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1.
A quantitative solid-phase microextraction, gas chromatography, flame ionization detector (SPME-GC-FID) method for low-molecular-weight
hydrocarbons from crude oil was developed and applied to live biodegradation samples. Repeated sampling was achieved through
headspace extractions at 30°C for 45 min from flasks sealed with Teflon Mininert. Quantification without detailed knowledge
of oil–water–air partition coefficients required the preparation of standard curves. An inverse relationship between retention
time and mass accumulated on the SPME fibre was noted. Hydrocarbons from C5 to C16 were dated and those up to C11 were quantified. Total volatiles were quantified using six calibration curves. Biodegradation of volatile hydrocarbons during
growth on crude oil was faster and more complete with a mixed culture than pure isolates derived therefrom. The mixed culture
degraded 55% of the compounds by weight in 4 days versus 30–35% by pure cultures of Pseudomonas aeruginosa, Rhodococcus globerulus or a co-culture of the two. The initial degradation rate was threefold higher for the mixed culture, reaching 45% degradation
after 48 h. For the mixed culture, the degradation rate of individual alkanes was proportional to the initial concentration,
decreasing from hexane to undecane. P. fluorescens was unable to degrade any of the low-molecular-weight hydrocarbons and methylcyclohexane was recalcitrant in all cases. Overall,
the method was found to be reliable and cost-effective. Journal of Industrial Microbiology & Biotechnology (2000) 25, 155–162.
Received 04 March 2000/ Accepted in revised form 25 June 2000 相似文献
2.
Jiménez N Viñas M Guiu-Aragonés C Bayona JM Albaigés J Solanas AM 《Applied microbiology and biotechnology》2011,91(3):823-834
A laboratory experiment was conducted to identify key hydrocarbon degraders from a marine oil spill sample (Prestige fuel oil), to ascertain their role in the degradation of different hydrocarbons, and to assess their biodegradation potential
for this complex heavy oil. After a 17-month enrichment in weathered fuel, the bacterial community, initially consisting mainly
of Methylophaga species, underwent a major selective pressure in favor of obligate hydrocarbonoclastic microorganisms, such as Alcanivorax and Marinobacter spp. and other hydrocarbon-degrading taxa (Thalassospira and Alcaligenes), and showed strong biodegradation potential. This ranged from >99% for all low- and medium-molecular-weight alkanes (C15–C27) and polycyclic aromatic hydrocarbons (C0- to C2- naphthalene, anthracene, phenanthrene, dibenzothiophene, and carbazole), to 75–98% for higher molecular-weight alkanes (C28–C40) and to 55–80% for the C3 derivatives of tricyclic and tetracyclic polycyclic aromatic hydrocarbons (PAHs) (e.g., C3-chrysenes), in 60 days. The numbers of total heterotrophs and of n-alkane-, aliphatic-, and PAH degraders, as well as the structures of these populations, were monitored throughout the biodegradation
process. The salinity of the counting medium affects the counts of PAH degraders, while the carbon source (n-hexadecane vs. a mixture of aliphatic hydrocarbons) is a key factor when counting aliphatic degraders. These limitations notwithstanding,
some bacterial genera associated with hydrocarbon degradation (mainly belonging to α- and γ-Proteobacteria, including the
hydrocarbonoclastic Alcanivorax and Marinobacter) were identified. We conclude that Thalassospira and Roseobacter contribute to the degradation of aliphatic hydrocarbons, whereas Mesorhizobium and Muricauda participate in the degradation of PAHs. 相似文献
3.
