Influence of easily degradable naturally occurring carbon substrates on biodegradation of monosubstituted phenols by aquatic bacteria

The influence of readily degradable, naturally occurring carbon substrates on the biodegradation of several monosubstitued phenols (m-cresol, m-aminophenol, p-chlorophenol) was examined. The natural substrate classes used were amino acids, carbohydrates, and fatty acids. Samples of the microbial community from Lake Michie, a mesotrophic reservoir, were adapted to different levels of representatives from each natural substrate class in chemostats. After an extended adaptation period, the ability of the microbial community to degrade the monosubstituted phenols was determined by using a radiolabeled substrate uptake and mineralization method. Several microbiological characteristics of the communities were also measured. Adaptation to increasing concentrations of amino acids, carbohydrates, or fatty acids enhanced the ability of the microbial community to degrade all three phenols. The stimulation was largest for m-cresol and m-aminophenol. The mechanism responsible for the enhancement of monosubstituted phenol metabolism was not clearly identified, but the observation that adaptation to amino acids also increased the biodegradation of glucose and, to a lesser extent, naphthalene suggests a general stimulation of microbial metabolism. This study demonstrates that prior exposure to labile, natural substrates can significantly enhance the ability of aquatic microbial communities to respond to xenobiotics.     

Adaptation to and biodegradation of xenobiotic compounds by microbial communities from a pristine aquifer

The ability of subsurface microbial communities to adapt to the biodegradation of xenobiotic compounds was examined in aquifer solids samples from a pristine aquifer. An increase in the rates of mineralization of radiolabeled substrates with exposure was used as an indication of adaptation. For some compounds, such as chlorobenzene and 1,2,4-trichlorobenzene, slight mineralization was observed but no adaptation was apparent during incubations of over 8 months. Other compounds demonstrated three patterns of response. For m-cresol, m-aminophenol, and aniline intermediate rates of biodegradation and a linear increase in the percent mineralized with time were observed. Phenol, p-chlorophenol, and ethylene dibromide were rapidly metabolized initially, with a nonlinear increase in the percent mineralized with time, indicating that the community was already adapted to the biodegradation of these compounds. Only p-nitrophenol demonstrated a typical adaptation response. In different samples of soil from the same layer in the aquifer, the adaptation period to p-nitrophenol varied from a few days to as long as 6 weeks. In most cases the concentration of xenobiotic added, over the range from a few nanograms to micrograms per gram, made no difference in the response. Most-probable-number counts demonstrated that adaptation is accompanied by an increase in specific degrader numbers. This study has shown that diverse patterns of response occur in the subsurface microbial community.     

Adaptation to and biodegradation of xenobiotic compounds by microbial communities from a pristine aquifer.

The ability of subsurface microbial communities to adapt to the biodegradation of xenobiotic compounds was examined in aquifer solids samples from a pristine aquifer. An increase in the rates of mineralization of radiolabeled substrates with exposure was used as an indication of adaptation. For some compounds, such as chlorobenzene and 1,2,4-trichlorobenzene, slight mineralization was observed but no adaptation was apparent during incubations of over 8 months. Other compounds demonstrated three patterns of response. For m-cresol, m-aminophenol, and aniline intermediate rates of biodegradation and a linear increase in the percent mineralized with time were observed. Phenol, p-chlorophenol, and ethylene dibromide were rapidly metabolized initially, with a nonlinear increase in the percent mineralized with time, indicating that the community was already adapted to the biodegradation of these compounds. Only p-nitrophenol demonstrated a typical adaptation response. In different samples of soil from the same layer in the aquifer, the adaptation period to p-nitrophenol varied from a few days to as long as 6 weeks. In most cases the concentration of xenobiotic added, over the range from a few nanograms to micrograms per gram, made no difference in the response. Most-probable-number counts demonstrated that adaptation is accompanied by an increase in specific degrader numbers. This study has shown that diverse patterns of response occur in the subsurface microbial community.     

