biodegradability has been hindered by a lack of reproducible testing methods. We developed a novel method to evaluate the biodegradability of plastic samples based on the monitoring of bacterial respiration in aqueous media via the quantification of CO2 produced, where the only carbon source available is from the polymer. Rhodococcus rhodochrous and Alcanivorax borkumensis were used as model organisms for soil and marine systems, respectively. Our results demonstrate that this approach is reproducible and can be used with a variety of plastics, allowing comparison of the relative biodegradability of the different materials. In the case of low-density polyethylene, the study demonstrated a clear correlation between the molecular weight of the sample and CO2 released, taken as a measure of biodegradability.

Thematic Issue on Metal(loid) Microbiology., Pristine marine environments are highly oligotrophic ecosystems populated by well‐established specialized microbial communities. Nevertheless, during oil spills, low‐abundant hydrocarbonoclastic bacteria bloom and rapidly prevail over the marine microbiota. The genus Alcanivorax is one of the most abundant and well‐studied organisms for oil degradation. While highly successful under polluted conditions due to its specialized oil‐degrading metabolism, it is unknown how they persist in these environments during pristine conditions. Here, we show that part of the Alcanivorax genus, as well as oils, has an enormous potential for biodegrading aliphatic polyesters thanks to a unique and abundantly secreted alpha/beta hydrolase. The heterologous overexpression of this esterase proved a remarkable ability to hydrolyse both natural and synthetic polyesters. Our findings contribute to (i) better understand the ecology of Alcanivorax in its natural environment, where natural polyesters such as polyhydroxyalkanoates (PHA) are produced by a large fraction of the community and, hence, an accessible source of carbon and energy used by the organism in order to persist, (ii) highlight the potential of Alcanivorax to clear marine environments from polyester materials of anthropogenic origin as well as oils, and (iii) the discovery of a new versatile esterase with a high biotechnological potential., VZ was supported by CONICYT‐BECAS CHILE/Doctorado Becas Chile en el Extranjero, Folio 72160583. JAC‐O was supported by the NERC Independent Research Fellowship NE/K009044/1, NERC research project NE/S005501/1, and Ramón y Cajal contract RYC‐2017‐22452 (funded by the Ministry of Science, Innovation and Universities, the National Agency of Research, and the European Social Fund). RB was supported by the MINECO project CTM2015‐70180‐R (FEDER co‐funding).

Major challenges hampering biotechnological applications of esterases include the requirement to accept nonnatural and chemically demanding substrates and the tolerance of the enzymes toward organic solvents which are often required to solubilize such substrates. We describe here a high-throughput screening strategy to identify novel organic-solvent-tolerant carboxylic ester hydrolases (CEs). Among these enzymes, CEs active against water-insoluble bulky substrates were identified. Our results thus contribute to fostering the identification and biotechnological application of CEs., Biocatalysis has emerged as an important tool in synthetic organic chemistry enabling the chemical industry to execute reactions with high regio- or enantioselectivity and under usually mild reaction conditions while avoiding toxic waste. Target substrates and products of reactions catalyzed by carboxylic ester hydrolases are often poorly water soluble and require organic solvents, whereas enzymes are evolved by nature to be active in cells, i.e., in aqueous rather than organic solvents. Therefore, biocatalysts that withstand organic solvents are urgently needed. Current strategies to identify such enzymes rely on laborious tests carried out by incubation in different organic solvents and determination of residual activity. Here, we describe a simple assay useful for screening large libraries of carboxylic ester hydrolases for resistance and activity in water-miscible organic solvents. We have screened a set of 26 enzymes, most of them identified in this study, with four different water-miscible organic solvents. The triglyceride tributyrin was used as a substrate, and fatty acids released by enzymatic hydrolysis were detected by a pH shift indicated by the indicator dye nitrazine yellow. With this strategy, we succeeded in identifying a novel highly organic-solvent-tolerant esterase from Pseudomonas aestusnigri. In addition, the newly identified enzymes were tested with sterically demanding substrates, which are common in pharmaceutical intermediates, and two enzymes from Alcanivorax borkumensis were identified which outcompeted the gold standard ester hydrolase CalB from Candida antarctica. IMPORTANCE Major challenges hampering biotechnological applications of esterases include the requirement to accept nonnatural and chemically demanding substrates and the tolerance of the enzymes toward organic solvents which are often required to solubilize such substrates. We describe here a high-throughput screening strategy to identify novel organic-solvent-tolerant carboxylic ester hydrolases (CEs). Among these enzymes, CEs active against water-insoluble bulky substrates were identified. Our results thus contribute to fostering the identification and biotechnological application of CEs.