Biodegradation of a crude oil by three microbial consortia of different origins and metabolic capabilities 总被引:6,自引:0,他引:6
Viñas M Grifoll M Sabaté J Solanas AM 《Journal of industrial microbiology & biotechnology》2002,28(5):252-260
Microbial consortia were obtained three by sequential enrichment using different oil products. Consortium F1AA was obtained
on a heavily saturated fraction of a degraded crude oil; consortium TD, by enrichment on diesel and consortium AM, on a mixture
of five polycyclic aromatic hydrocarbons [PAHs]. The three consortia were incubated with a crude oil in order to elucidate
their metabolic capabilities and to investigate possible differences in the biodegradation of these complex hydrocarbon mixtures
in relation to their origin. The efficiency of the three consortia in removing the saturated fraction was 60% (F1AA), 48%
(TD) and 34% (AM), depending on the carbon sources used in the enrichment procedures. Consortia F1AA and TD removed 100% of
n-alkanes and branched alkanes, whereas with consortium AM, 91% of branched alkanes remained. Efficiency on the polyaromatic
fraction was 19% (AM), 11% (TD) and 7% (F1AA). The increase in aromaticity of the polyaromatic fraction during degradation
of the crude oil by consortium F1AA suggested that this consortium metabolized the aromatic compounds primarily by oxidation
of the alkylic chains. The 500-fold amplification of the inocula from the consortia by subculturing in rich media, necessary
for use of the consortia in bioremediation experiments, showed no significant decrease in their degradation capability. Journal of Industrial Microbiology & Biotechnology (2002) 28, 252–260 DOI: 10.1038/sj/jim/7000236
Received 12 July 2001/ Accepted in revised form 11 November 2001 相似文献
4.
In this study biodegradation of hydrocarbons in thin oil films was investigated in seawater at low temperatures, 0 and 5 °C.
Heterotrophic (HM) or oil-degrading (ODM) microorganisms enriched at the two temperatures showed 16S rRNA sequence similarities
to several bacteria of Arctic or Antarctic origin. Biodegradation experiments were conducted with a crude mineral oil immobilized
as thin films on hydrophobic Fluortex adsorbents in nutrient-enriched or sterile seawater. Chemical and respirometric analysis
of hydrocarbon depletion showed that naphthalene and other small aromatic hydrocarbons (HCs) were primarily biodegraded after
dissolution to the water phase, while biodegradation of larger polyaromatic hydrocarbons (PAH) and C10–C36 n-alkanes, including n-hexadecane, was associated primarily with the oil films. Biodegradation of PAH and n-alkanes was significant at both 0 and 5°C, but was decreased for several compounds at the lower temperature. n-Hexadecane biodegradation at the two temperatures was comparable at the end of the experiments, but was delayed at 0°C. Investigations
of bacterial communities in seawater and on adsorbents by PCR amplification of 16S rRNA gene fragments and DGGE analysis indicated
that predominant bacteria in the seawater gradually adhered to the oil-coated adsorbents during biodegradation at both temperatures.
Sequence analysis of most DGGE bands aligned to members of the phyla Proteobacteria (Gammaproteobacteria) or Bacteroidetes. Most sequences from experiments at 0°C revealed affiliations to members of Arctic or Antarctic consortia, while no such
homology was detected for sequences from degradation experiment run at 5°C. In conclusion, marine microbial communities from
cold seawater have potentials for oil film HC degradation at temperatures ≤5°C, and psychrotrophic or psychrophilic bacteria
may play an important role during oil HC biodegradation in seawater close to freezing point. 相似文献
5.
Batch experiments were conducted to evaluate the biodegradation rates of limonene, α-pinene, γ-terpinene, terpinolene and
α-terpineol at 23 °C under aerobic conditions. Biodegradation was demonstrated by the depletion of monoterpene mass, CO2 production and a corresponding increase in biomass. Monoterpene degradation in liquid cultures devoid of soil followed Monod
kinetics. The maximum specific growth rate (μmax) was 0.02 h−1 and 0.06 h−1 and the half-velocity constant (K
s ) varied from 32 mg/l to 3 mg/l for the limonene and α-terpineol respectively. The recovery of monoterpenes by solvent extraction
from autoclaved and azide-amended soil-slurry samples decreased over time and ranged from 69% to 73% for 120 h of incubation
period. Although a significant fraction of monoterpene hydrocarbon could not be extracted, mineralization of these compounds
in the soil-slurry systems took place, as shown by CO2 production. The soil-normalized degradation rates for the hydrocarbon monoterpenes ranged from 0.6 μg g−1 h−1 to 2.1 μg g−1 h−1. A kinetic model – which combined monoterpene biodegradation in the liquid phase and net desorption – was developed and applied
to data obtained from soil-slurry assays.
Received: 10 September 1996 / Received revision: 16 December 1996 / Accepted: 10 January 1997 相似文献
6.
