Effect of inoculation on the biodegradation of wathered Prudhoe Bay crude oil

Summary Enrichment cultures from oil-contaminated beach material from Prince William Sound, Alaska, generated both a mixed bacterial community of indigenous, oil-degrading marine microorganisms and a pure culture oil-degrader, strain EI2V. The mixed and axenic cultures were used in comparative shake flask studies of inoculation on biodegradation of Prudhoe Bay crude oil. Within 12 h following inoculation of homogenized, oiled beach material with the mixed culture, total CO₂ production was increased 2-fold relative to a noninoculated control. Moreover, measurements of phenanthrene degradation (as determined by the release of¹⁴CO₂ from [9-¹⁴C]phenanthrene) showed a 2-or 3-fold greater degradation when inoculated with either strain EI2V or with the mixed culture, respectively. However, as medium was replaced by a simulated tidal cycle, the observed stimulation of CO₂ production decreased, and the addition of strain EI2V had no greater effect on total CO₂ production than the addition of inorganic nutrients alone. Chemical analysis of oil recovered after 7 days incubation also suggested that, while these cultures are capable of efficient biodegradation of Prudhoe Bay crude in liquid culture, inoculation of beach material with high numbers of these microorganisms had little effect on the rate and extent of biodegradation of weathered crude oil. Overall, the sustained stimulatory effect was no greater than that observed with the addition of inorganic nutrients alone.     

Biogenic methane production in formation waters from a large gas field in the North Sea

Methanogenesis was investigated in formation waters from a North Sea oil rimmed gas accumulation containing biodegraded oil, which has not been subject to seawater injection. Activity and growth of hydrogenotrophic methanogens was measured but acetoclastic methanogenesis was not detected. Hydrogenotrophic methanogens showed activity between 40 and 80°C with a temperature optimum (ca. 70°C) consistent with in situ reservoir temperatures. They were also active over a broad salinity range, up to and consistent with the high salinity of the waters (90 g l⁻¹). These findings suggest the methanogens are indigenous to the reservoir. The conversion of H₂ and CO₂ to CH₄ in methanogenic enrichments was enhanced by the addition of inorganic nutrients and was correlated with cell growth. Addition of yeast extract also stimulated methanogenesis. Archaeal 16S rRNA gene sequences recovered from enrichment cultures were closely related to Methanothermobacter spp. which have been identified in other high-temperature petroleum reservoirs. It has recently been suggested that methanogenic oil degradation may be a major factor in the development of the world’s heavy oils and represent a significant and ongoing process in conventional deposits. Although an oil-degrading methanogenic consortium was not enriched from these samples the presence and activity of communities of fermentative bacteria and methanogenic archaea was demonstrated. Stimulation of methanogenesis by addition of nutrients suggests that in situ methanogenic biodegradation of oil could be harnessed to enhance recovery of stranded energy assets from such petroleum systems.     

Evaluation of haloalkaliphilic sulfur-oxidizing microorganisms with potential application in the effluent treatment of the petroleum industry

Haloalkaliphilic sulfur-oxidizing mixed cultures for the treatment of alkaline–saline effluents containing sulfide were characterized and evaluated. The mixed cultures (IMP-PB, IMP-XO and IMP-TL) were obtained from Mexican alkaline soils collected in Puebla (PB), Xochimilco (XO) and Tlahuac (TL), respectively. The Ribosomal Intergenic Spacer Analysis (RISA) revealed bacteria related to Thioalkalibacterium and Thioalkalivibrio in IMP-XO and IMP-PB mixed cultures. Halomonas strains were detected in IMP-XO and IMP-TL. In addition, an uncultured Bacteroides bacterium was present in IMP-TL. Mixed cultures were evaluated at different pH and NaCl concentrations at 30°C. IMP-PB and IMP-TL expressed thiosulfate-oxidizing activity in the 7.5–10.5 pH range, whereas IMP-XO presented its maximal activity with 19.0 mg O₂ g_protein⁻¹ min⁻¹, at pH 10.6; it was not affected by NaCl concentrations up to 1.7 M. In continuous culture, IMP-XO showed a growth rate of 15 day⁻¹, productivity of 433.4 mg_protein l⁻¹ day⁻¹ and haloalkaliphilic sulfur-oxidizing activity was also detected up to 170 mM by means of N-methyl-diethanolamine (MDEA). Saline–alkaline soil samples are potential sources of haloalkaliphilic sulfur-oxidizing bacteria and the mixed cultures could be applied in the treatment of inorganic sulfur compounds in petroleum industry effluents under alkaline–saline conditions.     

