Accelerated biodegradation of petroleum hydrocarbon waste

Conventional landfarming approaches to bioremediation of refinery and other petroleum sludges are not acceptable environmentally and are banned in most North American jurisdictions. While initial bioreactor-based systems for treatment of these sludges required batch-cycle process-times of 1–3 months, an accelerated process has now been developed which can be completed in 10–12 days. In this process, up to 99% of total petroleum hydrocarbons are degraded and the sludges are converted from hazardous to non-hazardous according to the United States EPA's toxicity characteristic leachate procedure criteria. Understanding and exploiting mechanisms to improve hydrocarbon accession to the degrading microorganisms was a key development component of the process. Contrasting physiological mechanisms were observed for different component organisms of the mixed culture with respect to their associations with the hydrocarbon substrate; and the beneficial effects of using surfactants were demonstrated. The mixed culture used in the process exhibited a capacity for high-rate degradation of volatile organic carbons and the potential use of the culture as a liquid biofilter was demonstrated. The culture was also effective as an inoculant for the bioaugmentation of total petroleum hydrocarbon-contaminated soil and as a de-emulsifier of oilfield emulsions and could transform some other environmental contaminants which are not predominant components of crude oil.     

Accelerated biodegradation of cured cement paste by Thiobacillus species under simulation condition

Biodegradation is one of the most important types of cement deterioration. Complex microbial populations take part in the biodegradation process of cement-based materials. Studies in this field show that the sulfur-oxidizing bacteria, including Acidithiobacillus thiooxidans, due to sulfuric acid formation, play a key role in this process. In this study, with the accelerated leaching process of calcium hydroxide of cement paste, cured under running tap water and exposed to sterile biogenic sulfuric acid for 6 days, the surface pH of the cement was reduced to a more favorable level for bacterial growth. In this case, the growth of Thiobacillus proceeded in the presence of cured cement paste specimens. After 90 days of exposure to a semi-continuous culture of A. thiooxidans with its pH less than 2 and continuous removal of damaged layers the compressive strength, length and mass of the samples dropped by 96%, 11% and 43%, in the order given. The mechanism of degradation and the structure of degraded specimens were analyzed by test laboratory techniques such as, XRD, SEM and EDAX analyses.     

Accelerated evolutionary rate in sulfur-oxidizing endosymbiotic bacteria associated with the mode of symbiont transmission

The nearly neutral theory of molecular evolution predicts that the rate of nucleotide substitution should accelerate in small populations at sites under low selective constraint. We examined these predictions with respect to the relative population sizes for three bacterial life histories within chemolithoautotrophic sulfur-oxidizing bacteria: (1) free-living bacteria, (2) environmentally captured symbionts, and (3) maternally transmitted symbionts. Both relative rates of nucleotide substitution and relative ratios of loop, stem, and domain substitutions from 1,165 nt of the small-subunit 16S rDNA were consistent with expectations of the nearly neutral theory. Relative to free-living sulfur-oxidizing autotrophic bacteria, the maternally transmitted symbionts have faster substitution rates overall and also in low-constraint domains of 16S rDNA. Nucleotide substitition rates also differ between loop and stem positions. All of these findings are consistent with the predictions that these symbionts have relatively small effective population sizes. In contrast, the rates of nucleotide substitution in environmentally captured symbionts are slower, particularly in high-constraint domains, than in free-living bacteria.     