G. Misra S. G. Pavlostathis E. M. Perdue R. Araujo 《Applied microbiology and biotechnology》1996,45(6):831-838
Batch experiments were conducted to assess the biotransformation potential of four hydrocarbon monoterpenes (d-limonene, α-pinene, γ-terpinene, and terpinolene) and four alcohols (arbanol, linalool, plinol, and α-terpineol) under aerobic
conditions at 23°C. Both forest-soil extract and enriched cultures were used as inocula for the biodegradation experiments
conducted first without, then with prior microbial acclimation to the monoterpenes tested. All four hydrocarbons and two alcohols
were readily degraded. The increase in biomass and headspace CO2 concentrations paralleled the depletion of monoterpenes, thus confirming that terpene disappearance was the result of biodegradation
accompanied by microbial growth and mineralization. Plinol resisted degradation in assays using inocula from diverse sources,
while arbanol degraded very slowly. A significant fraction of d-limonene-derived carbon was accounted for as non-extractable, dissolved organic carbon, whereas terpineol exhibited a much
higher degree of utilization. The rate and extent of monoterpene biodegradation were not significantly affected by the presence
of dissolved natural organic matter.
Received: 27 November 1995/Received last revision: 15 March 1996/Accepted: 17 March 1996 相似文献
7.
Laboratory degradation studies of two indigeneously produced linear alkyl benzenes byNocardia amarae MB-11 isolated from soil showed an overall degradation of linear alkyl benzenes isomers to the extent of 57–70%. Degradation
of 2-phenyl isomers of linear alkyl benzenes was complete and faster than that of other phenyl position (C3–C7) isomers which
were degraded to the extent of 40–72% only. Length of alkyl side chains (C10–C14) had little or no impact on the degradation pattern. Major metabolities detected were 2-, 3-and 4-phenyl butyric acids, phenyl
acetic acid and cis, cis-muconic acid. Minor metabolites weretrans-cinnamic acid, 4-phenyl 3-butenoic acid and 3-phenyl pentanoic acid along with two unidentified hydroxy acids. On the basis
of the formation pattern of these metabolities, three catabolic pathways of linear alkyl benzenes isomers inNocardia amarae MB-11 were postulated. All the phenyl position (C2–C7) isomers of C10, C12, and C14 linear alkyl benzenes along with 3-phenyl and 5-phenyl isomers of C11 and C13 linear alkyl benzenes were degraded viacis,cis-muconic acid pathway. Other phenyl position isomers of C11 and C13 linear alkyl benzenes with phenyl substitution at even number carbon atoms were principally degraded via phenyl acetic acid
pathway whiletrans-cinnamic acid formation provided a minor pathway 相似文献
8.
陕北地区石油污染土壤中不动杆菌属的筛选、鉴定及降解性能 总被引:2,自引:0,他引:2
从陕北地区石油污染土壤中分离鉴定得到两株不动杆菌属(Acinetobacter sp.)的高效石油降解菌A.sp 1和A.sp 2,分别从盐浓度、pH值、氮源、磷源和接种量等因素进行研究以确定其最佳石油降解条件,并进一步通过GC-MS(Gas ChromatographyMass Spectrometer)方法分析其在最佳条件下对原油组分的不同降解性能。结果显示:A.sp 1在盐浓度为1%、pH值为6—7、磷源为KH2PO4和K2HPO4、氮源为尿素和接种量为4%的条件下,最高降解率可达到60%。A.sp 2在盐浓度为1%、pH值为7—9、磷源为KH2PO4和K2HPO4、氮源为硝酸铵和接种量为8%的条件下,最高降解率可达到67%。GC-MS分析结果表明,菌株A.sp 1对石油烃类C21—C25有明显的降解效果,菌株A.sp 2对石油烃类C20—C30的降解效果较好。 相似文献
9.
