In Situ Discrimination and Cultivation of Active Degraders in Soils by Genome-Directed Cultivation Assisted by SIP-Raman-Activated Cell Sorting.

The identification and in situ cultivation of functional yet uncultivable microorganisms are important to confirm inferences regarding their ecological functions. Here, we developed a new method that couples Raman-activated cell sorting (RACS), stable-isotope probing (SIP), and genome-directed cultivation (GDC)─namely, RACS-SIP-GDC─to identify, sort, and cultivate the active toluene degraders from a complex microbial community in petroleum-contaminated soil. Using SIP, we successfully identified the active toluene degrader Pigmentiphaga, the single cells of which were subsequently sorted and isolated by RACS. We further successfully assembled the genome of Pigmentiphaga based on the metagenomic sequencing of 13C-DNA and genomic sequencing of sorted cells, which was confirmed by gyrB gene comparison and average nucleotide identity determination. Additionally, the genotypes and phenotypes of this degrader were directly linked at the single-cell level, and its complete toluene metabolic pathways in petroleum-contaminated soil were reconstructed. Based on its unique metabolic properties uncovered by genome sequencing, we modified the traditional cultivation medium with antibiotics, amino acids, carbon sources, and growth factors (e.g., vitamins and metals), achieving the successful cultivation of RACS-sorted active degrader Pigmentiphaga sp. Our results implied that RACS-SIP-GDC is a state-of-the-art approach for the precise identification, targeted isolation, and cultivation of functional microbes from complex communities in natural habitats. RACS-SIP-GDC can be used to explore specific and targeted organic-pollution-degrading microorganisms at the single-cell level and provide new insights into their biodegradation mechanisms.

Synergistic/antagonistic toxicity characterization and source-apportionment of heavy metals and organophosphorus pesticides by the biospectroscopy-bioreporter-coupling approach.

Many anthropogenic chemicals are manufactured and eventually enter the surrounding environment, threatening food security and human health. Considering the additive or synergistic effects of pollutant mixtures, there is an expanding need for rapid, cost-effective and field-portable screening methods in environmental monitoring. This study used a recently developed biospectroscopy-bioreporter-coupling (BBC) approach to investigate the binary toxicity of Ag(I), Cr(VI) and four organophosphorus pesticides (dichlorvos, parathion, omethoate and monocrotophos). Ag(I) and Cr(VI) altered the toxicity mechanisms of pesticides, explained by the synergistic or antagonistic effect of Ag/Cr-induced cytotoxicity and pesticide-induced genotoxicity. The discriminating Raman spectral peaks associated with organophosphorus pesticides were 1585 and 1682 cm-1, but 750, 1004, 1306 and 1131 cm-1 were found in heavy metal and pesticide mixtures. More spectral alterations were related to pesticides rather than Ag(I) or Cr(VI), hinting at the dominant toxicity mechanisms of pesticides in mixtures. Ag(I) supplement significantly increased the levels of reactive oxygen species induced by organophosphorus pesticides, attributing to the increased permeability of cell membrane and entrance of toxic substances into the cells by the oligodynamic actions. This study lends deeper insights into the interactions between microbes and pollutant mixtures, offering clues to assess the cocktail effects of multiple pollutants comprehensively.

Dynamics of dominant rhizospheric microbial communities responsible for trichlorfon absorption and translocation in maize seedlings.

In this study, the available phosphorus (AP) and TCF concentrations in soils and maize (Zea mays) seedling tissues were measured in response to escalating TCF concentrations during 216 hr of culture. Maize seedlings growth considerably enhanced soil TCF degradation, reaching the highest of 73.2% and 87.4% at 216 hr in 50 and 200 mg/kg TCF treatments, respectively, and increased AP contents in all the seedling tissues. Soil TCF was majorly accumulated in seedling roots, reaching maximum concentration of 0.017 and 0.076 mg/kg in TCF-50 and TCF-200, respectively. The hydrophilicity of TCF might hinder its translocation to the aboveground shoot and leaf. Using bacterial 16 S rRNA gene sequencing, we found that TCF addition drastically lessened bacterial community interactions and hindered the complexity of their biotic networks in rhizosphere than in bulk soils, leading to the homogeneity of bacterial communities that were resistant or prone to TCF biodegradation. Mantel test and redundancy analysis suggested a significant enrichment of dominant species Massilia belonging to Proteobacteria phyla, which in turn affecting TCF translocation and accumulation in maize seedling tissues. This study provided new insight into the biogeochemical fate of TCF in maize seedling and the responsible rhizobacterial community in soil TCF absorption and translocation.

