Inorganic nutrient utilisation by "adapted" Pseudomonas putida strain used in the bioremediation of agricultural soil polluted with crude petroleum

Garden soil samples polluted with crude petroleum were bioremediated by inorganic nutrient monitoring with appropriate adjustment and inoculation with crude oil-adapted strain of Pseudomonasputida (PP) isolated from oil-impacted soils. Soil samples without PP inoculation served as the control samples to compare the abilities of the native soil microflora with the adapted PP strain in biodegrading crude oil pollutant. In the experimental samples, oil concentration and all the inorganic nutrient sources tested decreased more rapidly with a proportional increase in the population densities of both PP and the native soil microflora than were observed in the control samples. This trend was particularly strong for PO4(3-) and NO3- which eventually became limiting both in all the experimental samples and in some control samples. Inoculation of crude oil-impacted agricultural soils by oil -adapted PP strain with nutrient monitoring and adjustment can be effective as bioremediation methods of agricultural land upon pollution with petroleum or petroleum products.     

Investigation of bioremediation potential of zymogenous bacteria and fungi for crude oil degradation

Bioremediation potential of bacteria and fungi isolated from sludge samples has been investigated (Danube alluvium, Pančevo, Serbia). Total isolated microorganisms were divided into three parts. One part was added with actidione antifungicide. The second part was added with streptomycin antibiotic. The third part was without additives. Paraffinic type of crude oil was a substrate for assessment of bioremediation potential. The simulated oil biodegradation lasted 30, 60 and 90 days. Parallel with that, the experiments with blind trial were conducted. Extracts were isolated from the samples with chloroform in a separate funnel. They were assayed for the group composition (alkanes, aromatics, alcohols and fatty acids) by column chromatography. Alkane fraction was analysed by gas chromatography—mass spectrometry (GC–MS). The most intense oil degradation was achieved in the experiments with bacteria, somewhat weaker with consortium of fungi and bacteria, and the weakest bioremediation potential in these experiments was shown by fungi.     

Microbial community and functional genes in the rhizosphere of alfalfa in crude oil-contaminated soil

A rhizobox system constructed with crude oil-contaminated soil was vegetated with alfalfa (Medicago sativa L.) to evaluate the rhizosphere effects on the soil microbial population and functional structure, and to explore the potential mechanisms by which plants enhance the removal of crude oil in soil. During the 80-day experiment, 31.6% of oil was removed from the adjacent rhizosphere (AR); this value was 27% and 53%higher than the percentage of oil removed from the far rhizosphere (FR) and from the non-rhizosphere (NR), respectively. The populations of heterotrophic bacteria and hydrocarbon-degrading bacteria were higher in the AR and FR than in the NR. However, the removal rate of crude oil was positively correlated with the proportion of hydrocarbon-degrading bacteria in the rhizosphere. In total, 796, 731, and 379 functional genes were detected by microarray in the AR, FR, and NR, respectively. Higher proportions of functional genes related to carbon degradation and organic remediation, were found in rhizosphere soil compared with NR soil, suggesting that the rhizosphere selectively increased the abundance of these specific functional genes. The increase in water-holding capacity and decrease in pH as well as salinity of the soil all followed the order of AR>FR>NR. Canonical component analysis showed that salinity was the most important environmental factor influencing the microbial functional structure in the rhizosphere and that salinity was negatively correlated with the abundance of carbon and organic degradation genes.     

