Biomedical subjects

F Lépine

Publications and source records attributed to F Lépine.

At least 19 recordsLinked to original sources

PubMed2005

Desulfitobacterium hafniense is present in a high proportion within the biofilms of a high-performance pentachlorophenol-degrading, methanogenic fixed-film reactor.

We developed a pentachlorophenol (PCP)-degrading, methanogenic fixed-film reactor by using broken granular sludge from an upflow anaerobic sludge blanket reactor. This methanogenic consortium was acclimated with increasing concentrations of PCP. After 225 days of acclimation, the reactor was performing at a high level, with a PCP removal rate of 1,173 muM day(-1), a PCP removal efficiency of up to 99%, a degradation efficiency of approximately 60%, and 3-chlorophenol as the main chlorophenol residual intermediate. Analyses by PCR-denaturing gradient gel electrophoresis (DGGE) showed that Bacteria and Archaea in the reactor stabilized in the biofilms after 56 days of operation. Important modifications in the profiles of Bacteria between the original granular sludge and the reactor occurred, as less than one-third of the sludge DGGE bands were still present in the reactor. Fluorescence in situ hybridization experiments with probes for Archaea or Bacteria revealed that the biofilms were composed mostly of Bacteria, which accounted for 70% of the cells. With PCR species-specific primers, the presence of the halorespiring bacterium Desulfitobacterium hafniense in the biofilm was detected very early during the reactor acclimation period. D. hafniense cells were scattered in the biofilm and accounted for 19% of the community. These results suggest that the presence of PCP-dehalogenating D. hafniense in the biofilm was crucial for the performance of the reactor.

Anaerobiosis↗

PubMed2005

Molecular analysis of Desulfitobacterium frappieri pcp-1 involved in reductive dehalogenation of pentachlorophenol.

Desulfitobacterium are Gram positive, spore-forming, strictly anaerobic bacteria, that belong to the Firmicutes, Clostridia, Clostridiales, and Peptococcaceae. Most known members of the genus Desulfitobacterium have the ability to dechlorinate several halogenated compounds by a mechanism of reductive dehalogenation and use them as electron acceptors to generate energy (halorespiration). Desulfitobacteria are therefore perfect candidates to be used in bioremediation treatments of environment polluted with halogenated compounds. Understanding the physiology and the molecular mechanisms of these bacteria will help to develop better bioremediation systems. This report summarizes works that have been done in our laboratories with D. frappieri PCP-1 on reductive dehalogenases, genes encoding these dehalogenases and their expression, and the development of lab-scale PCP-degrading reactors using this bacterium.

Biodegradation, Environmental↗

PubMed2004

Observation of large quadrupolar effects in a slow photoelectron imaging experiment.

We have studied nondipolar effects in resonance-enhanced multiphoton ionization of Xe and have observed an azimuthal dependence of the photoelectron angular distribution on a quadrupole resonance, as well as a very large asymmetry with respect to the direction of the laser propagation close to the resonance, which is understood in terms of interference between dipole- and quadrupole-allowed ionization channels. The observed asymmetry in the photoelectron angular distribution provides insight into the ejection of slow photoelectrons near an ionization threshold.

Journal Article↗

PubMed2003

A stable isotope dilution assay for the quantification of the Pseudomonas quinolone signal in Pseudomonas aeruginosa cultures.

A stable isotope dilution method was developed to analyse 2-heptyl-3,4-dihydroxyquinoline, also called the Pseudomonas quinolone signal (PQS), directly in Pseudomonas aeruginosa cultures by liquid chromatography coupled to mass spectrometry (LC/MS). PQS, along with the isobaric 2-heptyl-4-hydroxyquinoline N-oxide (HQNO), were quantified in various Pseudomonas liquid cultures using a deuterated PQS analog as internal standard. The kinetic of production of these quinolines in a growing culture of P. aeruginosa PA14 showed that their production starts at the end of the logarithmic growth phase and is maximal at the onset of the stationary growth phase. The concentration of PQS reached a maximum at 13 mg/l and then decreased, while the HQNO concentration reached 18 mg/l and then remained stable. Culture supernatants of P. aeruginosa strains PAO1 and PA14 produced similar concentrations of PQS whereas no PQS or HQNO could be detected in culture supernatants of the P. aeruginosa strain PAK or in the other Pseudomonas species tested, including phytopathogenic pseudomonads.

Hydroxyquinolines↗

PubMed2003

Initial characterization of new bacteria degrading high-molecular weight polycyclic aromatic hydrocarbons isolated from a 2-year enrichment in a two-liquid-phase culture system.