Solano-Serena F Marchal R Huet T Lebeault JM Vandecasteele JP 《Applied microbiology and biotechnology》2000,54(1):121-125
The biodegradability under aerobic conditions of volatile hydrocarbons (4–6 carbons) contained in gasoline and consisting
of n-alkanes, iso-alkanes, cycloalkanes and alkenes, was investigated. Activated sludge was used as the reference microflora. The biodegradation
test involved the degradation of the volatile fraction of gasoline in closed flasks under optimal conditions. The kinetics
of biodegradation was monitored by CO2 production. Final degradation was determined by gas chromatographic analysis of all measurable hydrocarbons (12 compounds)
in the mixture after sampling the headspace of the flasks. The degradation of individual hydrocarbons was also studied with
the same methodology. When incubated individually, all hydrocarbons used as carbon sources, except 2,2-dimethylbutane and
2,3-dimethylbutane, were completely consumed in 30 days or less with different velocities and initial lag periods. When incubated
together as constituents of the light gasoline fraction, all hydrocarbons were metabolised, often with higher velocities than
for individual compounds. Cometabolism was involved in the degradation of dimethyl isoalkanes.
Received: 19 October 1999 / Received revision: 21 January 2000 / Accepted: 23 January 2000 相似文献
10.
M. Eriksson G. Dalhammar A.-K. Borg-Karlson 《Applied microbiology and biotechnology》1999,51(4):532-535
A hydrocarbon mixture containing p-xylene, naphthalene, Br-naphthalene and straight aliphatic hydrocarbons (C14 to C17) was aerobically degraded without lag phase by a natural uncontaminated potting soil at 20 °C and 6 °C. Starting concentrations
were approximately 46 ppm for the aromatic and 13 ppm for the aliphatic compounds. All aliphatic hydrocarbons were degraded
within 5 days at 20 °C, to levels below detection (ppb levels) but only down to 10% of initial concentration at 6 °C. Naphthalene
was degraded within 12 days at 20 °C and unaffected at 6 °C. At 20 °C p-xylene was degraded within 20 days, but no degradation occurred at 6 °C. Br-naphthalene was only removed down to 30% of initial
concentration at 20 °C, with no significant effect at 6 °C. The biodegradation was monitored with head space solid-phase microextraction
and gas chromatography–mass spectrometry.
Received: 5 October 1998 / Received revision: 4 December 1998 / Accepted: 5 December 1998 相似文献
11.
Enhanced biodegradation of phenanthrene using different inoculum types in a microbial fuel cell
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Environmental pollution by petroleum hydrocarbons from contaminated groundwater and soils is a serious threat to human health. Microbial fuel cells (MFCs) could be employed in the treatment of these recalcitrant pollutants with concomitant bioelectricity generation. In this study, the use of MFCs in biodegradation of phenanthrene, a model hydrocarbon, was investigated with respect to its biodegradation rate, biodegradation efficiency, and power production using a range of inocula (Shewanella oneidensis MR1 14063, Pseudomonas aeruginosa NCTC 10662, mixed cultures, and combinations thereof). All the inocula showed high potentials for phenanthrene degradation with a minimum degradation efficiency of 97%. The best overall performing inoculum was anaerobically digested sludge supplemented with P. aeruginosa NCTC 10662, having a degradation rate, maximum power density and chemical oxygen demand removal efficiency of 27.30 μM/d, 1.25 mW/m2 and 65.6%, respectively. Adsorption of phenanthrene on the carbon anode was also investigated; it conformed to a Type II adsorption isotherm and could be modelled using a modified Brunauer, Emmett and Teller model with a maximum monolayer capacity of 0.088 mg/cm2. This work highlights the possibility of using MFCs to achieve high degradation rates of phenanthrene through co‐metabolism and could potentially be used as a replacement of permeable reactive barriers for remediation of hydrocarbon‐contaminated groundwater. 相似文献
12.
Maneerat S Bamba T Harada K Kobayashi A Yamada H Kawai F 《Applied microbiology and biotechnology》2006,70(2):254-259
A marine bacterium, Myroides sp. SM1, can grow on weathered crude oil and show emulsification of it. The biosurfactant able to emulsify crude oil was
excreted in culture supernatant of Myroides sp. SM1 grown on marine broth, which was extracted with chloroform/methanol (1:1) at pH 7 and purified by normal and reverse
phase silica gel column chromatographies. The compound was ninhydrin-positive, and the chemical structure was elucidated by
nuclear magnetic resonance (NMR), infrared spectroscopy (IR), fast atom bombardment mass spectrometry, and gas chromatography–mass
spectrometry (GC-MS) to be a mixture of l-ornithine lipids, which were composed of l-ornithine and a different couple of iso-3-hydroxyfatty acid (C15–C17) and iso-fatty acid (C15 or C16) in a ratio of 1:1:1. The critical micelle concentration for a mixture of ornithine lipids was measured to be approximately
40 mg/l. A mixture of ornithine lipids exhibited emulsifying activity for crude oil in a broad range of pH, temperature, and
salinity and showed higher surface activity for oil displacement test than other several artificial surfactants and a biosurfactant,
surfactin. 相似文献
13.