A halotolerant Alcanivorax sp. strain with potential application in saline soil remediation

Biodegradation of petroleum compounds in saline environments seems intricate and needs more attention. In this study, tetracosane was used to enrich alkane-degrading bacteria from oil-contaminated saline soils. Among the isolates, strain Qtet3, with the highest 16s rRNA gene sequence similarity to Alcanivorax dieselolei B-5^T, was able to grow at a wide range of NaCl concentrations and was shown by GC analysis to degrade more than 90% of tetracosane in 10 days. This strain has at least two alkB genes and could grow on crude oil and diesel fuel, and utilize various pure aliphatic hydrocarbon substrates (from C₁₂ to C₃₄). Highly hydrophobic cell surfaces and lack of significant surface tension reduction in the media suggest that the main mechanism of the cells for accessing substrate is to attach directly to hydrocarbon particles. Application of this strain for remediating crude oil-contaminated soils irrigated with defined saline water demonstrated that this halotolerant bacterium could survive and grow in saline soils irrigated with NaCl solutions up to 5% w/v, with the highest hydrocarbon degradation of 26.1% observed at 2.5% NaCl. This strain is promising for future industrial applications especially in bioremediation of saline soils and wastes.     

Leucanthemum Vulgare Lam. Germination,Growth and Mycorrhizal Symbiosis Under Crude Oil Contamination

Oil contamination of soil limits plants’ access to water and nutrients. Leucanthemum vulgare colonized by mycorrhizae could provide an effective tool in remedying oil contamination. Seeds of L. vulgare were planted in pots containing soil mixed with petroleum at 0, 2.5, 5, 7.5, and 10% w/w and propagules of mycorrhizal fungi. Plants were grown under ambient conditions for 16 weeks. Seed germination data were collected weekly for three weeks. Mycorrhizal percentage, spore counts, length and weight of roots and shoots were determined after harvesting. Results showed significant differences in seed germination rates between oil-treated, mycorrhizal and non-mycorrhizal plants. The overall germination rate was greater at 7.5% w/w crude oil contamination (ρ = 0.05) in mycorrhizal and non-mycorrhizal pots with significant differences between their respective Root:Shoot ratios (both length and weight). Results of this research showed L. vulgare could be germinated and grown in crude oil contaminated soils and could be used to augment plant establishment as part of phytoremediation practices.     

The root surface phosphatases ofEriophorum vaginatum: Effects of temperature,pH, substrate concentration and inorganic phosphorus

Eriophorum vaginatum L. subsp.spissum (Fern.) Hult., a dominant plant in arctic tundra ecosystems, has acid phosphatase activity evenly distributed along its root surface from the root tip to a distance at least 16 cm from the tip. These root surface phosphatases have optimal activity from pH 3.5 to 4.0; mean soil pH for soil samples collected with roots was 3.69. Apparent energy of activation and Q₁₀ values (14.0 kcal mol⁻¹ and 2.2, respectively) do not provide evidence for temperature acclimation, but substantial phosphatase activity was measured at 1°C. Kinetic parameters determined for this root surface phosphatase were as follows: Km=9.23 mM, Vmax=1.61×10⁻³ μmoles mm⁻²h⁻¹. The presence of inorganic phosphorus in the assay medium did not inhibit root surface phosphatase activity except at very high concentrations (100 mM); even then, only slight inhibition was detected (7 to 19%). A comparison of hydrolysis rates with inorganic phosphate assimilation rates measured forE. vaginatum indicates that organic phosphate hydrolysis may occur at approximately one third the rate of inorganic phosphate absorption. Calculations show that inorganic phosphate produced by root surface phosphatase activity may satisfy 65% of the annual phosphate demand ofE. vaginatum. Since arctic tundra soils are typically higher in dissolved organic phosphorus compounds than in inorganic phosphate, root surface phosphatase activity may make a considerable contribution to the phosphate nutrition of this widespread and abundant arctic plant.     