Biodegradation of aniline, anthracene, chlornitrophen, fenitrothion and linear alkylbenzene sulphonate in pond water

Biodegradation of five chemicals (aniline, anthracene, chlornitrophen (CNP), fenitrothion (FNT) and linear alkylbenzene sulphonate (LAS)) by aquatic bacteria in three different types of ponds was determined according to the cultivation method developed by this group. The degradability toward these chemicals was varied among the ponds, except for LAS which was decomposed well in all samples. Higher degradability towards the two agrochemicals, CNT and FNT, was found in the pond surrounded by paddy fields, whereas aniline and anthracene were decomposed more rapidly in the pond located in the industrial area. Water from the pond in the botanical garden, with the least exposure to any chemicals, exhibited the lowest degradation toward all chemicals tested. There was no significant seasonal variation in the biodegradation of chemicals in these ponds. It was deduced that biodegradability toward certain chemicals could be a result of acclimatization of the microbial community by chemical contamination present and past, suggesting the possible use of biodegradation profiles as an indicator for chemical pollution in the aquatic environment.     

Influence of Naturally Occurring Humic Acids on Biodegradation of Monosubstituted Phenols by Aquatic Bacteria

Samples of the microbial community from Lake Michie, a mesotrophic reservoir in central North Carolina, were adapted to various levels (100 to 1,000 μg/liter) of natural humic acids in chemostats. The humic acids were extracted from water samples from Black Lake, a highly colored lake in the coastal plain of North Carolina. After adaptation, the microbial community was tested for its ability to degrade the monosubstituted phenols m-cresol, m-aminophenol, and p-chlorophenol. Adaptation to increasing levels of humic acids significantly reduced the ability of the microbial communities to degrade all three phenols. The decline in biodegradation was accompanied by a decrease in the number of specific compound degraders in the adapted communities. Short-term exposure of the community to increasing levels of humic acids had no significant effect on the ability of the community to degrade m-cresol. Thus the suppressive effect of humic acids on monosubstituted phenol metabolism was the result of long-term exposure to the humic materials. Increasing the levels of inorganic nutrients fed to the chemostats during the humic acid adaptation had little effect on the suppressive influence of the humic acids, indicating that nutrient limitation was probably not responsible for the metabolic suppression. The results of the study suggest that long-term exposure to humic acids can reduce the ability of microbial communities to respond to monosubstituted phenols.     

Use of field-based stable isotope probing to identify adapted populations and track carbon flow through a phenol-degrading soil microbial community

The goal of this field study was to provide insight into three distinct populations of microorganisms involved in in situ metabolism of phenol. Our approach measured 13CO2 respired from [13C]phenol and stable isotope probing (SIP) of soil DNA at an agricultural field site. Traditionally, SIP-based investigations have been subject to the uncertainties posed by carbon cross-feeding. By altering our field-based, substrate-dosing methodologies, experiments were designed to look beyond primary degraders to detect trophically related populations in the food chain. Using gas chromatography-mass spectrometry (GC/MS), it was shown that (13)C-labeled biomass, derived from primary phenol degraders in soil, was a suitable growth substrate for other members of the soil microbial community. Next, three dosing regimes were designed to examine active members of the microbial community involved in phenol metabolism in situ: (i) 1 dose of [13C]phenol, (ii) 11 daily doses of unlabeled phenol followed by 1 dose of [13C]phenol, and (iii) 12 daily doses of [13C]phenol. GC/MS analysis demonstrated that prior exposure to phenol boosted 13CO2 evolution by a factor of 10. Furthermore, imaging of 13C-treated soil using secondary ion mass spectrometry (SIMS) verified that individual bacteria incorporated 13C into their biomass. PCR amplification and 16S rRNA gene sequencing of 13C-labeled soil DNA from the 3 dosing regimes revealed three distinct clone libraries: (i) unenriched, primary phenol degraders were most diverse, consisting of alpha-, beta-, and gamma-proteobacteria and high-G+C-content gram-positive bacteria, (ii) enriched primary phenol degraders were dominated by members of the genera Kocuria and Staphylococcus, and (iii) trophically related (carbon cross-feeders) were dominated by members of the genus Pseudomonas. These data show that SIP has the potential to document population shifts caused by substrate preexposure and to follow the flow of carbon through terrestrial microbial food chains.     