Degradation and mineralization of petroleum by two bacteria isolated from coastal waters 总被引:14,自引:0,他引:14
Within the framework of a study on the oil biodegradation potential of the sea the ability of a Flavobacterium sp. and Brevibacterium sp. to metabolize a paraffinic crude oil and a chemically defined hydrocarbon mixture was investigated. Major components of the crude oil were identified by combination gas chromatography and mass spectrometry. The rate and extent of total hydrocarbon biodegradation was measured. In addition, CO2 evolution from the crude oil was continuously monitored in a shaker-mounted gas train arrangement. Degradation started after a 2 to 4 day lag period, and reached its maximum within two weeks. At this time up to 60% of the crude oil and 75% of the model hydrocarbon mixture, each added at the level of 1 ml per 100 ml artificial sea water, were degraded. Mineralization(conversion to CO2) was slightly lower due to formation of products and bacterial cell material. n-Paraffins were preferentially degraded as compared to branched chain hydrocarbons. Biodegradation of n-paraffins in the range of C12 to C20 was simultaneous; no diauxie effects were observed. 相似文献
14.
P. A. Vieira S. Faria R. B. Vieira F. P. De França V. L. Cardoso 《World journal of microbiology & biotechnology》2009,25(3):427-438
In order to optimize and evaluate the influence of nitrogen, phosphorus, and inoculum concentrations on the biodegradation
of hydrocarbon contaminated effluents, experiments based on central composite design (CCD) method were carried out for 3 days,
employing C1 mixed culture and intermittent aeration. The independent variables were nitrogen concentration (X
1), phosphorus concentration (X
2), and inoculum concentration (X
3) and the removal of total petroleum hydrocarbons (TPH) was the dependent variable. The optimized nutrients ratio (C:N:P = 100:20:2.7)
and inoculum concentration (1.32 g/l) provided TPH removal of 71.8% after processing for three days. Analysis using gas chromatography
identified five hydrocarbons classes: paraffins, isoparaffins, olefins, naphthenics, and aromatics. The naphthenic compounds
did not degrade as readily as the other hydrocarbons that were identified. The following degradation percentages were obtained:
87.1% for the paraffins, 77.7% for the isoparaffins, 78.6% for the olefins, 38.4% for the naphthenics, and 71.7% for the aromatics. 相似文献
15.
Statistical optimization of the biodegradation of two keratinous wastes directed by Bacillus subtilis recombinant cells was carried out by means of a response surface methodology. A Box–Behnken design was employed to predict
the optimal levels of three variables namely, keratin percent, incubation time and inoculum size. Analysis of variance revealed
that, only keratin percent had the highest significant effect. Canonical analysis and ridge max analysis were used to get
the optimal levels of the three predictors along with the optimum levels of the responses. The optimal sets of predicted and
validated levels of the three variables were [7.69% (w/v) feathers, 96.58 h and 1.28% (v/v) inoculum size] and [8% (w/v) feathers,
98.45 h, 3.9% (v/v) inoculum size] to achieve the highest levels of soluble proteins (1.25–1.7 mg/ml) and NH2-free amino groups (245.82–270.0 μmol leucine/ml), respectively upon using three optimized feathers-based media. These values
represented 83.67–100% and 100% adequacy for the models of soluble proteins and NH2-free amino groups, respectively. While, [8.23% (w/v) sheep wool, 5.52% (v/v) inoculum size and 46.58 h] and [8.33% (w/v)
sheep wool, 5.89% (v/v) inoculum size and 63.46 h] were the optimal sets of predicted and validated levels of the above variables
to achieve the highest yields of soluble proteins (3.4–4.6 mg/ml) and NH2-free amino groups (290.9–302.0 μmol leucine/ml), respectively upon using three optimized sheep wool-based media. These values
represented 100% adequacy for the models of soluble proteins and NH2-free amino groups. By the end of the optimization strategy, a fold enhancement (2.14–2.43 and 1.78–2.12) in the levels of
released soluble proteins and NH2-free amino groups, respectively was obtained upon using three optimized feathers-based media. However, a fold enhancement
(4.25–5.75 and 2.42–2.5) in the levels of soluble proteins and NH2-free amino groups, respectively was obtained upon using three optimized sheep wool-based media. Data would encourage pilot
scale optimization of the biodegradation of these wastes. 相似文献
16.