Biodegradation of petroleum hydrocarbons by filamentous fungi (Aspergillus ustus and Purpureocillium lilacinum) isolated from used engine oil contaminated soil

The use of microorganisms for remediation and restoration of hydrocarbons contaminated soils is an effective and economic solution. The current study aims to find out efficient telluric filamentous fungi to degrade petroleum hydrocarbons pollutants. Six fungal strains were isolated from used engine (UE) oil contaminated soil. Fungi were screened for their ability to degrade crude oil, diesel and UE oil using 2.6-dichlorophenol indophenol (DCPIP). Two isolates were selected, identified and registered at NCBI as Aspergillus ustus HM3.aaa and Purpureocillium lilacinum HM4.aaa. Fungi were tested for their tolerance to different concentration of petroleum oils using radial growth diameter assay. Hydrocarbons removal percentage was evaluated gravimetrically. The degradation kinetic of crude oil was studied at a time interval of 10 days. A.ustus was the most tolerant fungi to high concentration of petroleum oils in solid medium. Quantitative analysis showed that crude oil was the most degraded oil by both isolate; P. lilacinium and A. ustus removed 44.55% and 30.43% of crude oil, respectively. The two fungi were able to degrade, respectively, 27.66 and 21.27% of diesel and 14.39 and 16.00% of UE oil. As compared to the controls, these fungi accumulated high biomass in liquid medium with all petroleum oils. Likewise, crude oil removal rate constant (K) and half-lives (t_1/2) were 0.02 day⁻¹, 34.66 day and 0.015 day⁻¹, 46.21 day for P. lilacinium and A. ustus, respectively. The selected fungi appear interesting for petroleum oils biodegradation and their application for soil bioremediation require scale-up studies.     

Microbial activity and diversity during extreme freeze–thaw cycles in periglacial soils, 5400 m elevation, Cordillera Vilcanota, Perú

High-elevation periglacial soils are among the most extreme soil systems on Earth and may be good analogs for the polar regions of Mars where oligotrophic mineral soils abut with polar ice caps. Here we report on preliminary studies carried out during an expedition to an area where recent glacial retreat has exposed porous mineral soils to extreme, daily freeze–thaw cycles and high UV fluxes. We used in situ methods to show that inorganic nitrogen (NO₃ ⁻ and NH₄ ⁺) was being actively cycled even during a period when diurnal soil temperatures (5 cm depth) ranged from −12 to 27°C and when sub-zero, soil cooling rates reached 1.8°C h⁻¹ (the most rapid soil cooling rates recorded to date). Furthermore, phylogenetic analyses of microbial phylotypes present at our highest sites (5410 m above sea level) showed the presence of nitrifying bacteria of the genus Nitrospira and newly discovered nitrite-oxidizing Betaproteobacteria. These soils were overwhelmingly dominated (>70% of phylotypes) by photosynthetic bacteria that were related to novel cyanobacteria previously found almost exclusively in other plant-free, high-elevation soils. We also demonstrated that soils from our highest sites had higher potential for mineralizing glutamate and higher microbial biomass than lower elevation soils that had been more recently covered by ice. Overall, our findings indicate that a diverse and robustly functioning microbial ecosystem is present in these previously unstudied high-elevation soils.     