Degradation of methyl-phenanthrene isomers during bioremediation of soil contaminated by residual fuel oil

Phenanthrene and methyl-phenanthrenes are major aromatic pollutants originating in particular from fuel oil. Phenanthrene is usually degraded faster than methyl-phenanthrenes under geological and environmental conditions. Here, we report a preferential and accelerated biodegradation of methyl-phenanthrenes versus phenanthrene in soil contaminated by fuel oil. The polluted soil was mixed with sawdust and sand to form a homogenized biopile. The biopile was continuously sprayed with microbial consortia isolated from crude oil?Ccontaminated soil and treated by biosurfactants and nutritive substances for biostimulation. During a 6-month bioremediation experiment, a steady increase in the relative abundance of phenanthrene compared to methyl-phenathrenes was observed by gas chromatography?Cmass spectrometry. The increase was the highest for trimethyl-phenanthrenes, with a phenanthrene/trimethyl-phenanthrenes ratio increasing from 0.42 to 2.45. By contrast, the control, non-stimulated samples showed a ratio decrease from 0.85 to 0.11. Moreover, the results showed that the level of degradability depends on the number of methyl groups.     

Effect of Diesel Fuel on Growth of Selected Plant Species

Diesel oil is a complex mixture of hydrocarbons with an average carbon number of C8–C26. The majority of components consist of alkanes, both straight chained and branched and aromatic compounds including mono-, di- and polyaromatic hydrocarbons. Regardless of this complexity, diesel oil can be readily degraded by a number of soil microorganisms making it a likely candidate for bioremediation. The concept of using plants to enhance bioremediation, termed phytoremediation, is a relatively new area of scientific interest. It is particularly applicable to diesel oil contamination as diesel oil generally contaminates the top few metres of soil (surface soil) and contamination is not uniform throughout the site. By encouraging plants to grow on diesel oil contaminated soil, conditions are improved for the microbial degradation of the contaminant. During this study, establishing plants on diesel oil contaminated soil proved difficult. Diesel oil is phytotoxic to plants at relatively low concentrations. At concentrations below this phytotoxic level, the development of plants grown in diesel oil contaminated soil differs greatly from plants grown in uncontaminated soil. Tolerance of plants to diesel oil and ability to germinate in diesel oil contaminated soil varied greatly between plant species as well as within plant species. The broadest differences in germination were seen within the grasses with certain species thriving in low levels of contamination (e.g. Creeping bent) while others were intolerant of diesel oil contamination (e.g. Rough meadow grass). The herbs, legumes and commercial crops screened appeared to be largely unaffected by low levels of diesel oil contamination (25g dieselkg^–1). At the higher level of contamination (50g dieselkg^–1), half of the twenty two plants species screened failed to reach a germination rate equal to 50% of the control rate. Two species of grass failed to germinate at all at this contamination level. Plant species that successfully germinated and grew were studied further to determine the effect of diesel oil contamination on the later stages of plant development. This work investigates the effect of diesel oil on plant growth and development.     

Long-term effects of water-soluble fractions of Kuwait crude oil on the larval and juvenile development of the mud crab Eurypanopeus depressus

Larvae of the mud crab Eurypanopeus depressus (Smith) were reared in various concentrations of the water-soluble fraction of Kuwait crude oil. The 48-h TL_m (median tolerance limit) for Zoea Stage I was approximately 10 ppm total dissolved hydrocarbons and that for Zoea Stage II approximately 17 ppm. Chronic toxicity of more dilute solutions (4.3 and 8.7 ppm) was assessed independently for each larval stage and for subsequent developmental stages through Crab Stage 5. In the group continuously exposed to oil from hatching, there was differential mortality relative to controls in every larval stage and increased duration of intermolt periods was observed at every stage through Crab Stage 5. Mortality in groups not exposed until larvae had reached Zoea Stages III or IV was not greater than controls, suggesting that toxicity to advanced larval stages may be related to accumulation of toxic compounds by the larvae. Neither concentration (4.3 or 8.7 ppm) of crude oil caused increased mortality among juvenile crab stages regardless of the time of initial exposure. Increased occurrence of an extra and morphologically abnormal megalopa stage was associated with exposure to the crude oil. This has not been reported before.Communicated by I. Morris, West Boothbay Harbor     

渤海相近石油平台原油指纹鉴别规律初探

选择我国渤海相邻两个石油平台,不同油井6种原油,依据重复性限及t检验方法进行油源比对,对渤海相近平台原油指纹鉴别规律进行了初探性研究.结果表明,同一原油开采平台或相近的石油平台,原油指纹信息总体相差不大,仅部分组分表现出显著差异,如n-C17/姥鲛烷、n-C18/植烷在渤海相近石油平台原油中差异最为显著,此外,伽玛蜡烷、升藿烷、莫烷、C29甾烷、C19—C22的正构烷烃、菲与烷基化菲类、二苯并噻吩及烷基化二苯并噻吩类也表现出明显差异.由此加强对原油中该类组分筛查,将有助于石油平台原油油源鉴别.     