Microbial Community Changes Elicited by Exposure to Cyanobacterial Allelochemicals

An increasing body of evidence points out that allelopathy may be an important process shaping microbial communities in aquatic ecosystems. Cyanobacteria have well-documented allelopathic properties, mainly derived from the evaluation of the activity of allelopathic extracts or pure compounds towards monocultures of selected target microorganisms. Consequently, little is known regarding the community dynamics of microorganisms associated with allelopathic interactions. In this laboratory-based study, a Microcystis spp.-dominated microbial community from a freshwater lake was exposed, for 15 days, to exudates from the cyanobacterium Oscillatoria sp. strain LEGE 05292 in laboratory conditions. This cyanobacterium is known to produce the allelochemicals portoamides, which were among the exuded compounds. The community composition was followed (by means of polymerase chain reaction followed by denaturing gradient gel electrophoresis and microscopic analyses) and compared to that of a non-exposed situation. Following exposure, clear differences in the community structure were observed, in particular for cyanobacteria and unicellular eukaryotic taxa. Interestingly, distinct Microcystis genotypes present in the community were differentially impacted by the exposure, highlighting the fine-scale dynamics elicited by the exudates. These results support a role for cyanobacterial allelochemicals in the structuring of aquatic microbial communities.     

Adaptation of aquatic microbial communities to hg stress

The mechanism of adaptation to Hg in four aquatic habitats was studied by correlating microbially mediated Hg volatilization with the adaptive state of the exposed communities. Community diversity, heterotrophic activity, and Hg resistance measurements indicated that adaptation of all four communities was stimulated by preexposure to Hg. In saline water communities, adaptation was associated with rapid volatilization after an initial lag period. This mechanism, however, did not promote adaptation in a freshwater sample, in which Hg was volatilized slowly, regardless of the resistance level of the microbial community. Distribution of the mer operon among representative colonies of the communities was not related to adaptation to Hg. Thus, although volatilization enabled some microbial communities to sustain their functions in Hg-stressed environments, it was not mediated by the genes that serve as a model system in molecular studies of bacterial resistance to mercurials.     

Fourier transform infrared spectroscopy as a metabolite fingerprinting tool for monitoring the phenotypic changes in complex bacterial communities capable of degrading phenol

The coking process produces great volumes of wastewater contaminated with pollutants such as cyanides, sulfides and phenolics. Chemical and physical remediation of this wastewater removes the majority of these pollutants; however, these processes do not remove phenol and thiocyanate. The removal of these compounds has been effected during bioremediation with activated sludge containing a complex microbial community. In this investigation we acquired activated sludge from an industrial bioreactor capable of degrading phenol. The sludge was incubated in our laboratory and monitored for its ability to degrade phenol over a 48 h period. Multiple samples were taken across the time‐course and analysed by Fourier transform infrared (FT‐IR) spectroscopy. FT‐IR was used as a whole‐organism fingerprinting approach to monitor biochemical changes in the bacterial cells during the degradation of phenol. We also investigated the ability of the activated sludge to degrade phenol following extended periods (2–131 days) of storage in the absence of phenol. A reduction was observed in the ability of the microbial community to degrade phenol and this was accompanied by a detectable biochemical change in the FT‐IR fingerprint related to cellular phenotype of the microbial community. In the absence of phenol a decrease in thiocyanate vibrations was observed, reflecting the ability of these communities to degrade this substrate. Actively degrading communities showed an additional new band in their FT‐IR spectra that could be attributed to phenol degradation products from the ortho‐ and meta‐cleavage of the aromatic ring. This study demonstrates that FT‐IR spectroscopy when combined with chemometric analysis is a very powerful high throughput screening approach for assessing the metabolic capability of complex microbial communities.     

南极冰下水生态系统微生物与生源元素循环研究进展

南极大陆冰盖下存在液态水,形成了由冰下湖、冰下河/溪、冰封湖和冰架下水体等组成的冰下水生态系统,具有低温、黑暗和寡营养等极端的环境条件特征。微生物主导了南极冰下水生态系统,其具有丰富多样的种群构成、功能形式和独特的适应机制,在生源元素生物地球化学循环过程中起了重要作用。研究南极冰下微生物群落的生态特征及其参与的生源元素地球化学循环过程,可为揭示地球生命演化和探索外星生命提供指示,具有重要的科学意义。本文综述了南极冰下水生态系统的极端环境条件、冰下微生物的多样性、冰下微生物参与的生物地球化学循环以及冰下微生物的适极机理,最后基于研究现状展望了南极冰下微生物的未来研究方向。     