Suppression of nitrate formation within an exotic conifer plantation

Summary Nitrate-N losses to stream waters and soil inorganic N pools, nitrifying potentials and NO₃-N production rates were measured in 2 adjacent watersheds, one used as pasture and the other planted in exotic conifer forest (Pinus radiata D. Don). Estimated NO₃-N loss to stream waters draining the pine and pasture watersheds were 0.6kg ha⁻¹ y⁻¹ and 7.6 kg ha⁻¹ y⁻¹ respectively. Ammonium-N pool sizes were not significantly different between soils in the two watersheds but NO₃−N pools and nitrifying potentials were always lower in the pine watershed soil samples. Laboratory incubation experiments indicated that suppression of NO₃−N formation in pine watershed soils required the presence of live tree roots and was not due to the direct action of allelopathic chemicals on nitrifiers.     

Oxygen and nutrient dynamics of the upside down jellyfish (Cassiopea sp.) and its influence on benthic nutrient exchanges and primary production

The oxygen and nutrient dynamics of the zooxanthellate, upside down jellyfish (Cassiopea sp.), were determined both in situ and during laboratory incubations under controlled light conditions. In the laboratory, Cassiopea exhibited a typical Photosynthesis–Irradiance (P–I) curve with photosynthesis increasing linearly with irradiance, until saturation was reached at an irradiance of ~400 μE m⁻² s⁻¹, with photosynthetic compensation (photosynthesis = respiration) being achieved at an irradiance of ~50 μE m⁻² s⁻¹. Under saturating irradiation, gross photosynthesis attained a rate of almost 3.5 mmol O₂ kg WW⁻¹ h⁻¹, whereas the dark respiration rate averaged 0.6 mmol O₂ kg WW⁻¹ h⁻¹. Based upon a period of saturating irradiance of 9 h, the ratio of daily gross photosynthesis to daily respiration was 2.04. Thus, photosynthetic carbon fixation was not only sufficient to meet the carbon demand of respiration, but also to potentially support a growth rate of ~3% per day. During dark incubations Cassiopea was a relatively minor source of inorganic N and P, with the high proportion of NO _X (nitrate + nitrite) produced indicating that the jellyfish were colonised by nitrifying bacteria. Whereas, under saturating irradiance the jellyfish assimilated ammonium, NO _X and phosphate from the bathing water. However, the quantities of inorganic nitrogen assimilated were small by comparison to carbon fixation rates and the jellyfish would need to exploit other sources of nitrogen, such as ingested zooplankton, in order to maintain balanced growth. During in situ incubations the presence of Cassiopea had major effects on benthic oxygen and nutrient dynamics, with jellyfish occupied patches of sediment having 3.6-fold higher oxygen consumption and 4.5-fold higher ammonium regeneration rates than adjacent patches of bare sediment under dark conditions. In contrast at saturating irradiance, jellyfish enhanced benthic photosynthetic oxygen production almost 100-fold compared to the sediment alone and created a small sink for inorganic nutrients, whereas unoccupied sediment patches were sources of inorganic nutrients to the water column. Overall, Cassiopea greatly enhanced the spatial and temporal heterogeneity of benthic fluxes and processes by creating “hotspots” of high activities which switched between being sources or sinks for oxygen and nutrients over diurnal irradiance cycles, as the metabolism of the jellyfish swapped between heterotrophy and net autotrophy.     