黄河三角洲生态区土壤石油污染及其对碱蓬萌发的生态影响

基于黄河三角洲生态区内的油田生产区中石油污染土壤的调查取样,分析石油污染状况,并在实验室人工控制条件下,研究土壤中不同质量分数的石油污染对该区先锋植物——碱蓬（Salsola glauca Bung）的种子萌发影响。结果表明,油田区土壤受到了不同程度的石油污染;溢油处石油的质量分数最高达6.230 g·kg^-1。另外,萌发实验说明在土壤污染情况下种子萌发率具有不确定性,土壤石油污染质量分数为5～40 g·kg^-1时,对碱蓬种子的萌发无明显抑制作用。该研究对黄河三角洲生态区土壤石油类污染敏感指示指标的筛选和植物的生物修复具有重要意义。     

喀斯特生境下AMF侵染对任豆生长的影响

喀斯特地区土壤退化,植被定植更新困难,丛枝菌根真菌(Arbuscular mycorrhizal fungi,AMF)具有增强植物养分吸收能力和抵抗逆境胁迫能力。研究喀斯特生境下植物与AMF共生效果,选择优势菌种促进喀斯特植被恢复,对于提高植物定植成活率具有重要作用。以豆科植物任豆(Zenia insignis)幼苗为试验材料,盆栽条件下,选取喀斯特优势菌种-摩西球囊霉(Funneliformis mosseae)、根内球囊霉(Rhizophagus intraradices),2种菌根真菌混合菌剂进行接种,培养180 d,研究贫瘠喀斯特土壤生境和养分较高的滇柏林下土壤生境下AMF对任豆生长影响。结果表明:摩西球囊霉、根内球囊霉和混合接种均能侵染任豆根系,幼嫩根系更易侵染,木质化根系侵染率下降。接种摩西球囊霉,贫瘠喀斯特土壤生境下,株高、地径、地上生物量、地下生物量和总生物量分别提高68.92%、56.18%、83.90%、42.20%和67.34%;养分较高的滇柏林下喀斯特土壤生境下,株高、地上生物量、地下生物量和总生物量分别提高48.05%、6.77%、7.92%和8.89%;根内球囊霉处理接种效应低于摩西球囊霉和混合接种处理,对生物量增长为负效应,混合接种处理接种效应介于单接种之间,摩西球囊霉接种效果优于根内球囊霉和混合接种。摩西球囊霉在贫瘠喀斯特土壤生境下发挥的促生效应优于养分较高的喀斯特土壤,可作为喀斯特侵蚀区植被恢复菌根真菌干扰途径的优势菌种,混合接种作为接种剂具有单接种兼容效应。     

添加不同营养助剂对石油污染土壤生物修复的影响

实验室条件下研究NPK复合肥、诺沃肥和腐殖酸三种物质不同配比添加对石油污染土壤的生物修复效果.在60d的修复实验中,定期取样测定土壤含油量、总异养菌数、石油烃降解菌数和脱氢酶活性,并采用PCR-DGGE技术研究修复过程中微生物多样性变化.结果表明,NPK肥的添加和NPK肥-诺沃肥-腐殖酸复合添加能够提高土壤中微生物的数量、微生物多样性和脱氢酶活性.在含油量为84600mg.kg-1的土壤中,添加营养助剂的处理60d后石油烃降解率为31.3%—39.5%,不添加营养助剂的石油烃降解率仅为3.5%.NPK肥-诺沃肥-腐殖酸复合添加对石油烃的降解率要高于NPK肥的单独添加(高8%),其原因可能是诺沃肥-腐殖酸能够有效提高土著微生物的活性,增加微生物的多样性,增强石油烃的降解.     