Alcanivorax borkumensis is a bacterial community that dominates hydrocarbon-degrading communities around many oil spills. The physicochemical conditions that prompt bacterial binding to oil/water interfaces are not well understood. To provide key insights into this process, A. borkumensis cells were cultured either in a clean environment condition (dissolved organic carbon) or in an oil spill condition (hexadecane as the sole energy source). The ability of these bacteria to bind to the oil/water interface was monitored through interfacial tension measurements, bacterial cell hydrophobicity, and fluorescence microscopy. Our experiments show that A. borkumensis cells cultured in clean environment conditions remain hydrophilic and do not show significant transport or binding to the oil/water interface. In sharp contrast, bacteria cultured in oil spill conditions become partially hydrophobic and their amphiphilicity drives them to oil/water interfaces, where they reduce interfacial tension and form the early stages of a biofilm. We show that it is A. borkumensis cells that attach to the oil/water interface and not a synthesized biosurfactant that is released into solution that reduces interfacial tension. This study provides key insights into the physicochemical properties that allow A. borkumensis to adhere to oil/water interfaces.

Strain GI5T was isolated from a surface seawater sample collected from Garorim Bay (West Sea, Republic of Korea). The isolated strain was aerobic, Gram-stain-negative, rod-shaped, motile by means of a polar flagellum, negative for catalase and weakly positive for oxidase. The optimum growth pH, salinity and temperature were determined to be pH 7.5–8.0, 3 % NaCl (w/v) and 25 °C, respectively; the growth ranges were pH 6.0–9.0, 1–7 % NaCl (w/v) and 18–40 °C. The results of phylogenetic analysis of 16S rRNA gene sequences indicated that GI5T clustered within the family Alcanivoracaceae , and most closely with Alcanivorax dieselolei B-5T and Alcanivorax marinus R8-12T (91.9 % and 91.6 % similarity, respectively). The major cellular fatty acids in GI5T were C18 : 1ω7c/C18 : 1ω6c (44.45 %), C16 : 1ω6c/C16 : 1ω7c (14.17 %) and C16 : 0 (10.19 %); this profile was distinct from those of the closely related species. The major respiratory quinone of GI5T was Q-8. The main polar lipids were phosphatidylethanolamine and phosphatidylglycerol. Two putative alkane hydroxylase (alkB) genes were identified in GI5T. The G+C content of the genomic DNA of GI5T was determined to be 51.2 mol%. On the basis of the results of phenotypic, chemotaxonomic and phylogenetic studies, strain GI5T represents a novel species of a novel genus of the family Alcanivoracaceae, for which we propose the name Ketobacter alkanivorans gen. nov., sp. nov.; the type strain is GI5T (=KCTC 52659T=JCM 31835T).

A novel heterotrophic, Gram-stain-negative, aerobic, rod-shaped, pale yellow, non-motile and non-spore-forming bacterium, designated strain F2T, was isolated from marine sediment collected from the Weihai coast, Shandong Province, PR China. Optimal growth occurred at 33 °C (range, 10–37 °C), with 3.0–4.0 % (w/v) NaCl (1.0–8.0 %) and at pH 7.5–8.0 (pH 6.5–9.0). Q-8 was the sole respiratory quinone. The major polar lipids of strain F2T were phosphatidylmonomethylethanolamine, phosphatidylcholine, phosphatidylglycerol, phosphatidylethanolamine, two unidentified phospholipids and two unidentified polar lipids. The major cellular fatty acid in strain F2T was iso-C15 : 0. The genomic DNA G+C content of the strain was 48.1 mol%. Phylogenetic analysis based on 16S rRNA gene sequencing revealed that strain F2T is most closely related to Marinicella litoralis JCM 16154T (97.5 %) and Marinicella pacifica sw153T (96.0 %). Based on the results of our polyphasic analysis, we conclude that strain F2T represents a novel species of the genus Marinicella , for which the name Marinicella sediminis sp. nov. is proposed. The type strain of the new species is F2T (=KCTC 42953T=MCCC 1H00149T).