New insight into the mechanisms of autochthonous fungal bioaugmentation of phenanthrene in petroleum contaminated soil by stable isotope probing.

Autochthonous fungal bioaugmentation (AFB) is considered a reliable bioremediation approach for polycyclic aromatic hydrocarbon (PAH) contamination, but little is known about its mechanisms in contaminated soils. Here, a microcosm experiment was performed to explore the AFB mechanisms associated with two highly efficient phenanthrene degrading agents of fungi (with laccase-producing Scedosporium aurantiacum GIG-3 and non-laccase-producing Aspergillus fumigatus LJD-29), using stable-isotope-probing (SIP) and high-throughput sequencing. The results showed that each fungus markedly improved phenanthrene removal, and microcosms with both fungi exhibited the best phenanthrene removal performance among all microcosms. Additionally, AFB markedly shifted the composition of the microbial community, particularly the phenanthrene-degrading bacterial taxa. Interestingly, based on SIP results, strains GIG-3 and LJD-29 did not assimilate phenanthrene directly during AFB, but instead played key roles in the preliminary decomposition of phenanthrene though secretion of different extracellular enzymes to oxidize the benzene ring (GIG-3 bioaugmentation with laccase, and LJD-29 bioaugmentation with manganese and lignin peroxidases). In addition, all functional degraders directly involved in phenanthrene assimilation were indigenous bacteria, while native fungi rarely participated in the direct phenanthrene mineralization. Our findings provide a new mechanism of AFB with multiple fungi, and support AFB as a promising strategy for the in situ bioremediation of PAH-contaminated soil.

Response of soil microbial communities to petroleum hydrocarbons at a multi-contaminated industrial site in Lanzhou, China.

Total petroleum hydrocarbon (TPH) contamination poses threats to ecological systems and human health. Many studies have reported its negative impacts on soil microbes, but limited information is known about microbial change and response to multiple TPH contamination events. In this study, we investigated TPH contamination level, microbial community structure and functional genes at a multi-contaminated industrial site in Lanzhou, where a benzene spill accident caused the drinking water crisis in 2014. TPHs distribution in soils and groundwater indicated multiple TPH contamination events in history, and identified the spill location where high TPH level (6549 mg kg-1) and high ratio of low-molecular-weight TPHs (>80%) were observed. In contrast, TPH level was moderate (349 mg kg-1) and the proportion of low-molecular-weight TPHs was 44% in soils with a long TPH contamination history. After the spill accident, soil bacterial communities became significant diverse (p = 0.047), but the dominant microbes remained the same as Pseudomonadaceae and Comamonadaceae. The abundance of hydrocarbon-degradation related genes increased by 10-1000 folds at the site where the spill accident occurred in multi-contaminated areas and was significantly related to 2-ring PAHs. Such changes of microbial community and hydrocarbon-degradation related genes together indicated the resilience of soil indigenous microbes toward multiple contamination events. Our results proved the significant change of bacterial community and huge shift of hydrocarbon-degradation related genes after the spill accident (multiple contamination events), and provided a deep insight into microbial response at industrial sites with a long period of contamination history.

In-vitro toxicity assessment of Eucalyptus robusta Smith extracts via whole-cell bioreporter.