Biosorption of lead phosphates by lead-tolerant bacteria as a mechanism for lead immobilization

The study of metal-tolerant bacteria is important for bioremediation of contaminated environments and development of green technologies for material synthesis due to their potential to transform toxic metal ions into less toxic compounds by mechanisms such as reduction, oxidation and/or sequestration. In this study, we report the isolation of seven lead-tolerant bacteria from a metal-contaminated site at Zacatecas, México. The bacteria were identified as members of the Staphylococcus and Bacillus genera by microscopic, biochemical and 16S rDNA analyses. Minimal inhibitory concentration of these isolates was established between 4.5 and 7.0 mM of Pb(NO₃)₂ in solid and 1.0–4.0 mM of Pb(NO₃)₂ in liquid media. A quantitative analysis of the lead associated to bacterial biomass in growing cultures, revealed that the percentage of lead associated to biomass was between 1 and 37% in the PbT isolates. A mechanism of complexation/biosorption of lead ions as inorganic phosphates (lead hydroxyapatite and pyromorphite) in bacterial biomass, was determined by Fourier transform infrared spectroscopy and X-ray diffraction analyses. Thus, the ability of the lead-tolerant isolates to transform lead ions into stable and highly insoluble lead minerals make them potentially useful for immobilization of lead in mining waste.     

Microbial activity of trench leachates from shallow-land, low-level radioactive waste disposal sites.

Trench leachate samples collected anoxically from shallow-land, low-level radioactive waste disposal sites were analyzed for total aerobic and anaerobic populations, sulfate reducers, denitrifiers, and methanogens. Among the several aerobic and anaerobic bacteria isolated, only Bacillus sp., Pseudomonas sp., Citrobacter sp., and Clostridium sp. were identified. Mixed bacterial cultures isolated from the trench leachates were able to grow anaerobically in trench leachates, which indicates that the radionuclides and organic chemicals present were not toxic to these bacteria. Changes in concentrations of several of the organic constituents of the waste leachate samples were observed due to anaerobic microbial activity. Growth of a mixed culture of trench-water bacteria in media containing a mixture of radionuclides, 60Co, 85Sr, and 134,137Cs, was not affected at total activity concentrations of 2.6 X 10(2) and 2.7 X 10(3) pCi/ml.     

餐厨垃圾高温好氧生物减量菌种的筛选及特性

【背景】随着餐厨垃圾产生量的逐步提高,如何实现其快速降解,成为餐厨垃圾处理亟待解决的问题。餐厨垃圾的高温好氧生物减量技术是一种可以快速降解餐厨垃圾的有效方法。【目的】筛选能够适应餐厨垃圾环境且具有高效降解餐厨垃圾中有机物能力的菌株,以提高餐厨垃圾的降解效率和减量效果。【方法】采用温度梯度耐受性实验和餐厨垃圾浸出液高油高盐耐受性实验进行菌种初筛,并利用产酶培养基复筛及餐厨垃圾生物减量实验验证。【结果】通过初筛、复筛和功能验证,最终获得4株生物减量效果优良的菌株N3-1、C7、N3-3和G6-1,其对餐厨垃圾挥发性固体(volatile solid,VS)的降解率分别为36.95%、33.23%、32.83%和31.91%,是对照组的3.02、2.71、2.68和2.61倍。经鉴定,这4株菌分别属于热嗜油地芽孢杆菌(Geobacillus thermoleovorans)、史氏芽孢杆菌(Bacillus smithii)、热解木糖地芽孢杆菌(Geobacillus caldoxylosilyticus)和立陶宛地芽孢杆菌(Geobacillus lituanicus)。【结论】筛选出的4株菌均具有较强的餐厨垃圾原料适应性和高效的生物降解能力,为开发餐厨垃圾高温好氧复合菌剂奠定了基础,并为实现餐厨垃圾减量化、无害化处理和资源化利用提供了技术支持。     

嗜酸硫氧化细菌消解元素硫的研究进展

浸矿酸性环境下,金属硫化矿在Fe3+作用下,经过硫代硫酸盐途径或多聚硫化氢途径而分解的过程中导致大量元素硫的累积,进而可能在金属硫化矿表面形成疏水元素硫层,阻碍金属离子的进一步浸出。酸性环境下,惰性元素硫的消解必须借助嗜酸硫氧化细菌来实现。该消解过程包括嗜酸硫氧化细菌对元素硫的吸附、转运以及氧化转化等过程。本文对近年来嗜酸硫氧化细菌消解元素硫过程的相关研究进行了全面评述,认为有关嗜酸硫氧化细菌消解元素硫的分子机制的清晰阐述还有待人们通过对消解过程的各个环节的分子机制进行大量研究来实现。     