This study investigates the production of alkane hydroxylase, lipase and esterase by the marine hydrocarbon degrading bacteria Alcanivorax borkumensis. The focus of this study is the remediation of petroleum hydrocarbons, hexane, hexadecane and motor oil as model substrates. A. borkumensis showed an incremental growth on these substrates with a high cell count. Growth on motor oil showed highest alkane hydroxylase and lipase production of 2.62 U/ml and 71 U/ml, respectively, while growth on hexadecane showed the highest esterase production of 57.5 U/ml. The percentage of hexane, hexadecane, and motor oil degradation during A. borkumensis growth after 72 h, was around 80%, 81.5% and 75%, respectively. Zymogram showed two different bands with a molecular weight of approx. 52 and 40 kDa, respectively with lipase and esterase activity. Alkane hydroxylase reached optimum activity at pH 8.0 and 70 ± 1 °C for hexane and hexadecane and 75 ± 1 °C for motor oil. Lipase and esterase showed optimum activity at 35 ± 1 °C and 40 ± 1 °C, respectively and pH 7.0. The crude enzymes showed higher stability in a wide range of pH, but they were not thermostable at higher temperatures.

A taxonomic study was carried out on strain MT13131T, which was isolated from deep-sea sediment of the Indian Ocean during the screening of oil-degrading bacteria. The chain length range of n-alkanes (C8 to C32) oxidized by strain MT13131T was determined in this study. The bacterium was Gram-negative, oxidase- and catalase-positive, single rod shaped, and motile by peritrichous flagella. Growth was observed at salinities of 1–12 % and at temperatures of 10–42 °C. The isolate was capable of Tween 20, 40 and 80 hydrolysis, but incapable of gelatin, cellulose or starch hydrolysis. Phylogenetic analysis based on 16S rRNA gene sequences indicated that strain MT13131T belonged to the genus Alcanivorax , with highest sequence similarity to Alcanivorax marinus R8-12T (96.92 %), other species of genus Alcanivorax shared 92.96–96.69 % sequence similarity. The principal fatty acids were summed feature 3 (C16 : 1ω6c/ω7c), summed feature 8 (C18 : 1ω7c/ω6c), C16 : 0 and C12 : 0 3OH. The G+C content of the chromosomal DNA was 64.2 mol%. Phosphatidylglycerol, phosphatidylethanolamine, three aminolipids and three phospholipids were present. The combined genotypic and phenotypic data showed that strain MT13131T represents a novel species within the genus Alcanivorax , for which the name Alcanivorax mobilis sp. nov. is proposed, with the type strain MT13131T (=MCCC 1A11581T=KCTC 52985T).

Halloysites (tubular aluminosilicate) are introduced as inexpensive natural nanoparticles that form and stabilize oil-water emulsions. Pickering emulsification can proceed with energies low enough to be afforded by ocean turbulence and the stability of droplets extends over more than a week. The oil/water interface is shown to be roughened and bacteria, which are added for oil degradation, are better attached to such oil droplets than to droplets without halloysites. The metabolic activity of Alcanivorax borkumensis, alkanotrophic bacteria widely distributed in marine environments, is enhanced by halloysite addition. A halloysite-based dispersant system is therefore environmentally friendly and promising for further optimization. The key elements of the described formulations are natural clay nanotubes, which are abundantly available in thousands of tons, thus making this technology scalable for environmental remediation.

Alcanivorax belongs to the hydrocarbonoclastic group of bacteria that are known for their preferential growth on alkanes and other related compounds. Here we report the genomic features of Alcanivorax marinus strain NMRL4 (=MCC 4632) isolated from oil polluted seawater of the Arabian Sea. Its 4,062,055 bp genome with 66.1% GC content encodes for 3935 coding sequences. The genome annotations of strain NMRL4 revealed the presence of multiple hydrocarbon degradation genes suggestive of its wider hydrocarbon substrate range. The strain encodes for three alkane monooxygenases, two cytochrome P450 and two flavin binding monooxygenases for degradation of short and long-chain alkanes. The genome shows capabilities for scavenging of nutrients, biofilm formation at oil-water interfaces, chemotaxis, motility and habitat specific adaptation. The genomic insights showed that the strain NMRL4 is an ideal candidate for bioremediation of pollutant petroleum hydrocarbons from the marine environment.