假单胞菌AEBL3对土壤中呋喃丹的生物降解

从农药污染的农田土壤中分离到一株假单胞菌AEBL3，该菌能够以呋喃丹为唯一的碳源和氮源生长。使用AEBL3作为生物强化剂对模拟呋喃丹污染的土壤环境进行了生物修复试验。结果表明，接种AEBL3能够明显增加土壤中呋喃丹的降解率。降解菌AEBL3在土壤中具有一定的移动性，当从土壤表层加菌时，对0-7cm深土层中的呋喃丹都有很好的降解效果。在各种投加方式的比较中，以投加降解菌原液修复效果最好，缓冲液悬浮的菌细胞次之，砂粒混合物的效果最差。16S rRNA的变性梯度电泳(DGGE)结果揭示了生物修复过程中土壤微生物菌群结构中的动态变化。     

Phytotoxicity of volatile oil from Eucalyptus citriodora against some weedy species

A study was undertaken to explore the phytotoxicity of volatile essential oil from Eucalyptus citriodora Hook. against some weeds viz. Bidens pilosa, Amaranthus viridis, Rumex nepalensis, and Leucaena leucocephala in order to assess its herbicidal activity. Dose-response studies conducted under laboratory conditions revealed that eucalypt oils (in concentration ranging from 0.0012 to 0.06%) greatly suppress the germination and seedling height of test weeds. At 0.06% eucalypt oil concentration, none of the seed of test weeds germinated. Among the weed species tested, A. viridis was found to be the most sensitive and its germination was completed inhibited even at 0.03%. Not only the germination and seedling growth, even the chlorophyll content and respiratory activity in leaves of emerged seedlings were severely affected. In A. viridis chlorophyll content and respiratory activity were reduced by over 51% and 71%, respectively, even at a very low concentration of 0.06%. These results indicated an adverse effect of eucalypt oils on the photosynthetic and energy metabolism of the test weeds. A strong negative correlation was observed between the observed effect and the concentration of eucalypt oil. Based on the study, it can be concluded that oil from E. citriodora possess strong inhibitory potential against weeds that could be exploited for weed management.     

Effects of spacing and post-planting treatments on survival and growth of Fraxinus angustifolia seedlings

This study was conducted at a bottomland hardwood site with heavy textured soil in Akyazi, Turkey to determine the effect of initial spacing (3.0 x 3.0, 3.0 x 2.0, 2.5 x 1.6 and 2.5 x 1.2 m) and post-planting treatments (untreated check, moving, hoeing, disking, and hoeing plus disking) on early survival and growth of Fraxinus angustifolia Vahl. One-year old bare-root seedlings (70 +/- 5 cm in height) were hand-planted in December 2004. Through three years survival was perfect with a rate of 98% in all treatments. Spacing and the interaction between spacing and post-planting treatment did not significantly affect seedling growth through three years. However height and diameter growth increased overtime and differed significantly among post-planting treatments. The hoeing and hoeing plus disking treatments gave the highest growth, and resulted in about 31% increase in diameter and height increment, and in total diameter and height about 20%. These results suggest that post-planting treatments on bottomland sites with heavy textured soil give promising results.     

原油污染土壤的生物法修复效果研究

对原油污染土壤进行生物法修复试验,结果表明,当土壤中含油量分别为5%,10%,20%和45%时,生物降解20d后石油去除率分别达到44.8%,49.9%,47.2%和49.6%;55d后去除率分别达到80.9%,83.6%,87.8%和87.1%;150d后,石油去除率分别达到91.7%,92.6%,91.3%和90.6%.从GC和GC-MS分析结果看,土壤经修复处理后,各种油成分都出现了明显的降解,其中C28以上的分子链基本消失;从脱氢酶分析结果看,微生物活性起初有所上升,后来随着含油量的降低而下降.     