Aerobic biodegradation of nonylphenol by cold-adapted bacteria

Three strains capable of mineralizing nonylphenol as sole carbon source were isolated from a sample of contaminated soil and characterized as two Pseudomonas spp. and a Stenotrophomonas sp. The two Pseudomonas spp. expressed characteristics typical of psychrophiles growing optimally of 10 °C and capable of growing at 0 °C. The Stenotrophomonas sp. was more likely psychrotrophic because it had an optimal temperature between 14 and 22 °C although it was not capable of growing at 4 °C. At 14 °C, one of the Pseudomonas spp. exhibited the highest rate of degradation of nonylphenol (4.4 mg l^–1 d^–1), when compared with axenic or mixed cultures of the isolates. This study represents, to the best of our knowledge, the first reported case of cold-adapted microorganisms capable of mineralizing nonylphenol.     

The biodegradation of aromatic hydrocarbons by bacteria

Aromatic compounds of both natural and man-made sources abound in the environment. The degradation of such chemicals is mainly accomplished by microorganisms. This review provides key background information but centres on recent developments in the bacterial degradation of selected man-made aromatic compounds. An aromatic compound can only be considered to be biodegraded if the ring undergoes cleavage, and this is taken as the major criteria for inclusion in this review (although the exact nature of the enzymic ring-cleavage has not been confirmed in all cases discussed).The biodegradation of benzene, certain arenes, biphenyl and selected fused aromatic hydrocarbons, by single bacterial isolates, are dealt with in detail.     

The influence of microbial redox cycling on radionuclide mobility in the subsurface at a low-level radioactive waste storage site

As anaerobic microbial metabolism can have a major impact on radionuclide speciation and mobility in the subsurface, the solubility of uranium, technetium and radium was determined in microcosms prepared from sediments adjacent to the Drigg low-level radioactive waste storage site (UK). Both uranium (as U(VI);     ) and Tc (as Tc(VII);     ) were removed from groundwater concurrently with microbial Fe(III) reduction, presumably through reduction to insoluble U(IV) and Tc(IV), respectively, while Ra (Ra²⁺) that had rapidly sorbed onto mineral surfaces was not released following Fe(III) reduction. Biogenic Fe(II) minerals in reduced Drigg sediments were unable to reduce U(VI) abiotically but could reduce Tc(VII). Following addition of the oxidant nitrate to the reduced sediments, uranium was remobilized and released into solution, whereas technetium remained associated with an insoluble phase. A close relative of Pseudomonas stutzeri dominated the microbial communities under denitrifying conditions, reducing nitrate to nitrite in the microcosms, which was able to reoxidize Fe(II) and U(IV), with release of the latter into solution as U(VI). These data suggest that microbial Fe(III) reduction in the far-field at Drigg has the potential to decrease the migration of some radionuclides in the subsurface, and the potential for reoxidation and remobilization by nitrate, a common contaminant in nuclear waste streams, is radionuclide-specific.     

Sequential application of waste whey as a medium component for Kluyveromyces lactis cultivation and a co-feeder for lipase immobilization by CLEA method

Currently, much attention is paid to technologies which can be drivers of the circular economy across different sectors, in particular, to develop technologies for utilization or reusability of biocompatible materials from industrial waste. One of such is the milk whey, which is a cheap biobased raw material, the disposal of which is a major problem for the dairy industry. Our proposed and investigated technology is based on a continuous exploitation of the whey combining microbiology and biotechnology. Primarily, whey was used as a nutrition source for the cultivation of Kluyveromyces lactis with the aim to produce the targeted biocatalyst—lipase. During cultivation, the whey was transformed into the hydrolyzed form, which was further successfully applied as a protein feeder (external linker) for immobilization of lipase by cross-linked enzyme aggregate (CLEA) method. The first time use of whey as a co-feeder for immobilization of enzymes by CLEA method has shown promising results and increased the stability of lipases for temperature and organic solvents. Hydrolysis of rapeseed oil catalyzed with immobilized derivatives was obtained with 45–96% efficiency at non-optimized conditions. Additionally, the determined kinetic parameters indicated that the rate of p-nitrophenyl palmitate hydrolysis was not changed drastically after immobilization.