Eucalyptus is widely planted in China for wood industries, and there are increasing concerns about its ecotoxicity in the environment. This study explored the in-vitro toxicity of Eucalyptus extracts by assessing the impacts of water-soluble and dimethylsulfoxide (DMSO)-soluble fractions via a whole-cell bioreporter, Acinetobacter baylyi ADPWH_recA. Compounds identified in Eucalyptus extracts included one tannin, two phenolic acids, four terpenoids, four glycosides, and five flavonoids. The leaf extracts contained more biological-active components than barks and roots. Genotoxicity induced by Eucalyptus extracts was mainly associated with water extracts (e.g., flavonoids, phenolic acids) instead of DMSO extracts. The significant cytotoxicity was explained by programmed cell death (PCD), suggested by the results of propidium iodide (PI) and 2',7'-dichlorofluorescein-diacetate (DCFH-DA) assays. Generally, water-soluble fractions contributed more toxicities than DMSO-soluble fractions, particularly at high concentrations. A robust linear regression was built between the compromised toxicity and PCD index (Compromised toxicity = -2.192 × PCD index + 2.219; R2 = 0.8886), suggesting a PCD-dependent compromised toxicity which was greatly underestimated. Our results implied non-neglectable ecotoxicological risks of Eucalyptus extracts, hinting at the possible magnified ecological impacts of its large-scale plantation and the potential adverse outcomes to the surrounding ecosystems.

The spatiotemporal variations in microalgae communities in vertical waters of a subtropical reservoir.

The construction of cascade reservoirs increases eutrophication and exacerbates algal blooms and thus threatens water quality. Previous studies on the microalgae in reservoir have mainly focused on the spatio-temporal patterns of surface microalgae communities at the horizontal scale, while few studies have simultaneously considered the successions of microalgae in vertical profiles including the sediments and the effects of the nutrients release and microalgae in sediments on microalgae in upper waters. In this study, we investigated the effects of microalgae and physico-chemical parameters in waters and sediments on the successions of vertical microalgae communities in Xipi Reservoir, Southeast China. The seasonal variations in microalgae compositions decreased gradually from the surface water (the dominance of Cryptophyta and Chlorophyta in spring, Chlorophyta and Cyanophyta in summer, and relatively uniform in autumn and winter) to the sediment (the dominance of Bacillariophyta throughout the year), which was influenced by the variations of physico-chemical factors in different layers. The spatio-temporal variations in microalgae communities in waters was attributing to not only the heterogeneities of the stratification, and the physico-chemical factors such as water temperature, pH, and nutrient concentrations, especially for phosphorus in the water column, but also the combinations of phosphorus release and microalgae composition in sediments. Environmental changes would be especially problematic for microalgae groups such as Cryptophyta, Dinophyta and Chlorophyta that were sensitive to the changes of temperature and nutrients. Our results are helpful for an extensive understanding of the dynamics of microalgae communities in reservoir, and contribute to reservoir management for ensuring the safety of drinking water.

Impacts of groundwater level fluctuation on soil microbial community, alkane degradation efficiency and alkane-degrading gene diversity in the critical zone: Evidence from an accelerated water table fluctuation simulation.

Petroleum hydrocarbons are hazardous to ecosystems and human health, commonly containing n-alkanes and polycyclic aromatic hydrocarbons. Previous researches have studied alkane degraders and degrading genes under aerobic or anaerobic conditions, but seldom discussed them in the intermittent saturation zone which is a connective area between the vadose zone and the groundwater aquifer with periodic alteration of oxygen and moisture. The present study investigated the difference in alkane degradation efficiency, bacterial community, and alkane degrading gene diversity in aerobic, anaerobic, and aerobic-anaerobic fluctuated treatments. All biotic treatments achieved over 90% of n-alkane removal after 120 days of incubation. The removal efficiencies of n-alkanes with a carbon chain length from 16 to 25 were much higher in anaerobic scenarios than those in aerobic scenarios, explained by different dominant microbes between aerobic and anaerobic conditions. The highest removal efficiency was found in fluctuation treatments, indicating an accelerated n-alkane biodegradation under aerobic-anaerobic alternation. In addition, the copy numbers of the 16S rRNA gene and two alkB genes (alkB-P and alkB-R) declined dramatically when switched from aerobic to anaerobic scenarios and oppositely from anaerobic to aerobic conditions. This suggested that water level fluctuation could notably change the presence of aerobic alkane degrading genes. Our results suggested that alkane degradation efficiency, soil microbial community, and alkane-degrading genes were all driven by water level fluctuation in the intermittent saturation zone, helping better understand the effects of seasonal water table fluctuation on the biodegradation of petroleum hydrocarbons in the subsurface environment.