AIMS: To characterize some polycyclic aromatic hydrocarbons (PAH)-degrading microorganisms isolated from an enriched consortium degrading high molecular weight (HMW) PAHs in a two-liquid-phase (TLP) soil slurry bioreactor, and to determine the effect of low molecular weight (LMW) PAH on their growth and HMW PAH-degrading activity. METHODS AND RESULTS: Several microorganisms were isolated from a HMW-PAH (pyrene, chrysene, benzo[a]pyrene and perylene) degrading consortium enriched in TLP cultures using silicone oil as the organic phase. From 16S rRNA analysis, four isolates were identified as Mycobacterium gilvum B1 (99% identity),Bacillus pumilus B44 (99% identity), Microbacterium esteraromaticum B21 (98% identity), and to the genus Porphyrobacter B51 (96% identity). The two latter isolates have not previously been associated with PAH degradation. Isolate B51 grew strongly in the interfacial fraction in the presence of naphthalene vapours and phenanthrene compared with cultures without LMW PAHs. Benzo[a]pyrene was degraded in cultures containing a HMW PAH mixture but pyrene had no effect on its degradation. The growth of isolates B1 and B21 was improved in the aqueous phase than in the interfacial fraction for cultures with naphthalene vapours. Pyrene was required for benzo[a]pyrene degradation by isolate B1. For isolate B21, pyrene and chrysene were degraded only in cultures without naphthalene vapours. CONCLUSION: Consortium enriched in a TLP culture is composed of microorganisms with different abilities to grow at the interface or in the aqueous phase according to the culture conditions and the PAH that are present. Naphthalene vapours increased the growth of the microorganisms in TLP cultures but did not stimulate the HMW PAH degradation. SIGNIFICANCE AND IMPACT OF THE STUDY: New HMW PAH-degrading microorganisms and a better understanding of the mechanisms involved in HMW PAH degradation in TLP cultures.

Bacteria↗

PubMed2002

Photoionization microscopy.

We present the first experimental results of a technique called photoionization microscopy. Photoelectrons ejected in threshold photoionization of Xe are detected in a velocity map imaging apparatus, and interferences between various trajectories by which the electron moves from the atom to the detector are observed. The structure of the interference pattern, which contains the transverse component of the electronic wave function, evolves smoothly with the excess energy above the saddle point. The main observed features are interpreted within the framework of the semiclassical approximation.

Journal Article↗

PubMed2001

Geographic distribution of Desulfitobacterium frappieri PCP-1 and Desulfitobacterium spp. in soils from the province of Quebec, Canada.

The presence of indigenous Desulfitobacterium species in 44 soil samples taken from various sites in the southern part of the province of Quebec (Canada) and four from locations outside Quebec was investigated. Twenty-four of these soils were sampled from contaminated industrial sites. Indigenous Desulfitobacterium bacteria from soil samples were enriched by cultivation in anaerobic soil slurry culture. Total DNA was then extracted from these slurries and polymerase chain reaction (PCR) amplifications were performed with primers targeting 16S ribosomal RNA gene sequences of Desulfitobacterium spp. and of Desulfitobacterium frappieri PCP-1. A positive PCR signal was obtained in 31 soil slurry cultures. Resolution of single-strand DNAs of some of the PCR products by a single-strand conformational polymorphism protocol suggests that more than one species of Desulfitobacterium were present in the corresponding slurry cultures. These results suggest that Desulfitobacterium are ubiquitous in soils in the province of Quebec, especially in soils from the St. Lawrence valley and the southern part of the province.

Journal Article↗

PubMed2001

Hydrolysis of 4-hydroxybenzoic acid esters (parabens) and their aerobic transformation into phenol by the resistant Enterobacter cloacae strain EM.

Enterobacter cloacae strain EM was isolated from a commercial dietary mineral supplement stabilized by a mixture of methylparaben and propylparaben. It harbored a high-molecular-weight plasmid and was resistant to high concentrations of parabens. Strain EM was able to grow in liquid media containing similar amounts of parabens as found in the mineral supplement (1,700 and 180 mg of methyl and propylparaben, respectively, per liter or 11.2 and 1.0 mM) and in very high concentrations of methylparaben (3,000 mg liter(-1), or 19.7 mM). This strain was able to hydrolyze approximately 500 mg of methyl-, ethyl-, or propylparaben liter(-1) (3 mM) in less than 2 h in liquid culture, and the supernatant of a sonicated culture, after a 30-fold dilution, was able to hydrolyze 1,000 mg of methylparaben liter(-1) (6.6 mM) in 15 min. The first step of paraben degradation was the hydrolysis of the ester bond to produce 4-hydroxybenzoic acid, followed by a decarboxylation step to produce phenol under aerobic conditions. The transformation of 4-hydroxybenzoic acid into phenol was stoichiometric. The conversion of approximately 500 mg of parabens liter(-1) (3 mM) to phenol in liquid culture was completed within 5 h without significant hindrance to the growth of strain EM, while higher concentrations of parabens partially inhibited its growth.