C. S. Jeong H. N. Murthy E. J. Hahn H. L. Lee K. Y. Paek 《Acta Physiologiae Plantarum》2009,31(1):219-222
The effect of the root-inoculum size and axuin concentration on growth of adventitious roots and accumulation of ginsenosides
were studied during suspension cultures of ginseng (Panax ginseng C.A. Meyer). Of the various concentrations of indole-3-butyric acid (IBA) and γ-naphthaleneacetic acid (NAA) used as supplementary
growth regulators along with Murashige and Skoog medium, 25 μM IBA was found suitable for lateral root induction and growth,
as well as accumulation of ginsenosides. Inoculum size of 5 g L−1 was found suitable for optimal biomass (10.5 g L−1 dry biomass) and ginsenosides (5.4 mg g−1 DW) accumulation. Of the various length of root inocula tested (chopped to 1–3, 4–6, 7–10 mm and un-chopped), root inocula
of 7–10 mm was found suitable for biomass and ginsenoside accumulation. 相似文献
17.
The assessment of biodegradation activity in contaminated aquifers is critical to demonstrate the performance of bioremediation
and natural attenuation and to parameterize models of contaminant plume dynamics. Real time quantitative PCR (qPCR) was used
to target the catabolic bssA gene (coding for benzylsuccinate synthase) and a 16S rDNA phylogenetic gene (for total Bacteria) as potential biomarkers to infer on anaerobic toluene degradation rates. A significant correlation (P = 0.0003) was found over a wide range of initial toluene concentrations (1–100 mg/l) between toluene degradation rates and
bssA concentrations in anaerobic microcosms prepared with aquifer material from a hydrocarbon contaminated site. In contrast,
the correlation between toluene degradation activity and total Bacteria concentrations was not significant (P = 0.1125). This suggests that qPCR targeting of functional genes might offer a simple approach to estimate in situ biodegradation
activity, which would enhance site investigation and modeling of natural attenuation at hydrocarbon-contaminated sites. 相似文献
18.
Anna Dürig Irene Kouskoumvekaki Rebecca M. Vejborg Per Klemm 《Applied microbiology and biotechnology》2010,87(1):309-317
The fuel oxygenate, methyl tert-butyl ether (MTBE), although now widely banned or substituted, remains a persistent groundwater contaminant. Multidimensional
compound-specific isotope analysis (CSIA) of carbon and hydrogen is being developed for determining the extent of MTBE loss
due to biodegradation and can also potentially distinguish between different biodegradation pathways. Carbon and hydrogen
isotopic fractionation factors were determined for MTBE degradation in aerobic and anaerobic laboratory cultures. The carbon
isotopic enrichment factor (εC) for aerobic MTBE degradation by a bacterial consortium containing the aerobic MTBE-degrading bacterium, Variovorax paradoxus, was −1.1 ± 0.2‰ and the hydrogen isotope enrichment factor (εH) was −15 ± 2‰. This corresponds to an approximated lambda value (Λ = εH/εC) of 14. Carbon isotope enrichment factors for anaerobic MTBE-degrading enrichment cultures were −7.0 ± 0.2‰ and did not vary
based on the original inoculum source, redox condition of the enrichment, or supplementation with syringic acid as a co-substrate.