Correlation of bacterial community with phosphorus fraction drives discovery of Actinobacteria involved soil phosphorus transformation during the trichlorfon degradation.

Trichlorfon (TCF) is a broad-spectrum phosphorus (P)-containing pesticide, yet its effects on soil P fraction transformation and bacterial communities during the TCF degradation in soils is unknown. In this study, we investigated soil TCF degradation behavior at different contents of 50, 100 and 200 mg/kg, and analyzed residual TCF contents and metabolites by gas chromatography mass spectrometry after 216-h incubation. Our results suggested that TCF was gradually degraded in soils and was be initially hydrolyzed to dichlorvos via P-C bond cleavage and then other P-containing metabolites. By analyzing different P fractions and soil microbial community composition, we found significant increases of soil available phosphorus contents from 2.76 mg/kg (control) to 3.23 mg/kg (TCF-50), 5.12 mg/kg (TCF-100) and 5.72 mg/kg (TCF-200), respectively. Inorganic CaCl2-P was easily and instantly transformed to primary mineral inorganic P (Pi) forms of HCl-P and citrate-P, while the proportion of enzyme-P (a labile organic P) fluctuated throughout TCF degradation process. Soil available P contents and Pi fractions were significantly correlated with the relative abundance of Actinobacteria. These results highlighted that Actinobacteria is the dominant soil species utilizing TCF as P sources to increase its community richness, and subsequently affect the transformation of P fractions to regulate soil P cycle. Our study gives new understanding on the microorganisms can involve soil P transformation during organophosphorus pesticides degradation in soils, highlighting the importance of bacteria in P transformation and pesticides soil decontamination.

The catabolic pathways of in situ rhizosphere PAH degraders and the main factors driving PAH rhizoremediation in oil-contaminated soil.

Rhizoremediation is a potential technique for polycyclic aromatic hydrocarbon (PAH) remediation; however, the catabolic pathways of in situ rhizosphere PAH degraders and the main factors driving PAH rhizoremediation remain unclear. To address these issues, stable-isotope-probing coupled with metagenomics and molecular ecological network analyses were first used to investigate the phenanthrene rhizoremediation by three different prairie grasses in this study. All rhizospheres exhibited a significant increase in phenanthrene removal and markedly modified the diversity of phenanthrene degraders by increasing their populations and interactions with other microbes. Of all the active phenanthrene degraders, Marinobacter and Enterobacteriaceae dominated in the bare and switchgrass rhizosphere respectively; Achromobacter was markedly enriched in ryegrass and tall fescue rhizospheres. Metagenomes of 13 C-DNA illustrated several complete pathways of phenanthrene degradation for each rhizosphere, which clearly explained their unique rhizoremediation mechanisms. Additionally, propanoate and inositol phosphate of carbohydrates were identified as the dominant factors that drove PAH rhizoremediation by strengthening the ecological networks of soil microbial communities. This was verified by the results of rhizospheric and non-rhizospheric treatments supplemented with these two substances, further confirming their key roles in PAH removal and in situ PAH rhizoremediation. Our study offers novel insights into the mechanisms of in situ rhizoremediation at PAH-contaminated sites.

History of petroleum disturbance triggering the depth-resolved assembly process of microbial communities in the vadose zone.

Biogeochemical gradient forms in vadose zone, yet little is known about the assembly processes of microbial communities in this zone under petroleum disturbance. This study collected vadose zone soils at three sites with 0, 5, and 30 years of petroleum contamination to unravel the vertical microbial community successions and their assembly mechanisms. The results showed that petroleum hydrocarbons exhibited higher concentrations at the long-term contaminated site, showing negative impacts on some soil properties, retarding in the surface soils and decreasing along soil depth. Cultivable fraction of heterotrophic bacteria and microbial α-diversity decreased along depth in vadose zones with short-term/no contamination history, but exhibited an opposite trend with long-term contamination history. Petroleum contamination intensified the vertical heterogeneity of microbial communities based on the contamination time. Microbial co-occurrence network revealed the lowest species co-occurrence pattern at the long-term contaminated site. The distance-decay patterns and null model analysis together suggested distinct assembly mechanisms at three sites, where dispersal limitation (42-45%) was higher and variable and homogenizing selections were lower (37-38%) in vadose zones under petroleum disturbance than those in the uncontaminated vadose zone. Our findings help to better understand the subsurface biogeochemical cycles and bioremediation of petroleum-contaminated vadose zones.