Poly-hydroxybutyrate-co-hydroxyhexanoate (PHBH) is a biodegradable, water-insoluble polymer produced by specific bacteria. The monomers of PHBH are the hydroxyalkanoic acids 3-hydroxybutyrate (3HB) and 3-hydroxyhexanoate (3HH). Previously, we reported that 3HB and 3HH showed marked antibacterial activities against the shrimp pathogenic bacterium Vibrio penaeicida, and that addition of 5% (w/w) PHBH to the standard aquaculture diet significantly increased survival rate in kuruma shrimp (Marsupenaeus japonicus) after challenge by V. penaeicida, which we attributed to the degradation of PHBH to its monomers in the shrimp gut. In the present study, we isolated four strains of bacteria with high PHBH-degrading activity and evaluated their inhibitory effects on V. penaeicida with PHBH: one strain from shrimp gut contents (E1; Pseudoalteromonas shioyasakiensis/P. mariniglutinosa), two strains from coastal surface seawater (F1; P. shioyasakiensis/P. mariniglutinosa, and F5; Alcanivorax dieselolei/A. xenomutans), and one strain that was a contaminant in commercial PHBH powder (Y1; Bacillus pseudofirmus). Strains E1, F1, and Y1 showed strong PHBH-degrading activity within 24 h of inoculation to PHBH-containing agar plates. Although none of the isolates alone had any effect on the growth of V. penaeicida, when cultured with E1 or F1 and PHBH, the growth of V. penaeicida was markedly suppressed. Incubation with E1 and PHBH resulted in a gradual reduction in the concentration of V. penaeicida from 2 days after the start of incubation until the concentration was 1.2% of that in the control (V. penaeicida alone). Incubation with F1 and PHBH resulted in a rapid reduction in the concentration of V. penaeicida from 2 days after the start of incubation until the concentration was only 0.32% of that of the control. Compared with strains E1 and F1, Y1 showed similar PHBH-degrading activity but did not show any suppressive effect on the growth of V. penaeicida until 5 days after the start of incubation. In addition, this suppressive effect was relatively weak compared with that of the other two strains, suggesting that Y1 can quickly degrade PHBH but that it takes several days to produce monomers. Together, these results suggest that addition to the aquaculture diet of PHBH and PHBH-degrading bacteria that rapidly degrade PHBH to its monomers may speed up degradation of PHBH to its monomers in the shrimp gut, and that it would increase resistance to infection mortality by V. penaeicida in kuruma shrimp.

Identification of microorganisms that contribute to the whole microbial community is important. In this study, dynamic changes in bioaugmentation process in diesel-polluted seawater collected from two different sites were assessed via simulation experiments. Ultraviolet spectrophotometry and analysis using the molecular operating environment software revealed that the degradation rate of diesel due to bioaugmentation was higher than 70 % after 45 days because of the formation of hydrogen bonds among biosurfactants and diesel components. Community structure and functional genes were analysed via high-throughput sequencing. Results showed that community diversity recovered during bioaugmentation. Principal coordinate analysis showed that the difference in microbial community between the two sites was considerably smaller than that when diesel was added and bioaugmentation was conducted. After bioaugmentation, the main families playing key roles in degradation that became dominant were Alcanivoracaceae, Rhodobiaceae, and Rhodospirillaceae. Moreover, the abundance of functional genes remarkably increased at two different sites.

A taxonomic study was carried out on strain 19-m-6T, which was isolated from deep sea sediment of the South China Sea during the screening of alkane-degrading bacteria. The isolate was Gram-reaction-negative, and oxidase- and catalase- positive. On the basis of 16S rRNA gene sequence similarity, strain 19-m-6T was shown to belong to the genus Alcanivorax , related to Alcanivorax jadensis T9T (97.5 %), Alcanivorax hongdengensis A-11-3T (97.3 %), A. lcanivorax borkumensis SK2T (96.6 %) and seven other species of the genus Alcanivorax (93.9–95.4 %). Average nucleotide identity values between strain 19-m-6T and A. jadensis T9T, A. hongdengensis A-11-3T and A. borkumensis SK2T were 85.12, 85.87 and 84.35 %, respectively. The estimated DNA–DNA hybridization values between strain 19-m-6T and these three type strains were 22.0, 22.6 and 21.2 %, respectively. Four alkane hydroxylase (alkB) genes were obtained from the draft genome sequence. The G+C content of the chromosomal DNA was 56.44 mol%. The major fatty acids were C16 : 0, C18 : 1 ω7c and summed feature 3 (C16 : 1 ω6c and/or C16 : 1 ω7c). The polar lipids were phosphatidylglycerol, phosphatidylethanolamine, one aminolipid, three phospholipids, two glycolipids and two aminophospholipids. According to its phenotypic features, fatty acid composition and 16S rRNA gene sequence, the novel strain fitted well into the genus Alcanivorax , but could be clearly distinguished from all other known Alcanivorax species described to date. The nameAlcanivorax nanhaiticus sp. nov. is thus proposed, with 19-m-6T (=MCCC 1A05629T=KCTC 52137T) as the type strain.