     

Kinetics of biodegradation of p-nitrophenol by different bacteria

Three bacterial species, i.e., Ralstonia sp. SJ98, Arthrobacter protophormiae RKJ100, and Burkholderia cepacia RKJ200, have been examined for their efficiency and kinetics behavior toward PNP degradation. All the three bacteria utilized PNP as the sole source of carbon, nitrogen, and energy. The rates of radiolabeled [U-(14)C]PNP degradation by all the bacteria were higher in the nitrogen-free medium compared to the medium with nitrogen. The apparent K(m) values of PNP degradation by SJ98, RKJ100, and RKJ200 were 0.32, 0.28, and 0.23 mM, respectively, as determined from the Michaelis-Menten curves. The maximum rates of PNP degradation (V(max)) according to Lineweaver-Burk's plots were 11.76, 7.81, and 3.84 micromol PNP degraded/min/mg dry biomass, respectively. The interpretation drawn from the Lineweaver-Burk's plots showed that the PNP degradation by SJ98 was stimulated by 4-nitrocatechol and 1, 2,4-benzenetriol. Benzoquinone and hydroquinone inhibited PNP degradation by RKJ100 noncompetitively and competitively, respectively, whereas in the case of RKJ200, benzoquinone and hydroquinone inhibited PNP degradation in an uncompetitive manner. beta-Ketoadipate did not affect the rate of PNP degradation in any case.     

Aerobic biodegradation of propylene glycol by soil bacteria

Propylene glycol (PG) is a main component of aircraft deicing fluids and its extensive use in Northern airports is a source of soil and groundwater contamination. Bacterial consortia able to grow on PG as sole carbon and energy source were selected from soil samples taken along the runways of Oslo Airport Gardermoen site (Norway). DGGE analysis of enrichment cultures showed that PG-degrading populations were mainly composed by Pseudomonas species, although Bacteroidetes were found, as well. Nineteen bacterial strains, able to grow on PG as sole carbon and energy source, were isolated and identified as different Pseudomonas species. Maximum specific growth rate of mixed cultures in the absence of nutrient limitation was 0.014 h^?1 at 4 °C. Substrate C:N:P molar ratios calculated on the basis of measured growth yields are in good agreement with the suggested values for biostimulation reported in literature. Therefore, the addition of nutrients is suggested as a suitable technique to sustain PG aerobic degradation at the maximum rate by autochthonous microorganisms of unsaturated soil profile.     

Accelerated biodegradation of atrazine by a microbial consortium is possible in culture and soil

A mixed enrichment culture of microorganisms capable of accelerated mineralization of atrazine was isolated from soil treated with successive applications of the herbicide. Liquid cultures of this consortium, in the presence of simple carbon sources, mineralized 96% of the applied atrazine (0.56 mM) within 7 days. Atrazine mineralization in culture is initiated with the formation of the metabolite hydroxyatrazine. In soil treated with atrazine at a concentration of 0.14 mM (concentration is based on total soil mass), and then inoculated with the microbial consortium, the parent compound was completely transformed in 25 days. After 30 days of incubation, 60% of the applied atrazine was accounted for as¹⁴CO₂. As was found with the liquid cultures, hydroxyatrazine was the major metabolite. After 145 days, soil extractable hydroxyatrazine declined to zero and 86% of the applied atrazine was accounted for as¹⁴CO₂. No metabolites, other than hydroxyatrazine, were recovered from either the liquid culture or soil inoculated with the consortium. The use of the mixed microbial culture enhanced mineralization more than 20 fold as compared to uninoculated soil.     