Biodegradation, Environmental↗

PubMed2001

Separation of a phenol carboxylating organism from a two-member, strict anaerobic co-culture.

In a culture converting phenol to benzoic acid under anaerobic conditions and previously described as being constituted of only a Clostridium-like strain 6, another bacterium (strain 7) was observed. Each organism was enriched by centrifugation on a Percoll gradient. Strain 6 was purified by dilution and plating. Strain 7 did not grow on solid media, but a strain 7 culture, cleared of strain 6, was obtained by subculturing in the presence of ampicillin and by dilution. In fresh medium, phenol was transformed by the reconstituted co-culture but not by each strain alone. In a supernatant from a co-culture or from a strain 6 culture, strain 7 alone transformed phenol but not strain 6. Maintenance of an active strain 7 in fresh medium instead of co-culture supernatant became possible when phenol was replaced by 4-hydroxybenzoate (4-OHB), which is decarboxylated to phenol before being transformed to benzoate. Even with 4-OHB, the use of co-culture (or strain 6 culture) supernatant resulted in faster transformation activity and growth rate. A phylogenetic analysis placed strain 7 in a cluster of uncultivated or nonisolated bacteria (92-96% homology). Strain 7 is also related to Desulfotomaculum, Desulfitobacterium, Desulfosporosinus, Moorella, and Sporotomaculum genera (87-92% homology).

Bacteria, Anaerobic↗

PubMed2000

Mass spectrometry monitoring of rhamnolipids from a growing culture of Pseudomonas aeruginosa strain 57RP.

Two rapid and simple methods for the characterisation and quantification of rhamnolipids produced by a growing culture of the Pseudomonas aeruginosa strain 57RP were developed. Two rhamnolipids were purified and their response factors determined. The various rhamnolipids produced were then measured using liquid chromatography/mass spectrometry. The culture supernatants were injected directly, without prior purification, in a HPLC equipped with a C(18) reverse-phase column. The complete profile of rhamnolipid congeners produced during a 2 week cultivation period was monitored. In order to shorten the analysis time, another method was developed which did not require chromatographic separation of the rhamnolipids prior to their detection. Quantification of rhamnolipids using the direct infusion method gave results very similar to those obtained with HPLC separation. These two methods were very well correlated with the standard colorimetric orcinol method. The rhamnolipid profiles obtained show that the various rhamnolipid congeners are secreted simultaneously, and that their relative proportion remained unchanged throughout the cultivation period.

Glycolipids↗

PubMed2000

Effects of bioaugmentation strategies in UASB reactors with a methanogenic consortium for removal of phenolic compounds.

The removal of phenol, ortho- (o-) and para- (p-)cresol was studied with two series of UASB reactors using unacclimatized granular sludges bioaugmented with a consortium enriched against these substances. The parameters studied were the amount of inoculum added to the sludges and the method of immobilization of the inoculum. Two methods were used, adsorption to the biomass or encapsulation within calcium alginate beads. In the bioaugmentation by adsorption experiment, and with a 10% inoculum, complete phenol removal was obtained after 36 d, while 178 d were required in the control reactor. For p-cresol, 95% removal was obtained in the bioaugmented reactor on day 48 while 60 d were required to achieve 90% removal in the control reactor. For o-cresol, the removals were only marginally better with the bioaugmented reactors. Tests performed with the reactors biomass under non-limiting substrate concentrations showed that the specific activities of the bioaugmented biomasses were larger than the original biomass for phenol, and p-cresol even after 276 of operations, showing that the inoculum bacteria successfully colonized the sludge granules. Immobilization of the inoculum by encapsulation in calcium alginate beads, was performed with 10% of the inoculum. Results showed that the best activities were obtained when the consortium was encapsulated alone and the beads added to the sludges. This reactor presented excellent activity and the highest removal of the various phenolic compounds a few days after start-up. After 90 d, a high-phenolic compounds removal was still observed, demonstrating the effectiveness of the encapsulation technique for the start-up and maintenance of high-removal activities.

Adsorption↗

PubMed2000

Optimization of high-molecular-weight polycyclic aromatic hydrocarbons' degradation in a two-liquid-phase bioreactor.