Significant influence of phosphorus resources on the growth and alkaline phosphatase activities of Microcystis aeruginosa.

It is well-accepted that phosphorus, particularly orthophosphate, is a determinant factor in aquatic eutrophication. However, numerous kinds of phosphorus sources exist in real world scenario, and limited studies have characterized the pairwise relationships among abundant different phosphorus sources and the physiological behaviour of algae. The present study developed a high-throughput assay to investigate the effects of 59 different phosphorus sources (equal initial concentration of total phosphorus) on the growth and alkaline phosphatase (AKP) activities of Microcystis aeruginosa, a model cyanobacteria whose predominance holds sway in lake eutrophication. M. aeruginosa cultivated with nucleoside monophosphates (NMPs) had higher growth, relative AKP activities and residual orthophosphate, which were positively intercorrelated. Oppositely, non-NMPs cultivation of M. aeruginosa led to negative relationships between the relative AKP activities and their growth or residual orthophosphate. These results indicated distinct mechanisms for M. aeruginosa to utilize different phosphorus sources in real-world scenario, and both phosphorus source and content are determinant factors on the growth and physiological behaviour of M. aeruginosa. Given the complicated and vast phosphorus pool in the natural environment, phosphorus resources might significantly alter the abundance and physiological behaviour of M. aeruginosa and other bloom-forming algae, then influence the phytoplanktonic community structure and affect the possibility and intensity of algal bloom. Our work hints the underestimation of the restriction factors in lake eutrophication and provides a new tool to study the driven forces of phytoplanktonic community dynamics as phosphorus from both internal and external sources.

Applying Raman Microspectroscopy to Evaluate the Effects of Nutrient Cations on Alkane Bioavailability to Acinetobacter baylyi ADP1.

Contamination with petroleum hydrocarbons causes extensive damage to ecological systems. On oil-contaminated sites, alkanes are major components; many indigenous bacteria can access and/or degrade alkanes. However, their ability to do so is affected by external properties of the soil, including nutrient cations. This study used Raman microspectroscopy to study how nutrient cations affect alkanes' bioavailability to Acinetobacter baylyi ADP1 (a known degrader). Treated with Na, K, Mg, and Ca at 10 mM, A. baylyi was exposed to seven n-alkanes (decane, dodecane, tetradecane, hexadecane, nonadecane, eicosane, and tetracosane) and one alkane mixture (mineral oil). Raman spectral analysis indicated that bioavailability of alkanes varied with carbon chain lengths, and additional cations altered the bacterial response to n-alkanes. Sodium significantly increased the bacterial affinity toward decane and dodecane, and K and Mg enhanced the bioavailability of tetradecane and hexadecane. In contrast, the bacterial response was inhibited by Ca for all alkanes. Similar results were observed in mineral oil exposure. Our study employed Raman spectral assay to offer a deep insight into how nutrient cations affect the bioavailability of alkanes, suggesting that nutrient cations can play a key role in influencing the harmful effects of hydrocarbons and could be optimized to enhance the bioremediation strategy.

Geo-distribution pattern of microbial carbon cycling genes responsive to petroleum contamination in continental horizontal oilfields.

Contamination significantly affects soil microbial community structures, and the metabolisms of organic contaminants might particularly alter soil carbon cycling by shaping microbial carbon cycling genes. Although numerous studies have discussed the impacts of petroleum contamination on soil bacterial communities and relevant degrading genes, there is no work addressing how soil carbon cycling genes are affected by petroleum contamination. In this study, 77 soil samples were collected from five typical oilfields horizontally located in China to explore the influence of environmental variables and petroleum contamination on microbial carbon cycling genes. Results from Geochip suggested a geographic-determined distribution of carbon cycling genes. Although no significant correlation was observed between carbon cycling genes and soil physio-chemical properties for all soils, some relationships were identified in specific oilfield. Principle component analysis indicated that soil physio-chemical properties, rather than petroleum contamination disturbance, are the key factors determining the degree of sample dispersion, whereas environmental variables predominantly control the degree of sample aggregation. Co-occurrence ecological network analysis revealed a more complex interactions of all functional genes in petroleum-contaminated soils, and carbon cycling genes were grouped with nitrogen related genes in petroleum-contaminated communities. Soil moisture and heterogeneity were identified as the main drivers for the abundance and diversity of carbon cycling genes, particularly in petroleum-contaminated soils. These results are attributing to the fewer impacts of petroleum contamination on the diversity of carbon cycling genes than soil physio-chemical properties, and soil carbon cycling genes are mainly driven by geographic location and petroleum contamination together. Our findings provide deeper insight into the influence of petroleum contamination in soil microbial functions related to carbon cycling.