Kinetics of anaerobic biodegradation of glycerol by sulfate-reducing bacteria

A kinetic model has been developed and kinetic parameters of anaerobic degradation of glycerol, an abundant by-product of biofuel manufacturing, by a consortium of sulfate reducing bacteria (SRB) in a closed system have been determined. The following main species of SRB has been identified in the consortium: Desulfovibrio baarsii, Desulfomicrobium sp., and Desufatomaculum sp. The proposed model included processes of glycerol degradation, sulfate reduction, and inhibition by metabolic products, as well as effects of pH and temperature. The suggested equation for the anaerobic glycerol degradation was based on Edward and Andrew’s equation. The following kinetic parameters of the anaerobic glycerol degradation were obtained for the initial glycerol concentration from 0.15 to 4 ml/l and sulfate concentration of 2760 mg/l at 22°C: maximum specific growth rate of SRB μ_max = 0.56 day⁻¹, economic coefficient of ashless biomass from glycerol of 0.08 mol SRB/mol COC, and yield of ashless biomass from sulfate of 0.020 mol SRB/mol SO₄. It was shown that the optimum molar ratio of $ {{C_{Gl} } \mathord{\left/ {\vphantom {{C_{Gl} } {C_{SO_4 } }}} \right. \kern-\nulldelimiterspace} {C_{SO_4 } }} $ {{C_{Gl} } \mathord{\left/ {\vphantom {{C_{Gl} } {C_{SO_4 } }}} \right. \kern-\nulldelimiterspace} {C_{SO_4 } }} for SRB growth was 0.8. Initial boundary concentration of inhibition by undissociated hydrogen sulfide was 70 mg/l. Dependence of the specific growth rate of bacteria on the temperature was approximated by the Arrhenius equation in the temperature range of 20–30°C with the goodness of fit R² = 0.99.     

Enhanced-rate biodegradation of organophosphate neurotoxins by immobilized nongrowing bacteria

Pesticide wastes generated from livestock dipping operations containing the organophosphate (OP) insecticide coumaphos (CP) are well suited for disposal by biodegradation since they are highly concentrated (approximately 1 g/L), generally contained, and lack additional toxic components. In this study, a significantly enhanced efficiency of degrading CP in cattle dip waste (CDW) is reported using a dense, nongrowing cell population that functions without the addition of nutrients required for growing cell cultures. A recombinant strain of Escherichia coli containing the opd gene for organophosphate hydrolase (OPH), which is capable of active hydrolysis of OP neurotoxins including CP, was cultivated in a rich medium containing all essential nutrients. Cells were harvested and utilized in lab scale experiments in the form of either freely suspended cells or cells immobilized within a macroporous gel matrix, poly(vinyl alcohol) (PVA) cryogel. Significantly higher degradation rates were achieved with either suspended or immobilized OPH(+) cells compared to rates with the microbial consortium naturally present in CDW. Of the two nongrowing cell systems, the detoxification rate with immobilized cells was approximately twice that of freely suspended cells, and kinetic studies demonstrated that a higher maximum reaction rate was achieved with the immobilized cell system. A comparative study using both the CDW and pure CP substrates with free cells indicated that the CDW contained one or more factors that reduced the bioavailability of CP. The immobilized cells retained their activity over a 4-month period of use and storage, demonstrating both sustained catalytic activity and long-term mechanical stability.     

Bacterial bioluminescence as a bioassay for mycotoxins.

The use of bacterial bioluminescence as a toxicological assay for mycotoxins was tested with rubratoxin B, zearalenone, penicillic acid, citrinin, ochratoxin A, PR-toxin, aflatoxin B1, and patulin. The concentrations of mycotoxins causing 50% light reduction (EC50) in Photobacterium phosphoreum were determined immediately and at 5 h after reconstitution of the bacteria from a freeze-dried state. Generally, less toxins were required to obtain an EC50 at 5 h. The effects of the above mycotoxins on bioluminescence were determined after 5, 10, 15, and 20 min of incubation with the bacterial suspensions. The concentration of rubratoxin B necessary to elicit an EC50 increased with time, whereas the concentration of citrinin, penicillic acid, patulin, and PR-toxin necessary decreased with time. There was very little change in the concentration of zearalenone, aflatoxin B1, and ochratoxin A required to elicit an EC50 with time. The bacterial bioluminescence assay was most sensitive to patulin and least sensitive to rubratoxin B.     