n-Alkane uptake and utilisation by Streptomyces strains

Streptomyces strains isolated from the Kuwait Burgan oil field were defined as S. griseoflavus, S. parvus, and S. plicatus utilised n-hexadecane, n-octadecane (purified fractions of mineral oil), kerosene, and crude oil as sole carbon and energy sources. The strains were incubated with n-alkanes and increase of the fatty acid content with chain length equivalent to the employed n-alkanes was observed. Signal transducing GTP-binding proteins (GBPs) play an important role in n-alkane uptake in streptomycetes. Specific activators of GBPs increased the uptake of hydrocarbons. Using the hydrophobic fluorescent dye diphenylhexatrien (DPH) as a probe, it was found that the microviscosity of the hydrophobic inner region of the cellular membrane is significantly lower in hydrocarbon utilisers than in non-utilisers. This difference probably reflects differences in the fatty acid composition of the strains. When cultures were grown in n-alkane containing media, electron microscopy revealed that the hydrocarbon utilisers showed less-electron dense areas as inclusions in the cytoplasm. Soil samples inoculated with Streptomyces strains eliminated hydrocarbons much faster than those not containing these strains, serving as control. When inorganic medium was supplied with n-hexadecane-1-¹⁴C as sole carbon and energy source, radioactive CO₂ was detected. Since streptomycetes have not been used until now for oil elimination, though they are known as abundant soil bacteria tolerating extreme conditions, their possible use for bioremediation of hydrocarbon contaminated soils is discussed.     

The effects of nutrient supply,predominantly addition of iron,and rhizobial inoculation on the tolerance of Lupinus pilosus genotypes to a calcareous soil

Two glasshouse experiments were conducted to examine the effects of nutrient supply and rhizobial inoculation on the performance of Lupinus pilosus genotypes differing in tolerance to calcareous soils. In experiment 1, plants were grown for 84 days in a calcareous soil (50% CaCO₃; soil water content 90% of field capacity) at four nutrient treatments (no-added nutrients, added nutrients without Fe, added nutrients with soil applied FeEDDHA, added nutrients with foliar applied FeSO₄). In experiment 2, plants were grown for 28 days with supply of NH₄NO₃ without inoculation or inoculated with Bradyrhizobium sp. (Lupinus). Chlorosis in the youngest leaves was a good indicator of the relative tolerance of the genotypes to the calcareous soil in both experiments, except the treatment with FeEDDHA at 5 mg kg^–1 soil which was toxic to all genotypes. Chlorosis scores correlated with chlorophyll meter readings and chlorophyll concentrations. The foliar application of FeSO₄ did not fully alleviate chlorotic symptoms despite concentrations of active or total Fe in the youngest leaves being increased. Adding nutrients and chemical nitrogen did not change the severity of chlorosis or improve the growth of the plant. The nutrient supply did not alter the ranking of tolerance of genotypes to the calcareous soil. The results suggest that nutrient deficiency or poor nodulation was not a major cause of poor plant growth on calcareous soils and that bicarbonate may exert a direct effect on chlorophyll synthesis. The mechanism for tolerance is likely to be related to an ability to exclude bicarbonate or prevent its transport to the leaves.     

Degradation of hydrocarbons and biosurfactant production by Pseudomonas sp. strain LP1

Pseudomonas sp. strain LP1, an organism isolated on the basis of its ability to grow on pyrene, was assayed for its degradative and biosurfactant production potentials when growing on crude, diesel and engine oils. The isolate exhibited specific growth rate and doubling time of 0.304 days⁻¹ and 2.28 days, respectively on crude oil (Escravos Light). The corresponding values on diesel were 0.233 days⁻¹ and 2.97 days, while on engine oil, were 0.122 days⁻¹ and 5.71 days. The organism did not show significant biosurfactant production towards crude oil and diesel, but readily produced biosurfactant on engine oil. The highest Emulsification index (E₂₄) value for the biosurfactant produced by LP1 on engine oil was 80.33 ± 1.20, on day 8 of incubation. Biosurfactant production was growth-associated. The surface-active compound which exhibited zero saline tolerance had its optimal activity at 50°C and pH 2.0.     