Biomonitoring of continuous microbial community adaptation towards more efficient phenol-degradation in a fed-batch bioreactor

The anaerobic degradation of phenol was studied in a fed-batch culture. Nitrate was added as electron acceptor and phenol was provided three times, to a final concentration of 200 mg/l. Randomly amplified polymorphic DNA (RAPD) and terminal fraction fragment length polymorphism (T-RFLP) were used and compared in order to monitor the microbial succession in the reactor. Phenol degradation started after an initial lag phase of 14 days and was then completed within a few days. In addition, the duration of the lag phase was shortened and the degradation rate was increased after each phenol amendment. Nitrate reduction correlated with microbial growth and phenol depletion, confirming that the degradation was carried out anaerobically. Results from the DNA analysis showed that the structure of the microbial community changed after each phenol amendment. This study confirms the potential for anaerobic degradation of environmental pollutants and also confirms that microbial acclimation towards faster degradation rates occurred upon repeated substrate amendments. Furthermore, both of the DNA-based techniques described the phenol degradation-linked community shifts with similar general results. RAPD is a faster, simpler technique that gives a higher resolution and consequently reflects the shifts in the microbial community structure better, whereas T-RFLP is more suitable for phylogenetic studies.     

Change in bacterial community during biodegradation of aniline

The response of river water microbial communities to chemical compounds was monitored under laboratory conditions using aniline as a model. Bacteria were collected from unpolluted and polluted sites. Bacterial abundance (plate and total direct counting) and its relation to aniline biodegradation was examined. Colony hybridization with 16S rRNA oligonucleotide probes was used to study the changes in microbial community structure during biodegradation of aniline. The changes in bacterial abundance and community structure were related to biodegradation of aniline. Burkholderia–Pseudomonas (rRNA group III), an authentic Alcaligenes group became dominant despite the initial differences in the microbial communities, suggesting that these genera are the main aniline degraders in the aquatic environment.     

Effects of Picoxystrobin and 4-n-Nonylphenol on Soil Microbial Community Structure and Respiration Activity

There is widespread use of chemical amendments to meet the demands for increased productivity in agriculture. Potentially toxic compounds, single or in mixtures, are added to the soil medium on a regular basis, while the ecotoxicological risk assessment procedures mainly follow a chemical by chemical approach. Picoxystrobin is a fungicide that has caused concern due to studies showing potentially detrimental effects to soil fauna (earthworms), while negative effects on soil microbial activities (nitrification, respiration) are shown to be transient. Potential mixture situations with nonylphenol, a chemical frequently occurring as a contaminant in sewage sludge used for land application, infer a need to explore whether these chemicals in mixture could alter the potential effects of picoxystrobin on the soil microflora. The main objective of this study was to assess the effects of picoxystrobin and nonylphenol, as single chemicals and mixtures, on soil microbial community structure and respiration activity in an agricultural sandy loam. Effects of the chemicals were assessed through measurements of soil microbial respiration activity and soil bacterial and fungal community structure fingerprints, together with a degradation study of the chemicals, through a 70 d incubation period. Picoxystrobin caused a decrease in the respiration activity, while 4-n-nonylphenol caused an increase in respiration activity concurring with a rapid degradation of the substance. Community structure fingerprints were also affected, but these results could not be directly interpreted in terms of positive or negative effects, and were indicated to be transient. Treatment with the chemicals in mixture caused less evident changes and indicated antagonistic effects between the chemicals in soil. In conclusion, the results imply that the application of the fungicide picoxystrobin and nonylphenol from sewage sludge application to agricultural soil in environmentally relevant concentrations, as single chemicals or in mixture, will not cause irreversible effects on soil microbial respiration and community structure.     