Reverse dissimilatory sulfite reductase as phylogenetic marker for a subgroup of sulfur-oxidizing prokaryotes

Sulfur-oxidizing prokaryotes (SOP) catalyse a central step in the global S-cycle and are of major functional importance for a variety of natural and engineered systems, but our knowledge on their actual diversity and environmental distribution patterns is still rather limited. In this study we developed a specific PCR assay for the detection of dsrAB that encode the reversely operating sirohaem dissimilatory sulfite reductase (rDSR) and are present in many but not all published genomes of SOP. The PCR assay was used to screen 42 strains of SOP (most without published genome sequence) representing the recognized diversity of this guild. For 13 of these strains dsrAB was detected and the respective PCR product was sequenced. Interestingly, most dsrAB -encoding SOP are capable of forming sulfur storage compounds. Phylogenetic analysis demonstrated largely congruent rDSR and 16S rRNA consensus tree topologies, indicating that lateral transfer events did not play an important role in the evolutionary history of known rDSR. Thus, this enzyme represents a suitable phylogenetic marker for diversity analyses of sulfur storage compound-exploiting SOP in the environment. The potential of this new functional gene approach was demonstrated by comparative sequence analyses of all dsrAB present in published metagenomes and by applying it for a SOP census in selected marine worms and an alkaline lake sediment.     

Thermophilic biodegradation of BTEX by two consortia of anaerobic bacteria

Two thermophilic anaerobic bacterial consortia (ALK-1 and LLNL-1), capable of degrading the aromatic fuel hydrocarbons, benzene, toluene, ethylbenzene, and the xylenes (BTEX compounds), were developed at 60 °C from the produced water of ARCO'S Kuparuk oil field at Alaska and the subsurface water at the Lawrence Livermore National Laboratory gasoline-spill site, respectively. Both consortia were found to grow at 45–75 °C on BTEX compounds as their sole carbon and energy sources with 50 °C being the optimal temperature. With 3.5 mg total BTEX added to sealed 50-ml serum bottles, which contained 30 ml mineral salts medium and the consortium, benzene, toluene, ethylbenze, m-xylene, and an unresolved mixture of o- and p-xylenes were biodegraded by 22%, 38%, 42%, 40%, and 38%, respectively, by ALK-1 after 14 days of incubation at 50 °C. Somewhat lower, but significant, percentages of the BTEX compounds also were biodegraded at 60 °C and 70 °C. The extent of biodegradation of these BTEX compounds by LLNL-1 at each of these three temperatures was slightly less than that achieved by ALK-1. Use of [ring-¹⁴C]toluene in the BTEX mixture incubated at 50 °C verified that 41% and 31% of the biodegraded toluene was metabolized within 14 days to water-soluble products by ALK-1 and LLNL-1, respectively. A small fraction of it was mineralized to ¹⁴CO₂. The use of [U-¹⁴C]benzene revealed that 2.6%–4.3% of the biodegraded benzene was metabolized at 50 °C to water-soluble products by the two consortia; however, no mineralization of the degraded [U-¹⁴C]benzene to ¹⁴CO₂ was observed. The biodegradation of BTEX at all three temperatures by both consortia was tightly coupled to sulfate reduction as well as H₂S generation. None was observed when sulfate was omitted from the serum bottles. This suggests that sulfate-reducing bacteria are most likely responsible for the observed thermophilic biodegradation of BTEX in both consortial cultures. Received: 12 July 1996 / Received revision: 31 December 1996 / Accepted: 31 January 1997