Roles of sulfur compounds in growth and alkaline phosphatase activities of Microcystis aeruginosa under phosphorus deficiency stress.

Microcystis aeruginosa is one of the most common algae found in eutrophicated water bodies. Alkaline phosphatase (AKP) can be produced by Microcystis aeruginosa to utilize organic phosphates under phosphorus deficiency stress, thereby AKP can be regarded as an important indicator for algal growth. Sulfur compounds are ubiquitous in waters, while investigation on the interactions between sulfur compounds and Microcystis aeruginosa is limited. In this work, we introduced 33 types of sulfur compounds to culture Microcystis aeruginosa, and the results demonstrated that algal growth is positively related to AKP activities. Toxicity of organic sulfur compounds was further evaluated using Toxicity Estimation Software Tool based on quantitative structure-activity relationship prediction. The algal growth results exhibited strong correlation to the toxicity endpoints suggesting the organic sulfur compounds inhibits the algal growth as toxic matters. K-means cluster analyses have been carried out subsequently via Python based on the results of algal growth and AKP activities of each sample and statistically, the sulfur compounds can be adequately clustered into 2 groups. According to clustering results, sulfonic acids exhibit low toxicity while sulfur amino acids can be considered as more toxic compounds. Graphical abstract Varied sulfur compounds (33 types) were investigated to find out the interactions between them and Microcystis aeruginosa, a common alga. K-means cluster and correlation analyses demonstrate that algal growth and alkaline phosphatase activities exhibited strong correlation to the predicted toxicity endpoints.

A whole-cell bioreporter assay for quantitative genotoxicity evaluation of environmental samples.

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.

Separating and characterizing functional alkane degraders from crude-oil-contaminated sites via magnetic nanoparticle-mediated isolation.

Uncultivable microorganisms account for over 99% of all species on the planet, but their functions are yet not well characterized. Though many cultivable degraders for n-alkanes have been intensively investigated, the roles of functional n-alkane degraders remain hidden in the natural environment. This study introduces the novel magnetic nanoparticle-mediated isolation (MMI) technology in Nigerian soils and successfully separates functional microbes belonging to the families Oxalobacteraceae and Moraxellaceae, which are dominant and responsible for alkane metabolism in situ. The alkR-type n-alkane monooxygenase genes, instead of alkA- or alkP-type, were the key functional genes involved in the n-alkane degradation process. Further physiological investigation via a BIOLOG PM plate revealed some carbon (Tween 20, Tween 40 and Tween 80) and nitrogen (tyramine, l-glutamine and d-aspartic acid) sources promoting microbial respiration and n-alkane degradation. With further addition of promoter carbon or nitrogen sources, the separated functional alkane degraders significantly improved n-alkane biodegradation rates. This suggests that MMI is a promising technology for separating functional microbes from complex microbiota, with deeper insight into their ecological functions and influencing factors. The technique also broadens the application of the BIOLOG PM plate for physiological research on functional yet uncultivable microorganisms.

Assessing the impacts of phosphorus inactive clay on phosphorus release control and phytoplankton community structure in eutrophic lakes.