This study aimed to develop a new assay based on the whole cell hybridization in order to monitor alkane hydroxylase genes (alkB system) of the marine bacterium Alcanivorax borkumensis SK2(T) commonly reported as the predominant microorganism responsible for the biodegradation of n-alkanes which are the major fraction of petroleum hydrocarbons. The assay based on the whole cell hybridization targeting alkB2 gene was successfully developed and calibrated on a pure culture of Alcanivorax borkumensis SK2(T) with a detection efficiency up to 80%. The approach was further successfully validated on hydrocarbon-contaminated seawater and provided cells abundance (6.74E+04alkB2-carryingcellsmL(-1)) higher of about one order of magnitude than those obtained by qPCR (4.96E+03alkB2genecopiesmL(-1)). This study highlights the validity of the assay for the detection at single cell level of key-functional genes involved in the biodegradation of n-alkanes.

Bacterial cytochrome P450 enzymes in cytochrome P450 (CYP)153 family were recently reported as fatty acid ω-hydroxylase. Among them, CYP153As from Marinobacter aquaeolei VT8 (CYP153A33), Alcanivorax borkumensis SK2 (CYP153A13), and Gordonia alkanivorans (CYP153A35) were selected, and their specific activities and product yields of ω-hydroxy palmitic acid based on whole cell reactions toward palmitic acid were compared. Using CamAB as redox partner, CYP153A35 and CYP153A13 showed the highest product yields of ω-hydroxy palmitic acid in whole cell and in vitro reactions, respectively. Artificial self-sufficient CYP153A35-BMR was constructed by fusing it to the reductase domain of CYP102A1 (i.e., BM3) from Bacillus megaterium, and its catalytic activity was compared with CYP153A35 and CamAB systems. Unexpectedly, the system with CamAB resulted in a 1.5-fold higher yield of ω-hydroxy palmitic acid than that using A35-BMR in whole cell reactions, whereas the electron coupling efficiency of CYP153A35-BM3 reductase was 4-fold higher than that of CYP153A35 and CamAB system. Furthermore, various CamAB expression systems according to gene arrangements of the three proteins and promoter strength in their gene expression were compared in terms of product yields and productivities. Tricistronic expression of the three proteins in the order of putidaredoxin (CamB), CYP153A35, and putidaredoxin reductase (CamA), i.e., A35-AB2, showed the highest product yield from 5 mM palmitic acid for 9 h in batch reaction owing to the concentration of CamB, which is the rate-limiting factor for the activity of CYP153A35. However, in fed-batch reaction, A35-AB1, which expressed the three proteins individually using three T7 promoters, resulted with the highest product yield of 17.0 mM (4.6 g/L) ω-hydroxy palmitic acid from 20 mM (5.1 g/L) palmitic acid for 30 h.

The modified polyelectrolyte-magnetite nanocoating was applied to functionalize the cell walls of oil decomposing bacteria Alcanivorax borkumensis. Cationic coacervate of poly(allylamine) and 20 nm iron oxide nanoparticles allowed for a rapid single-step encapsulation process exploiting electrostatic interaction with bacteria surfaces. The bacteria were covered with rough 70-100-nm-thick shells of magnetite loosely bound to the surface through polycations. This encapsulation allowed for external manipulations of A. borkumensis with magnetic field, as demonstrated by magnetically facilitated cell displacement on the agar substrate. Magnetic coating was naturally removed after multiple cell proliferations providing next generations of the cell in the native nonmagnetic form. The discharged biosurfactant vesicles indicating the bacterial functionality (150 ± 50 nm lipid micelles) were visualized with atomic force microscopy in the bacterial biofilms.

Bioelectrochemical system is an emerging technology for the remediation of soils contaminated by petroleum hydrocarbons. However, performance of such systems can be limited by the inefficient mass transport in soil. Here we report a new method of sand amendment, which significantly increases both oxygen and proton transports, resulting to increased soil porosity (from 44.5% to 51.3%), decreased Ohmic resistance (by 46%), and increased charge output (from 2.5 to 3.5Cg(-1)soil). The degradation rates of petroleum hydrocarbons increased by up to 268% in 135d. The degradation of n-alkanes and polycyclic aromatic hydrocarbons with high molecular weight was accelerated, and denaturing gradient gel electrophoresis showed that the microbial community close to the air-cathode was substantially stimulated by the induced current, especially the hydrocarbon degrading bacteria Alcanivorax. The bioelectrochemical stimulation imposed a selective pressure on the microbial community of anodes, including that far from the cathode. These results suggested that sand amendment can be an effective approach for soil conditioning that will enhances the bioelectrochemical removal of hydrocarbons in contaminated soils.