Effect of arbuscular mycorrhizal (AM) fungus and plant growth promoting rhizomicroorganisms (PGPR's) on medicinal plant Solanum viarum seedlings

A green house nursery study was conducted to assess the interaction between arbuscular mycorrhizal (AM) fungus, Glomus aggregatum and some plant growth promoting rhizomicrooganisms (PGPR's), Bacillus coagulans and Trichoderma harzianum, in soil and their consequent effect on growth, nutrition and content of secondary metabolities of Solanum viarum seedlings. Triple inoculation of G. aggregatum+B. coagulans+T. harzainum with Solanum viarum in a green house nursery study resulted in maximum plant biomass (plant height 105 cm and plant dry weight 12.17 g), P, Fe, Zn, Cu and Mn and secondary metabolities [total phenols (129.6 microg g(-1) f.wt.), orthodihydroxy phenols (90.6 microg g(-1) f.wt.), flavonoids (3.94 microg g(-1) f.wt.), alkaloids (5.05 microg g(-1) f.wt.), saponins (5.05 microg g(-1) f.wt.) and tannins (0.324 microg g(-1) f.wt.)] of S. viarum seedlings. The mycorrhizal root colonization and spore numbers in the root zone soil of the inoculated plants increased. The enzyme activity namely acid phosphatase (53.44 microg PNP g(-1) soil), alkaline phosphatase (40.95 microg PNP g(-1) soil) and dehydrogenase (475.5 microg PNP g(-1) soil) and total population of B. coagulans (12.5x10(4) g(-1)) and T. harzianum (12.4 x 10(4) g(-1)), in the root zone soil was found high in the triple inoculation with G. aggregatum+B. coagulans+T. harzianum that proved to be the best microbial consortium.     

Decrease of net carbon flux in two species of mussels caused by extracts of crude oil

Effects of crude oil and salinity stress on the metabolism of two common filter-feeding animals (Mytilus edulis and Modiolus demissus) have been investigated. Carbon budgets have been calculated for each species under a variety of combinations of oil content and salinity. Both reduced salinity and the presence of crude oil tend to decrease the net carbon flux for each species; stresses from each source interacted in their effects on experimental animals. Although similar responses to oil were shown by each species, Mytilus edulis appeared to be slightly more resistant to oil than Modiolus demissus.     

天津地区表层土壤和河流沉积物中甾烷与五环三萜烷的分布特征及污染源分析

分析了天津地区表层土壤和具有代表性的河流沉积物中甾烷和五环三萜烷系列化合物的组成与分布特征，讨论了这类化合物的来源及环境意义。分析表明，在天津地区不同环境功能区表层土壤和河流沉积物中均检测到了甾烷和五环三萜烷系列化合物，但样品之间其含量存在明显的差别；表层土壤与河流沉积物中甾烷、五环三萜烷化合物的组成与石油中的基本一致，样品间这类化合物在饱和烃中的相对含量与正烷烃CPI之间有较好的负相关关系，表明它们主要来源于石油及副产品，可以根据样品中甾、萜化合物与正构烷烃的比值来反映饱和烃中石油烃污染源的贡献。论文根据样品中甾烷、五环三萜烷化合物与正构烷烃的比值，并结合正烷烃CPI参数初步分析了天津地区不同环境功能区表层土壤和河流沉积物中饱和烃污染物的污染源。     

Hydrogen peroxide (H2O2) requirement for the oxidation of crude oil in contaminated soils by a modified Fenton's reagent

The oxidation of soil organic matter (SOM) and total petroleum hydrocarbon were investigated in two soils at eight different hydrogen peroxide (H₂O₂) concentrations to determine the optimal H₂O₂ dosage for the efficient remediation of soils contaminated by crude oil with minimal SOM removal. In our study, H₂O₂ concentrations up to 1100 mM increased the SOM destruction up to 10%–15% in the two soils while no improvement of the crude oil removal efficiencies was observed. The results indicate that the destruction of SOM significantly limits the oxidation of crude oil because SOM might consume H₂O₂ more effectively than crude oil at H₂O₂ concentrations above 1100 mM. In addition, H₂O₂ concentrations higher than 1100 mM were not expected for both soils because of the extremely rapid H₂O₂ decomposition, and low H₂O₂ utilization, of both soils.     