Toxicity and Bioaccumulation of Petroleum Mixtures with Alkyl PAHs in Earthworms

The toxicities of three oil products with boiling-point ranges representative of petroleum hydrocarbons were tested on earthworms (Eisenia fetida) to investigate the correlation between bioaccumulated concentrations of polycyclic aromatic hydrocarbons (PAHs) and toxicity. The toxicities to earthworms were in the sequence: kerosene > diesel > bunker-C. After 14 days, the LC50s of the soils contaminated with kerosene, diesel, and bunker-C were 1079, 9135, and 15,609 mg/kg, respectively. Analysis of the body residue concentrations of PAHs in the earthworms showed that the accumulation of alkyl PAHs predominated that of the 16 priority PAHs. Principal component analysis (PCA) identified 12 PAHs, including four alkylated naphthalenes, as the oil constituents that affected mortality in the kerosene-contaminated soil. For the diesel-contaminated soil, eight PAHs were identified, including dibenzothiophene. It was not clear which compounds affected mortality in the bunker-C soil. Across the three series, biota-to-soil accumulation factors (BSAFs) ranged from 10^–2.05 to 10^3.98, and generally increased as the hydrophobicity (K_ow) or molecular weight of the alkyl PAHs increased. The toxicity endpoints of each oil product can be used as reference values in the risk assessment of soils contaminated with petroleum, and individual PAHs screened out have implications for future toxicity assessment of petroleum hydrocarbons.     

The effect of water exchange on bacterioplankton depletion and inorganic nutrient dynamics in coral reef cavities

We studied the effect of water exchange on the depletion (or accumulation) of bacterioplankton, dissolved organic matter and inorganic nutrients in small open framework cavities (50–70 l) at 15 m depth on the coral reef along Curaçao, Netherlands Antilles. The bacterioplankton removal rate in cavities increased with increasing water exchange rates up to a threshold of 0.0045 s⁻¹, reaching values of 50–100 mg C m⁻² total interior cavity surface area (CSA) per day. Beyond the threshold, bacterioplankton removal dropped. The cryptic community is apparently adapted to the average water exchange in these cavities (0.0041 s⁻¹). Dissolved inorganic nitrogen (DIN), nitrate + nitrite (NO _x ) in particular, accumulated in cavity water and the accumulation decreased with increasing water exchange. Net NO _x effluxes exceeded net DIN effluxes from cavities (average efflux rate of 1.9 mmol NO _x vs. 0.8 mmol DIN m⁻² interior CSA per day). The difference is ascribed to net ammonium losses (NH₄) in cavities at reef concentrations >0.025 μM NH₄, possibly due to enhanced nitrification. Dissolved inorganic phosphate accumulated in cavities, but was not related to water exchange. The cryptic biota in cavities depend on water exchange for optimization of consumption of bacterioplankton and removal of inorganic nitrogen. Coral cavities are an evident sink of bacterioplankton and a source of NO _x and PO ₄ ³⁻ .     

Accumulation of Hydrocarbons by Maize (Zea mays L.) in Remediation of Soils Contaminated with Crude Oil

This study has investigated the use of screened maize for remediation of soil contaminated with crude oil. Pots experiment was carried out for 60 days by transplanting maize seedlings into spiked soils. The results showed that certain amount of crude oil in soil (≤2 147 mg·kg^?1) could enhance the production of shoot biomass of maize. Higher concentration (6 373 mg·kg^?1) did not significantly inhibit the growth of plant maize (including shoot and root). Analysis of plant shoot by GC-MS showed that low molecular weight polycyclic aromatic hydrocarbons (PAHs) were detected in maize tissues, but PAHs concentration in the plant did not increase with higher concentration of crude oil in soil. The reduction of total petroleum hydrocarbon in planted soil was up to 52.21–72.84%, while that of the corresponding controls was only 25.85–34.22% in two months. In addition, data from physiological and biochemical indexes demonstrated a favorable adaptability of maize to crude oil pollution stress. This study suggested that the use of maize (Zea mays L.) was a good choice for remediation of soil contaminated with petroleum within a certain range of concentrations.     