A Gram-stain-negative, motile, rod-shaped bacterium, designated SW127T, was isolated from a seawater sample collected from the Indian Ocean. Strain SW127T was aerobic, catalase- and oxidase-positive, and grew at 8–42 °C (optimum, 30 °C), at pH 5.5–9.0 (optimum, pH 7.5) and in the presence of 0.5–11.0 % (w/v) NaCl (optimum, 3.0–4.0 %). Comparative analysis of 16S rRNA gene sequences indicated that strain SW127T belonged to the genus Alcanivorax , and closely related to Alcanivorax pacificus MCCC 1A00474T (96.7 % sequence similarity). The predominant cellular fatty acids of strain SW127T were C16 : 0 and summed feature 8 (comprising C18 : 1ω7c and/or C18 : 1ω6c). Strain SW127T contained phosphatidylethanolamine and phosphatidylglycerol as the major polar lipids. The genomic DNA G+C content of strain SW127T was 62.8 mol%. On the basis of its phenotypic characteristics and phylogenetic data, strain SW127T represents a novel species of the genus Alcanivorax , for which the name Alcanivorax indicus sp. nov. is proposed. The type strain is SW127T (=CGMCC 1.16233T=KCTC 62652T).

Summary Alcanivorax borkumensis SK2T is an important obligate hydrocarbonoclastic bacterium (OHCB) that can dominate microbial communities following marine oil spills. It possesses the ability to degrade branched alkanes which provides it a competitive advantage over many other marine alkane degraders that can only degrade linear alkanes. We used LC–MS/MS shotgun proteomics to identify proteins involved in aerobic alkane degradation during growth on linear (n‐C14) or branched (pristane) alkanes. During growth on n‐C14, A. borkumensis expressed a complete pathway for the terminal oxidation of n‐alkanes to their corresponding acyl‐CoA derivatives including AlkB and AlmA, two CYP153 cytochrome P450s, an alcohol dehydrogenase and an aldehyde dehydrogenase. In contrast, during growth on pristane, an alternative alkane degradation pathway was expressed including a different cytochrome P450, an alcohol oxidase and an alcohol dehydrogenase. A. borkumensis also expressed a different set of enzymes for β‐oxidation of the resultant fatty acids depending on the growth substrate utilized. This study significantly enhances our understanding of the fundamental physiology of A. borkumensis SK2T by identifying the key enzymes expressed and involved in terminal oxidation of both linear and branched alkanes. It has also highlights the differential expression of sets of β‐oxidation proteins to overcome steric hinderance from branched substrates.

Most plastics are released to the environment in landfills and around 32% end up in the sea, inducing large ecological and health impacts. The plastics constitute a physical substrate and potential carbon source for microorganisms. The present study compares the structures of bacterial communities from floating plastics, sediment-associated plastics and sediments from the Mediterranean Sea. The 16S rRNA microbiome profiles of surface and sediment plastic-associated microbial biofilms from the same geographic location differ significantly, with the omnipresence of Bacteroidetes and Gammaproteobacteria. Our research confirmed that plastisphere hosts microbial communities were environmental distinct niche. In parallel, this study used environmental samples to investigate the enrichment of potential plastic-degrading bacteria with Low Density PolyEthylene (LDPE), PolyEthylene Terephthalate (PET) and PolyStyrene (PS) plastics as the sole carbon source. In this context, we showed that the bacterial community composition is clearly plastic nature dependent. Hydrocarbon-degrading bacteria such as Alcanivorax, Marinobacter and Arenibacter genera are enriched with LDPE and PET, implying that these bacteria are potential players in plastic degradation. Finally, our data showed for the first time the ability of Alcanivorax borkumensis to form thick biofilms specifically on LDPE and to degrade this petroleum-based plastic.