The hydrogen enrichment factors of cultures without syringic acid were insignificant, however a strong hydrogen enrichment
factor of −41 ± 3‰ was observed for cultures which were fed syringic acid during MTBE degradation. The Λ = 6 obtained for NYsyr cultures might be diagnostic for the stimulation of anaerobic MTBE degradation by methoxylated compounds
by an as yet unknown pathway and mechanism. The stable-isotope enrichment factors determined in this study will enhance the
use of CSIA for monitoring anaerobic and aerobic MTBE biodegradation in situ. 相似文献
19.
Stimulation of the multiplication of Micrococcus luteus by an autocrine growth factor 总被引:12,自引:0,他引:12
Galina V. Mukamolova Svetlana S. Kormer Douglas B. Kell A. S. Kaprelyants 《Archives of microbiology》1999,172(1):9-14
Viable cells of Micrococcus luteus secrete a proteineous growth factor (Rpf) which promotes the resuscitation of dormant, nongrowing cells to yield normal,
colony-forming bacteria. When washed M. luteus cells were used as an inoculum, there was a pronounced influence of Rpf on the true lag phase and cell growth on lactate
minimal medium. In the absence of Rpf, there was no increase in colony-forming units for up to 10 days. When the inoculum
contained less than 105 cells ml–1, macroscopically observable M. luteus growth was not obtained in succinate minimal medium unless Rpf was added. Incubation of M. luteus in the stationary phase for 100 h resulted in a failure of the cells to grow in lactate minimal medium from inocula of small
size although the viability of these cells was close to 100% as estimated using agar plates made from lactate minimal medium
or rich medium. The underestimation of viable cells by the most-probable-number (MPN) method in comparsion with colony-forming
units was equivalent to the requirement that at least 105 cells grown on succinate medium, 103 cells from old stationary phase, or approximately 10–500 washed cells are required per millilitre of inoculum for growth
to lead to visible turbidity. The addition of Rpf in the MPN dilutions led to an increase of the viable cell numbers estimated
to approximately the same levels as those determined by colony-forming units. Thus, a basic principle of microbiology –“one
cell-one culture”– may not be applicable in some circumstances in which the metabolic activity of “starter” cells is not sufficient
to produce enough autocrine growth factor to support cell multiplication.
Received: 7 December 1998 / Accepted: 7 April 1999 相似文献
20.
In a previous study, a marine isolate Clostridium sp. EDB2 degraded 2,4,6,8,10,12-hexanitro-2,4,6,8,10,12-hexaazaisowurtzitane (CL-20) under anaerobic conditions (Bhushan B, Halasz A, Thiboutot S, Ampleman G, Hawari J (2004c) Chemotaxis-mediated biodegradation of cyclic nitramine explosives RDX, HMX, and CL-20 by Clostridium sp. EDB2. Biochem Biophys Res Commun 316:816–821); however, the enzyme responsible for CL-20 degradation was not known. In the present study, we isolated and purified an enzyme, from strain EDB2, responsible for CL-20 degradation. The enzyme was membrane-associated and NADH-dependent and had a molecular weight of 56 kDa (with SDS-PAGE). N-terminal amino acid sequence of enzyme revealed that it belonged to dehydrogenase class of enzymes. The purified enzyme degraded CL-20 at a rate of 18.5 nmol/h mg protein under anaerobic conditions. Carbon and nitrogen mass balance of the products were 100 and 64%, respectively. In LC–MS–MS studies, we detected three different initial metabolites from CL-20, i.e., mono-nitroso derivative, denitrohydrogenated product, and double-denitrated isomers with molecular weight of 422, 393, and 346 Da, corresponding to presumed empirical formulas of C6H6N12O11, C6H7N11O10, and C6H6N10O8, respectively. Identity of all the three metabolites were confirmed by using ring-labeled [15N]CL-20 and the nitro-group-labeled [15NO2]CL-20. Taken together, the above data suggested that the enzyme degraded CL-20 via three different routes: Route A, via two single electron transfers necessary to release two nitro-groups from CL-20 to produce two double-denitrated isomers; Route B, via a hydride transfer necessary to produce a denitrohydrogenated product; and Route C, via transfer of two redox equivalents to CL-20 necessary to produce a mono-nitroso derivative of CL-20. This is the first biochemical study which showed that CL-20 degradation can be initiated via more than one pathway. 相似文献