外来入侵植物美洲商陆提取物的化感活性

研究了外来入侵植物美洲商陆(Phytolacca americana)提取物对黄花草木樨(Melilotus officinalis)、大白菜(Brassica pekinensis)、北美车前(Plantago virginica)和马唐(Digitaria sanguinalis)4种植物种子萌发和幼苗生长的影响。结果表明,与对照相比,1 000～4 000、4 000和2 000～4 000μg·mL-1美洲商陆醇提取物分别对大白菜、北美车前和马唐种子萌发率具有显著抑制作用,1 000～4 000μg.mL-1水提取物对马唐种子萌发率也具有显著抑制作用,最大抑制率分别为32.22%、46.56%、32.69%和28.57%。250～500μg·mL-1醇提取物和250～4 000μg·mL-1水提取物对黄花草木樨幼苗根长,250～1 000μg·mL-1醇提取物和250～4 000μg·mL-1水提取物对黄花草木樨幼苗苗高,以及500μg·mL-1水提取物对黄花草木樨幼苗干质量均具有显著促进作用,最大促进率分别为21.32%、37.89%、12.94%、32.07%和18.81%。1 000～4 000μg·mL-1醇提取物和4 000μg·mL-1水提取物对大白菜幼苗根长,2 000～4 000μg·mL-1醇提取物和水提取物对大白菜幼苗苗高,4 000μg·mL-1水提取物对大白菜幼苗干质量具有显著抑制作用,最大抑制率分别为37.00%、20.95%、25.63%、22.30%和13.67%。250～1 000μg·mL-1水提取物对北美车前幼苗根长,250～4 000μg·mL-1水提取物对北美车前苗高具有显著促进作用,而1 000～4 000μg·mL-1醇提取物对北美车前幼苗根长,4 000μg·mL-1醇提取物对北美车前苗高和干质量具有显著抑制作用,最大抑制率分别为58.06%、29.72%和18.36%。500～4 000、1 000～4 000和2 000～4 000μg·mL-1醇提取物分别对马唐幼苗根长、苗高及干质量,4 000μg·mL-1水提取物对马唐幼苗根长具有显著抑制作用,最大抑制率分别为70.31%、44.65%、42.43%和35.50%。由此可见,美洲商陆对大白菜、北美车前和马唐种子萌发和幼苗生长具有较强的化感抑制作用,对入侵植物黄花草木樨具有化感促进作用,这些都直接或间接地增强了美洲商陆的入侵性。     

污泥堆肥利用中AM对稗草生长及Cu、Pb吸收的影响

在滨海盐渍土中分别混加质量分数为0%、20%、40%和60%的污泥堆肥,作为盆栽基质,研究摩西球囊霉（Glomusmosseae）和根内球囊霉（Glomus intraradices）2种AM真菌对稗草（Echinochloa crusgalli）生长及其吸收Cu、Pb的影响,分别以不接种AM真菌的处理为各自的对照。结果显示：添加污泥堆肥处理中稗草接种苗菌根侵染率均显著高于纯盐泽土处理。同时,随着盐渍土中污泥堆肥含量增加稗草生物量上升,其中在含有40%和60%污泥堆肥处理中接种AM真菌稗草的地上及地下部生物量显著高于未接种苗。接种AM真菌显著提高了稗草Cu、Pb富集总量;接种AM真菌显著提高了稗草地下部Cu富集量,却降低了地下部Pb累积量,提高了Pb向地上部的转运,增加了地上部Pb累积量。这些结果表明污泥堆肥中接种AM真菌可以促进稗草的生长和提高对重金属Cu、Pb的富集能力。