In the oligotrophic environment of the Mariana Trench, alkanes as carbohydrates are important for the ecosystem, but their spatial and periodic spreading in deep waters has never been reported. Alkane-degrading bacteria such as Alcanivorax spp. are biological signals of the alkane distribution. In the present study, Alcanivorax was abundant in some waters, at depths of up to 6,000 m, in the Mariana Trench. Genomic, transcriptomic, and chemical analyses provide evidence for the presence and activities of Alcanivorax jadensis in deep sea zones. The periodic spreading of alkanes, probably from the subductive plates, might have fundamentally modified the local microbial communities, as well as perhaps the deep sea microenvironment., In subduction zones, serpentinization and biological processes may release alkanes to the deep waters, which would probably result in the rapid spread of Alcanivorax. However, the timing and area of the alkane distribution and associated enrichment of alkane-degrading microbes in the dark world of the deep ocean have not been explored. In this study, we report the richness (up to 17.8%) of alkane-degrading bacteria, represented by Alcanivorax jadensis, in deep water samples obtained at 3,000 to 6,000 m in the Mariana Trench in two cruises. The relative abundance of A. jadensis correlated with copy numbers of functional almA and alkB genes, which are involved in alkane degradation. In these water samples, we detected a high flux of alkanes, which probably resulted in the prevalence of A. jadensis in the deep waters. Contigs of A. jadensis were binned from the metagenomes for examination of alkane degradation pathways and deep sea-specific pathways, which revealed a lack of nitrate and nitrite dissimilatory reduction in our A. jadensis strains. Comparing the results for the two cruises conducted close to each other, we suggest periodic release of alkanes that may spread widely but periodically in the trench. Distribution of alkane-degrading bacteria in the world’s oceans suggests the periodic and remarkable contributions of Alcanivorax to the deep sea organic carbon and nitrogen sources. IMPORTANCE In the oligotrophic environment of the Mariana Trench, alkanes as carbohydrates are important for the ecosystem, but their spatial and periodic spreading in deep waters has never been reported. Alkane-degrading bacteria such as Alcanivorax spp. are biological signals of the alkane distribution. In the present study, Alcanivorax was abundant in some waters, at depths of up to 6,000 m, in the Mariana Trench. Genomic, transcriptomic, and chemical analyses provide evidence for the presence and activities of Alcanivorax jadensis in deep sea zones. The periodic spreading of alkanes, probably from the subductive plates, might have fundamentally modified the local microbial communities, as well as perhaps the deep sea microenvironment.

The marine hydrocarbonoclastic bacterium Alcanivorax borkumensis is well known for its ability to successfully degrade various mixtures of n-alkanes occurring in marine oil spills. For effective growth on these compounds, the bacteria possess the unique capability not only to incorporate but also to modify fatty intermediates derived from the alkane degradation pathway. High efficiency of both these processes provides better competitiveness for a single bacteria species among hydrocarbon degraders. To examine the efficiency of A. borkumensis to cope with different sources of fatty acid intermediates, we studied the growth rates and membrane fatty acid patterns of this bacterium cultivated on diesel, biodiesel and rapeseed oil as carbon and energy source. Obtained results revealed significant differences in both parameters depending on growth substrate. Highest growth rates were observed with biodiesel, while growth rates on rapeseed oil and diesel were lower than on the standard reference compound (hexadecane). The most remarkable observation is that cells grown on rapeseed oil, biodiesel, and diesel showed significant amounts of the two polyunsaturated fatty acids linoleic acid and linolenic acid in their membrane. By direct incorporation of these external fatty acids, the bacteria save energy allowing them to degrade those pollutants in a more efficient way. Such fast adaptation may increase resilience of A. borkumensis and allow them to strive and maintain populations in more complex hydrocarbon degrading microbial communities.

This study investigates motor oil (3, 5, 7.5 and 10% (v v−1)) as a sole carbon source for the production of Alcanivorax borkumensis in shake flasks and a 5 L bench-scale fermenter in comparison to the standard media. Shake flask studies showed a significant and higher cell growth (p=0.000038), lipase (p = 0.006900) and alkane hydroxylase production (p = 0.000921) by Alcanivorax borkumensis when motor oil was used as the substrate. Based on Tukey post-hoc tests, 5% motor oil concentration was selected as the optimal substrate concentration. The 5 L fermenter experiments conducted using motor oil at 5% (v v−1) concentration, under controlled conditions exhibited significant and higher alkane hydroxylase and lipase activities (55.6 U mL−1 (p = 0.018418) and 208.30 U mL−1 (p = 0.020087), respectively) as compared with those of motor oil at 3% (v v−1) and n-hexadecane at 3% (v v−1) concentration which was used as control. Cell growth was significantly higher when motor oil (3 or 5%) was used as a substrate (p = 0.024705). Enzymatic degradation tested on two different polycyclic aromatic hydrocarbons (PAHs) contaminated groundwaters showed 37.4% removal after 5 days with a degradation rate of 196.6 ppb day−1 and 82.8% removal after 10 days with a degradation rate of 217.54 ppb day-1 for the 1st site and an almost complete biodegradation with 95% removal and 499.02 ppb day−1 removal rate after only 5 days for the 2nd site.