采煤沉陷区不同造林树种恢复土壤酚酸物质对土壤微生物的影响

酚酸物质是影响微生物群落和结构的最重要因子之一,研究酚酸物质在不同造林树种土壤中的变化规律及其与微生物群落结构的关系,有助于更好地了解和揭示采煤沉陷区恢复造林条件下微生物群落变化的机制.本研究在双鸭山宝山采煤沉陷区的撂荒地基础上营造三针一阔(红松、落叶松、樟子松和杨树)人工林,测定这4种造林地土壤酚类物质、11种酚酸物质和微生物群落结构.结果表明: 复合态酚含量总体表现为人工林显著高于撂荒地,其中,落叶松人工林和杨树人工林的复合态酚含量较高,落叶松人工林和红松人工林的总酚含量显著高于撂荒地,红松人工林的水溶性酚含量显著高于撂荒地;在11种酚酸物质中,阿魏酸、松香酸、β-谷甾醇、齐墩果酸、莽草酸、亚油酸和硬脂酸的含量在人工林土壤中较高.土壤酚类物质与土壤微生物生物量不存在显著的相关关系,个别的酚酸物质与土壤微生物的相关关系显著,其中,阿魏酸、松香酸和β-谷甾醇对土壤微生物生物量有明显的促进作用,与真菌和真菌/细菌存在显著的正相关关系.杨树人工林的酚酸物质含量较高,说明营造杨树人工林对采煤沉陷区的土壤恢复有益.     

Increasing concentrations of phenol progressively affect anaerobic digestion of cellulose and associated microbial communities

Performance stability is a key issue when managing anaerobic digesters. However it can be affected by external disturbances caused by micropollutants. In this study the influence of phenol on the methanization of cellulose was evaluated through batch toxicity assays. Special attention was given to the dynamics of microbial communities by means of automated ribosomal intergenic spacer analysis. We observed that, as phenol concentrations increased, the different steps of anaerobic cellulose digestion were unevenly and progressively affected, methanogenesis being the most sensitive: specific methanogenic activity was half-inhibited at 1.40 g/L of phenol, whereas hydrolysis of cellulose and its fermentation to VFA were observed at up to 2.00 g/L. Depending on the level of phenol, microbial communities resisted either through physiological or structural adaptation. Thus, performances at 0.50 g/L were maintained in spite of the microbial community’s shift. However, the communities’ ability to adapt was limited and performances decreased drastically beyond 2.00 g/L of phenol.     

微生物燃料电池降解苯酚过程中微电场对阴极微生物多样性的影响

[背景]苯酚废水作为一种毒性强、难降解的废水而备受关注.目前,微生物燃料电池(microbial fuel cell,MFC)已经广泛用于苯酚废水的降解,MFC的产电效果和苯酚的降解效率与反应器内的微生物群落有着密切关系.[目的]为了提高MFC的产电效果及对有害物质的降解能力,需要对MFC中苯酚的降解和微生物群落结构进...     

Biodegradation of tert-butylphenyl diphenyl phosphate

The biodegradation of tert-butylphenyl diphenyl phosphate (BPDP) was examined in microcosms containing sediment and water from five different ecosystems as part of our studies to elucidate the environmental fate of phosphate ester flame retardants. Biodegradation of [14C]BPDP was monitored in the environmental microcosms by measuring the evolution of 14CO2. Over 37% of BPDP was mineralized after 8 weeks in microcosms from an ecosystem which had chronic exposure to agricultural chemicals. In contrast, only 1.7% of BPDP was degraded to 14CO2 in samples collected from a noncontaminated site. The exposure concentration of BPDP affected the percentage which was degraded to 14CO2 in microcosms from the two most active ecosystems. Mineralization was highest at a concentration of 0.1 mg of BPDP and was inhibited with 10- and 100-fold higher concentrations of BPDP in these microcosms. Indigenous heterotrophic and BPDP-utilizing microbial populations and phosphoesterase enzyme activities were highest in sediments which had the highest biodegradation of BPDP. We observed adaptive increases in both microbial populations and phosphoesterase enzymes in some sediments acclimated to BPDP. Chemical analyses of the residues in the microcosms indicated undegraded BPDP and minor amounts of phenol, tert-butylphenol, diphenyl phosphate, and triphenyl phosphate as biodegradation products. These data suggest that the microbial degradation of BPDP results from at least three catabolic processes and is highest when low concentrations of BPDP are exposed to sediment microorganisms of eutrophic ecosystems which have high phosphotri- and diesterase activities and previous exposure to anthropogenic chemicals.     

Biodegradation of tert-butylphenyl diphenyl phosphate.

