A pyrene-degrading bacterial consortium was obtained from deep-sea sediments of the Pacific Ocean. The consortium degraded many kinds of polycyclic aromatic hydrocarbons (PAHs), including naphthalene, phenanthrene, pyrene, acenaphthene, fluorene, anthracene, fluoranthene, 2-methylnaphthalene and 2,6-dimethylnaphthalene, but it did not grow with chrysene and benzo[alpha]pyrene. With methods of plate cultivation and polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE), 72 bacteria belonging to 22 genera were detected from this consortium. Among the detected bacteria, the following genera frequently occurred: Flavobacterium, Cycloclasticus, Novosphingobium, Halomonas, Achromobacter, Roseovarius and Alcanivorax. The first two genera showed the strongest bands in denaturing gradient gel electrophoresis (DGGE) profiles and appeared in all PAH treatments. By now, only one isolate designated P1 was confirmed to be a pyrene degrader. It was identified to be Cycloclasticus spirillensus (100%). Although P1 can degrade pyrene independently, other bacteria, such as Novosphingobium sp. (Band 14), Halomonas sp. (Band 16) and an unidentified bacterium (Band 35), were involved in pyrene degradation in some way; they persist in the consortium in the test of dilution to extinction if only the consortium was motivated with pyrene. However, the secondary most important member Flavobacterium sp. evaded from the community at high dilutions. As a key member of the consortium, P1 distinguished itself by both cell morphology and carbon source range among the isolates of this genus. Based on intermediate analyses of pyrene degradation, P1 was supposed to take an upper pathway different from that previously reported. Together with the results of obtained genes from P1 homology with those responsible for naphthalene degradation, its degradation to pyrene is supposed to adopt another set of genes unique to presently detected. Summarily, an efficient pyrene-degrading consortium was obtained from the Pacific Ocean sediment, in which Cycloclasticus bacterium played a key role. This is the first report to exploit the diversity of pyrene-degrading bacteria in oceanic environments.

The bacteria involved in the biodegradation of polycyclic aromatic hydrocarbons (PAHs) in deep sea subsurface environments are largely unknown. In order to reveal their biodiversity, sediments from 2.2 m under the bottom surface at a water depth of 3542 m were sampled on the Middle Atlantic Ridge with a gravity column sampler. The sediments were promptly enriched with either crude oil or a mixture of PAHs (naphthalene, phenanthrene and pyrene) as the sole carbon source, and further enriched with the PAH mixture mentioned above in the lab. The resulting consortia were named C2CO and C2PPN respectively. Their bacterial composition was analysed with plate cultivation, PCR-DGGE and 16S rDNA library analysis. On plates, isolates belonging to Pseudoalteromonas, Halomonas, Marinobacter, Thalassospira and Tistrella dominated the culturable populations. With PCR-DGGE, five major bands closely related to Cycloclasticus, Alteromonas, Thalassospira, Alcanivorax and Rhodospirillaceae were detected in consortium C2CO, while only one major band of Cycloclasticus was detected in consortium C2PPN. In addition, the dynamics of community structure in response to aromatic substrate alterations were examined. As a result, three ribotypes of Cycloclasticus were detected by 16S rDNA library analysis, one which played a key role in phenanthrene degradation; two Alteromonas bacteria dominated the naphthalene reselected consortium. Although bacteria of the two genera grew as the main members of the communities, none of them were isolated, probably owing to their poor cultivability. These results confirm that bacteria of Cycloclasticus are important obligate PAH degraders in marine environments, and coexist with other degrading bacteria that inhabit the deep subsurface sediment of the Atlantic. This supports the view that PAH accumulation and bioattenuation occur in remote areas consistently and continuously.