Porewater Stoichiometry of Terminal Metabolic Products, Sulfate, and Dissolved Organic Carbon and Nitrogen in Estuarine Intertidal Creek-bank Sediments

Porewater equilibration samplers were used to obtain porewater inventories of inorganic nutrients (NH₄⁺, NO_x, PO₄³⁻), dissolved organic carbon (DOC) and nitrogen (DON), sulfate (SO₄²⁻), dissolved inorganic carbon (DIC), hydrogen sulfide (H₂S), chloride (Cl⁻), methane (CH₄) and reduced iron (Fe²⁺) in intertidal creek-bank sediments at eight sites in three estuarine systems over a range of salinities and seasons. Sulfate reduction (SR) rates and sediment particulate organic carbon (POC) and nitrogen (PON) were also determined at several of the sites. Four sites in the Okatee River estuary in South Carolina, two sites on Sapelo Island, Georgia and one site in White Oak Creek, Georgia appeared to be relatively pristine. The eighth site in Umbrella Creek, Georgia was directly adjacent to a small residential development employing septic systems to handle household waste. The large data set (>700 porewater profiles) offers an opportunity to assess system-scale patterns of porewater biogeochemical dynamics with an emphasis on DOC and DON distributions. SO₄²⁻ depletion (SO₄²⁻)_Dep was used as a proxy for SR, and (SO₄²⁻)_Dep patterns agreed with measured (³⁵S) patterns of SR. There were significant system-scale correlations between the inorganic products of terminal metabolism (DIC, NH₄⁺ and PO₄³⁻) and (SO₄²⁻)_Dep, and SR appeared to be the dominant terminal carbon oxidation pathway in these sediments. Porewater inventories of DIC and (SO₄²⁻)_Dep indicate a 2:1 stoichiometry across sites, and the C:N ratio of the organic matter undergoing mineralization was between 7.5 and 10. The data suggest that septic-derived dissolved organic matter with a C:N ratio below 6 fueled microbial metabolism and SR at a site with development in the upland. Seasonality was observed in the porewater inventories, but temperature alone did not adequately describe the patterns of (SO₄²⁻)_Dep, terminal metabolic products (DIC, NH₄⁺, PO₄³⁻), DOC and DON, and SR observed in this study. It appears that production and consumption of labile DOC are tightly coupled in these sediments, and that bulk DOC is likely a recalcitrant pool. Preferential hydrolysis of PON relative to POC when overall organic matter mineralization rates were high appears to drive the observed patterns in POC:PON, DOC:DON and DIC:DIN ratios. These data, along with the weak seasonal patterns of SR and organic and inorganic porewater inventories, suggest that the rate of hydrolysis limits organic matter mineralization in these intertidal creek-bank sediments.     

Bioremediation of diesel-oil-contaminated alpine soils at low temperatures

Bioremediation of two diesel-oil-contaminated alpine subsoils, differing in soil type and bedrock, was investigated in laboratory experiments at 10 °C after supplementation with an inorganic fertilizer. Initial diesel oil contamination of 4000 mg kg⁻¹ soil dry matter (dm) was reduced to 380–400 mg kg⁻¹ dm after 155 days of incubation. In both soils, about 30 % of the diesel oil contamination (1200 mg kg⁻¹ dm) was eliminated by abiotic processes. The residual decontamination (60 %–65 %) could be attributed to microbial degradation activities. In both soils, the addition of a cold-adapted diesel-oil-degrading inoculum enhanced biodegradation rates only slightly and temporarily. From C/N and N/P ratios (determined by measuring the contents of total hydrocarbons, NH₄ ⁺ N, NO₃ ⁻ N and PO₄ ³⁻ P) of soils␣it could be deduced that there was no nutrient deficiency during the whole incubation period. Soil biological activities (basal respiration and dehydrogenase activity) corresponded to the course of biodegradation activities in the soils. Received: 9 September 1996 / Accepted: 7 December 1996     

Microbial Activity and Community Composition during Bioremediation of Diesel-Oil-Contaminated Soil: Effects of Hydrocarbon Concentration,Fertilizers, and Incubation Time