The biodegradation of tert-butylphenyl diphenyl phosphate (BPDP) was examined in microcosms containing sediment and water from five different ecosystems as part of our studies to elucidate the environmental fate of phosphate ester flame retardants. Biodegradation of [14C]BPDP was monitored in the environmental microcosms by measuring the evolution of 14CO2. Over 37% of BPDP was mineralized after 8 weeks in microcosms from an ecosystem which had chronic exposure to agricultural chemicals. In contrast, only 1.7% of BPDP was degraded to 14CO2 in samples collected from a noncontaminated site. The exposure concentration of BPDP affected the percentage which was degraded to 14CO2 in microcosms from the two most active ecosystems. Mineralization was highest at a concentration of 0.1 mg of BPDP and was inhibited with 10- and 100-fold higher concentrations of BPDP in these microcosms. Indigenous heterotrophic and BPDP-utilizing microbial populations and phosphoesterase enzyme activities were highest in sediments which had the highest biodegradation of BPDP. We observed adaptive increases in both microbial populations and phosphoesterase enzymes in some sediments acclimated to BPDP. Chemical analyses of the residues in the microcosms indicated undegraded BPDP and minor amounts of phenol, tert-butylphenol, diphenyl phosphate, and triphenyl phosphate as biodegradation products. These data suggest that the microbial degradation of BPDP results from at least three catabolic processes and is highest when low concentrations of BPDP are exposed to sediment microorganisms of eutrophic ecosystems which have high phosphotri- and diesterase activities and previous exposure to anthropogenic chemicals.     

Adaptation of Aquatic Microbial Communities to Hg2+ Stress

The mechanism of adaptation to Hg²⁺ in four aquatic habitats was studied by correlating microbially mediated Hg²⁺ volatilization with the adaptive state of the exposed communities. Community diversity, heterotrophic activity, and Hg²⁺ resistance measurements indicated that adaptation of all four communities was stimulated by preexposure to Hg²⁺. In saline water communities, adaptation was associated with rapid volatilization after an initial lag period. This mechanism, however, did not promote adaptation in a freshwater sample, in which Hg²⁺ was volatilized slowly, regardless of the resistance level of the microbial community. Distribution of the mer operon among representative colonies of the communities was not related to adaptation to Hg²⁺. Thus, although volatilization enabled some microbial communities to sustain their functions in Hg²⁺-stressed environments, it was not mediated by the genes that serve as a model system in molecular studies of bacterial resistance to mercurials.     

Use of Field-Based Stable Isotope Probing To Identify Adapted Populations and Track Carbon Flow through a Phenol-Degrading Soil Microbial Community

The goal of this field study was to provide insight into three distinct populations of microorganisms involved in in situ metabolism of phenol. Our approach measured ¹³CO₂ respired from [¹³C]phenol and stable isotope probing (SIP) of soil DNA at an agricultural field site. Traditionally, SIP-based investigations have been subject to the uncertainties posed by carbon cross-feeding. By altering our field-based, substrate-dosing methodologies, experiments were designed to look beyond primary degraders to detect trophically related populations in the food chain. Using gas chromatography-mass spectrometry (GC/MS), it was shown that ¹³C-labeled biomass, derived from primary phenol degraders in soil, was a suitable growth substrate for other members of the soil microbial community. Next, three dosing regimes were designed to examine active members of the microbial community involved in phenol metabolism in situ: (i) 1 dose of [¹³C]phenol, (ii) 11 daily doses of unlabeled phenol followed by 1 dose of [¹³C]phenol, and (iii) 12 daily doses of [¹³C]phenol. GC/MS analysis demonstrated that prior exposure to phenol boosted ¹³CO₂ evolution by a factor of 10. Furthermore, imaging of ¹³C-treated soil using secondary ion mass spectrometry (SIMS) verified that individual bacteria incorporated ¹³C into their biomass. PCR amplification and 16S rRNA gene sequencing of ¹³C-labeled soil DNA from the 3 dosing regimes revealed three distinct clone libraries: (i) unenriched, primary phenol degraders were most diverse, consisting of α-, β-, and γ-proteobacteria and high-G+C-content gram-positive bacteria, (ii) enriched primary phenol degraders were dominated by members of the genera Kocuria and Staphylococcus, and (iii) trophically related (carbon cross-feeders) were dominated by members of the genus Pseudomonas. These data show that SIP has the potential to document population shifts caused by substrate preexposure and to follow the flow of carbon through terrestrial microbial food chains.