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A Gram-negative, rod-shaped, slightly halophilic and facultatively anaerobic bacterium, designated strain D15-8W(T), was isolated from the sediment of the South China Sea. Growth was found to occur optimally at 25 °C, between pH 7.0 and 8.0 and with 1-5 % (w/v) NaCl. The strain was observed to utilize a variety of organic substrates and polycyclic aromatic hydrocarbons as sole carbon sources. The G+C content of the genomic DNA was determined to be 58.7 %. The predominant respiratory quinone was found to be Q-9. The significant fatty acids were determined to be C(16:0), C(16:1) ω9c, C(18:1) ω9c, C(12:0) and C(14:0) 3OH. Analysis of 16S rRNA gene sequences showed that strain D15-8W(T) fits within the phylogenetic cluster of the genus Marinobacter and is most closely related to Marinobacter segnicrescens CGMCC 1.6489(T), Marinobacter bryozoorum DSM 15401(T), Marinobacter lacisalsi CECT 7297(T) and Marinobacter daqiaonensis CGMCC1.9167(T). The DNA-DNA hybridization values between strain D15-8W(T) and the type strains of the most closely related species were 42.3 % (CGMCC 1.6489(T)), 39.8 % (DSM 15401(T)), 37.3 % (CECT 7297(T)) and 35.2 % (CGMCC1.9167(T)). The results of this polyphasic study indicate that strain D15-8W(T) represents a novel species of the genus Marinobacter, for which the name Marinobacter nanhaiticus sp. nov. is proposed. The type strain is D15-8W(T) (=CGMCC 1.11019(T)=KCTC 23749(T)).

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Whole-cell bioreporters have emerged as promising tools for genotoxicity evaluation, due to their rapidity, cost-effectiveness, sensitivity and selectivity. In this study, a method for detecting genotoxicity in environmental samples was developed using the bioluminescent whole-cell bioreporter Escherichia coli recA::luxCDABE. To further test its performance in a real world scenario, the E. coli bioreporter was applied in two cases: i) soil samples collected from chromium(VI) contaminated sites; ii) crude oil contaminated seawater collected after the Jiaozhou Bay oil spill which occurred in 2013. The chromium(VI) contaminated soils were pretreated by water extraction, and directly exposed to the bioreporter in two phases: aqueous soil extraction (water phase) and soil supernatant (solid phase). The results indicated that both extractable and soil particle fixed chromium(VI) were bioavailable to the bioreporter, and the solid-phase contact bioreporter assay provided a more precise evaluation of soil genotoxicity. For crude oil contaminated seawater, the response of the bioreporter clearly illustrated the spatial and time change in genotoxicity surrounding the spill site, suggesting that the crude oil degradation process decreased the genotoxic risk to ecosystem. In addition, the performance of the bioreporter was simulated by a modified cross-regulation gene expression model, which quantitatively described the DNA damage response of the E. coli bioreporter. Accordingly, the bioluminescent response of the bioreporter was calculated as the mitomycin C equivalent, enabling quantitative comparison of genotoxicities between different environmental samples. This bioreporter assay provides a rapid and sensitive screening tool for direct genotoxicity assessment of environmental samples.

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A case study to investigate the relationship between antibacterial activity and quorum sensing mechanisms was carried out on a sponge-associated bacterium with remarkable biological activities: Pseudoalteromonas sp. NJ6-3-1. The dependence of active substance production on cell density was studied under various growth conditions. Bacteria NJ6-3-1 was found to start producing antibacterial compounds only when cell density reached the threshold value of OD 630 =0.4. To simulate the competitive real marine environment, NJ6-3-1 at low cell density (OD 630 value below the required threshold value) was cocultured with the terrestrial bacterium Staphylococcus aureus. Antibacterial activity assays indicated the existence of some signal molecules in the metabolites of S. aureus that could induce NJ6-3-1 to produce antibacterial substances even at low cell density. Three diketopiperazines (DKPs) as metabolites and potential autoinducers of NJ6-3-1 were synthesized and co-cultured with low density NJ6-3-1. The antibacterial activity assay showed that one of these DKPs-cyclo-(L-Phe-L-Val)-was the autoinducer and could indeed induce NJ6-3-1 to produce antibacterial substances under low cell density. Our results thus provide preliminary support to the hypothesis that the antibacterial activity of NJ6-3-1 is controlled by the quorum sensing system in both an intra-species and an inter-species manner.