Addressing the challenge that phosphorus is the key factor and cause for eutrophication, we evaluated the phosphorus release control performance of a new phosphorus inactive clay (PIC) and compared with Phoslock®. Meanwhile, the impacts of PIC and Phoslock® on phytoplankton abundance and community structure in eutrophic water were also discussed. With the dosage of 40 mg/L, PIC effectively removed 97.7% of total phosphorus (TP) and 98.3% of soluble reactive phosphorus (SRP) in eutrophic waters. In sediments, Fe/Al-phosphorus and organic phosphorus remained stable whereas Ca-phosphorus had a significant increase of 13.1%. The results indicated that PIC may form the active overlay at water-sediment interface and decrease the bioavailability of phosphorus. The phytoplankton abundance was significantly reduced by PIC and decreased from (1.0-2.4) × 107 cells/L to (1.3-4.3) × 106 cells/L after 15 d simultaneous experiment. The phytoplankton community structure was also altered, where Cyanobacteria and Bacillariophyceae were the most inhibited and less dominant due to their sensitivity to phosphorus. After PIC treatment, the residual lanthanum concentration in water was 1.44-3.79 µg/L, and the residual aluminium concentration was low as 101.26-103.72 µg/L, which was much less than the recommended concentration of 200 µg/L. This study suggests that PIC is an appropriate material for phosphorus inactivation and algal bloom control, meaning its huge potential application in eutrophication restoration and management.

Toxicity assessment and modelling of Moringa oleifera seeds in water purification by whole cell bioreporter.

Moringa oleifera has been used as a coagulation reagent for drinking water purification, especially in developing countries such as Malawi. This research revealed the cytoxicity and genotoxicity of M. oleifera by Acinetobacter bioreporter. The results indicated that significant cytoxicity effects were observed when the powdered M. oleifera seeds concentration is from 1 to 50 mg/L. Through direct contact, ethanolic-water extraction and hexane extraction, the toxic effects of hydrophobic and hydrophilic components in M. oleifera seeds were distinguished. It suggested that the hydrophobic lipids contributed to the dominant cytoxicity, consequently resulting in the dominant genotoxicity in the water-soluble fraction due to limited dissolution when the M. oleifera seeds granule concentration was from 10 to 1000 mg/L. Based on cytoxicity and genotoxicity model, the LC50 and LC90 of M. oleifera seeds were 8.5 mg/L and 300 mg/L respectively and their genotoxicity was equivalent to 8.3 mg mitomycin C per 1.0 g dry M. oleifera seed. The toxicity of M. oleifera has also remarkable synergistic effects, suggesting whole cell bioreporter as an appropriate and complementary tool to chemical analysis for environmental toxicity assessment.

Whole cell bioreporter application for rapid detection and evaluation of crude oil spill in seawater caused by Dalian oil tank explosion.

Accidents involving the release of crude oil to seawater pose serious threat to human and animal health, fisheries and marine ecosystems. A whole cell bioreporter detection method, which has unique advantages for the rapid evaluation on toxicity and bioavailability, is a useful tool to provide environmental risk assessments at crude oil-contaminated sites. Acinetobacter baylyi ADPWH_alk and ADPWH_recA are chromosomally-based alkane and genotoxicity bioreporters which can be activated to express bioluminescence in the presence of alkanes and genotoxic compounds. In this study, we applied Acinetobacter ADPWH_alk and ADPWH_recA bioreporters to examine six seawater and six sediment samples around the Dalian Bay four weeks after an oil tank explosion in Dalian, China in 2010, and compared the results with samples from the same sites one year after. The results of bioreporter detection suggest that seawater and sediments from five sites (DB, NT, JSB, XHP and FJZ) four weeks after the oil-spill were contaminated by the crude oil with various extents of genotoxicity. Among these six sites, DB and NT had high oil contents and genotoxicity, and JSB had high oil content but low genotoxicity in comparison with an uncontaminated site LSF, which is located at other side of the peninsula. These three sites (DB, NT and JSB) with detectable genotoxicity are within 30 km away from the oil spill point. The far-away two sites XHP (38.1 km) and FJZ (31.1 km) were lightly contaminated with oil but no genotoxicity suggesting that they are around the contamination boundary. Bioreporter detection also indicates that all six sites were clean one year after the oil-spill as the alkane and genotoxicity were below detection limit. This study demonstrates that bioreporter detection can be used as a rapid method to estimate the scale of a crude oil spill accident and to evaluate bioavailability and genotoxicity of contaminated seawater and sediments, which are crucial to risk assessment and strategic decision-making for environmental management and clean-up.