We investigated the influence of three factors—diesel oil concentration [2500, 5000, 10,000, 20,000 mg total petroleum hydrocarbons (TPH) kg⁻¹ soil], biostimulation (unfertilized, inorganic fertilization with NPK nutrients, or oleophilic fertilization with Inipol EAP22), and incubation time—on hydrocarbon removal, enzyme activity (lipase), and microbial community structure [phospholipid fatty acids (PLFA)] in a laboratory soil bioremediation treatment. Fertilization enhanced TPH removal and lipase activity significantly (P ≤ 0.001). The higher the initial contamination, the more marked was the effect of fertilization. Differences between the two fertilizers were not significant (P > 0.05). Microbial communities, as assessed by PLFA patterns, were primarily influenced by the TPH content, followed by fertilization, and the interaction of these two factors, whereas incubation time was of minor importance. This was demonstrated by three-factorial analysis of variance and multidimensional scaling analysis. Low TPH content had no significant effect on soil microbial community, independent of the treatment. High TPH content generally resulted in increased PLFA concentrations, whereby a significant increase in microbial biomass with time was only observed with inorganic fertilization, whereas oleophilic fertilization (Inipol EAP22) tended to inhibit microbial activity and to reduce PLFA contents with time. Among bacteria, PLFA indicative of the Gram-negative population were significantly (P ≤ 0.05) increased in soil samples containing high amounts of diesel oil and fertilized with NPK after 21–38 days of incubation at 20°C. The Gram-positive population was not significantly influenced by TPH content or biostimulation treatment.     

DOC and DIC in Flowpaths of Amazonian Headwater Catchments with Hydrologically Contrasting Soils

Organic and inorganic carbon (C) fluxes transported by water were evaluated for dominant hydrologic flowpaths on two adjacent headwater catchments in the Brazilian Amazon with distinct soils and hydrologic responses from September 2003 through April 2005. The Ultisol-dominated catchment produced 30% greater volume of storm-related quickflow (overland flow and shallow subsurface flow) compared to the Oxisol-dominated catchment. Quickflow fluxes were equivalent to 3.2 ± 0.2% of event precipitation for the Ultisol catchment, compared to 2.5 ± 0.3% for the Oxisol-dominated watershed (mean response ±1 SE, n = 27 storms for each watershed). Hydrologic responses were also faster on the Ultisol watershed, with time to peak flow occurring 10 min earlier on average as compared to the runoff response on the Oxisol watershed. These different hydrologic responses are attributed primarily to large differences in saturated hydraulic conductivity (K _s). Overland flow was found to be an important feature on both watersheds. This was evidenced by the response rates of overland flow detectors (OFDs) during the rainy season, with overland flow intercepted by 54 ± 0.5% and 65 ± 0.5% of OFDs for the Oxisol and Ultisol watersheds respectively during biweekly periods. Small volumes of quickflow correspond to large fluxes of dissolved organic C (DOC); DOC concentrations of the hydrologic flowpaths that comprise quickflow are an order of magnitude higher than groundwater flowpaths fueling base flow (19.6 ± 1.7 mg l⁻¹ DOC for overland flow and 8.8 ± 0.7 mg l⁻¹ DOC for shallow subsurface flow versus 0.50 ± 0.04,mg l⁻¹ DOC in emergent groundwater). Concentrations of dissolved inorganic C (DIC, as dissolved CO₂–C plus HCO₃⁻–C) in groundwater were found to be an order of magnitude greater than quickflow DIC concentrations (21.5 mg l⁻¹ DIC in emergent groundwater versus 1.1 mg l⁻¹ DIC in overland flow). The importance of deeper flowpaths in the transport of inorganic C to streams is indicated by the 40:1 ratio of DIC:DOC for emergent groundwater. Dissolved CO₂–C represented 92% of DIC in emergent groundwater. Results from this study illustrate a highly dynamic and tightly coupled linkage between the C cycle and the hydrologic cycle for both Ultisol and Oxisol landscapes: organic C fluxes strongly tied to flowpaths associated with quickflow, and inorganic C (particularly dissolved CO₂) transported via deeper flowpaths.