The structures of two bacterial antiporters that act as multidrug resistance efflux pumps, MtrF and YdaH, resemble each other and that of the sodium-coupled succinate symporter VcINDY., Membrane transporter proteins catalyze the passage of a broad range of solutes across cell membranes, allowing the uptake and efflux of crucial compounds. Because of the difficulty of expressing, purifying, and crystallizing integral membrane proteins, relatively few transporter structures have been elucidated to date. Although every membrane transporter has unique characteristics, structural and mechanistic similarities between evolutionarily diverse transporters have been identified. Here, we compare two recently reported structures of membrane proteins that act as antimicrobial efflux pumps, namely MtrF from Neisseria gonorrhoeae and YdaH from Alcanivorax borkumensis, both with each other and with the previously published structure of a sodium-dependent dicarboxylate transporter from Vibrio cholerae, VcINDY. MtrF and YdaH belong to the p-aminobenzoyl-glutamate transporter (AbgT) family and have been reported as having architectures distinct from those of all other families of transporters. However, our comparative analysis reveals a similar structural arrangement in all three proteins, with highly conserved secondary structure elements. Despite their differences in biological function, the overall “design principle” of MtrF and YdaH appears to be almost identical to that of VcINDY, with a dimeric quaternary structure, helical hairpins, and clear boundaries between the transport and scaffold domains. This observation demonstrates once more that the same secondary transporter architecture can be exploited for multiple distinct transport modes, including cotransport and antiport. Based on our comparisons, we detected conserved motifs in the substrate-binding region and predict specific residues likely to be involved in cation or substrate binding. These findings should prove useful for the future characterization of the transport mechanisms of these families of secondary active transporters.

A Gram-reaction-negative, rod-shaped marine bacterium, designated MEBiC08158T, was isolated from sediments collected from Taean County, Korea, near the Hebei Spirit tanker oil spill accident. 16S rRNA gene sequence analysis revealed that strain MEBiC08158T was closely related to Alcanivorax marinus R8-12T (99.5 % similarity) but was distinguishable from other members of the genus Alcanivorax (93.7–97.1 %). The DNA–DNA hybridization value between strain MEBiC08158T and A. marinus R8-12T was 58.4 %. Growth of strain MEBiC08158T was observed at 15–43 °C (optimum 37–40 °C), at pH 6.0–9.5 (optimum pH 7.0–8.0) and with 0.5–16 % NaCl (optimum 1.5–3.0 %). The dominant fatty acids were C16 : 0, C19 : 0 cyclo ω8c, C12 : 0, C18 : 1ω7c, C12 : 0 3-OH and summed feature 3 (comprising C15 : 0 2-OH and/or C16 : 1ω7c). Several phenotypic characteristics differentiate strain MEBiC08158T from phylogenetically close members of the genus Alcanivorax. Therefore, strain MEBiC08158T should be classified as representing a novel species of the genus Alcanivorax, for which the name Alcanivorax gelatiniphagus sp. nov. is proposed. The type strain is MEBiC08158T ( = KCCM 42990T = JCM 18425T).

Oil spill microcosms experiments were carried out to evaluate the effect of bioemulsificant exopolysaccharide (EPS2003) on quick stimulation of hydrocarbonoclastic bacteria. Early hours of oil spill, were stimulated using an experimental seawater microcosm, supplemented with crude oil and EPS2003 (SW+OIL+EPS2003); this system was monitored for 2 days and compared to control microcosm (only oil-polluted seawater, SW+OIL). Determination of bacterial abundance, heterotrophic cultivable and hydrocarbon-degrading bacteria were carried out. Community composition of marine bacterioplankton was determined by 16S rRNA gene clone libraries. Data obtained indicated that bioemulsificant addition stimulated an increase of total bacterial abundance and, in particular, selection of bacteria related to Alcanivorax genus; confirming that EPS2003 could be used for the dispersion of oil slicks and could stimulate the selection of marine hydrocarbon degraders thus increasing bioremediation process.