A microbial consortium degrading the high-molecular-weight polycyclic aromatic hydrocarbons (HMW PAHs) pyrene, chrysene, benzo[a]pyrene and perylene in a two-liquid-phase reactor was studied. The highest PAH-degrading activity was observed with silicone oil as the water-immiscible phase; 2,2,4,4,6,8, 8-heptamethylnonane, paraffin oil, hexadecane and corn oil were much less, or not efficient in improving PAH degradation by the consortium. Addition of surfactants (Triton X-100, Witconol SN70, Brij 35 and rhamnolipids) or Inipol EAP22 did not promote PAH biodegradation. Rhamnolipids had an inhibitory effect. Addition of salicylate, benzoate, 1-hydroxy-2-naphtoic acid or catechol did not increase the PAH-degrading activity of the consortium, but the addition of low-molecular-weight (LMW) PAHs such as naphthalene and phenanthrene did. In these conditions, the degradation rates were 27 mg l-1 d-1 for pyrene, 8.9 mg l-1 d-1 for chrysene, 1.8 mg l-1 d-1 for benzo[a]pyrene and 0.37 mg l-1 d-1 for perylene. Micro-organisms from the interface were slightly more effective in degrading PAHs than those from the aqueous phase.

Benzo(a)pyrene↗

PubMed2000

Monitoring of Desulfitobacterium frappieri PCP-1 in pentachlorophenol-degrading anaerobic soil slurry reactors.

Anaerobic biodegradation of pentachlorophenol (PCP) was studied in rotative bioreactors containing 200 g of PCP-contaminated soil and 250 ml of liquid medium. Reactors were bioaugmented with cells of Desulfitobacterium frappieri strain PCP-1, a bacterium able to dehalogenate PCP to 3-chlorophenol. Cells of strain PCP-1 were detected by quantitative PCR for at least 21 days in reactors containing 500 mg of PCP per kg of soil but disappeared after 21 days in reactors with 750 mg of PCP per kg of soil. Generally, PCP was completely removed in less than 9 days in soils contaminated with 189 mg of PCP per kg of soil. Sorption of PCP to soil organic matter reduced its toxicity and enhanced the survival of strain PCP-1. In some non-inoculated reactors, the indigenous microorganisms of some soils were also able to degrade PCP. These results suggest that anaerobic dechlorination of PCP in soils by indigenous PCP-degrading bacteria, or after augmentation with D. frappieri PCP-1, should be possible in situ and ex situ when the conditions are favourable for the survival of the degrading microorganisms.

Anaerobiosis↗

PubMed2000

Purification and characterization of a 4-hydroxybenzoate decarboxylase from an anaerobic coculture.

The oxygen-sensitive 4-hydroxybenzoate decarboxylase (4OHB-DC) activity from a phenol-carboxylating coculture, consisting of Clostridium-like strain 6 and an unidentified strain 7, was studied. Assays done with cell extracts showed that the optimal pH was 5.0-6.5 and the Km was 5.4 mM. The activity decreased by 50% in the presence of 5 mM EDTA, and it was restored and even enhanced by the addition of Mg++, Mn++, Zn++, or Ca++. After purification, the molecular mass of the enzyme was estimated as 420 kDa by gel chromatography, and as 119 kDa by SDS-PAGE, suggesting a homotetrameric structure. Its pI was 5.6. The N-terminal amino acid sequence showed 95% and 76% homology with the pyruvate-flavodoxin oxidoreductase (nifJ gene product) from Enterobacter agglomerans and Klebsiella pneumoniae, respectively. The purified enzyme also slowly catalyzed the reverse reaction, that is the phenol carboxylation. These characteristics suggest that this enzyme is different from other known decarboxylases. This includes the 4OHB-DC from Clostridium hydroxybenzoicum, which is the only one that had been purified before.

Amino Acid Sequence↗

PubMed1999

Liquid chromatography/mass spectrometry analysis of mixtures of rhamnolipids produced by Pseudomonas aeruginosa strain 57RP grown on mannitol or naphthalene.

Liquid chromatography/mass spectrometry using electrospray ionisation was used to analyse rhamnolipids produced by a Pseudomonas aeruginosa strain with mannitol or naphthalene as carbon source. Identification and quantification of 28 different rhamnolipid congeners was accomplished using a reverse-phase C(18) column and a 30 min chromatographic run. Isomeric rhamnolipids that were not chromatographically resolved could be identified by interpretation of their mass spectra and their relative proportions estimated. The most abundant rhamnolipid produced on mannitol contained two rhamnoses and two 3-hydroxydecanoic acid groups. The most abundant rhamnolipid produced from naphthalene contained two rhamnoses and one 3-hydroxydecanoic acid group.

Chromatography, Liquid↗

PubMed1999

Removal of phenolic compounds from a petrochemical effluent with a methanogenic consortium.

A methanogenic consortium was used to degrade phenol and ortho- (o-) cresol from a specific effluent of a petrochemical refinery. This effluent did not meet the local environmental regulations for phenolic compounds (178 mg/L), oils and greases (61 mg/L), ammoniacal nitrogen (75 mg/L) or sulfides (3.2 mg/L). The consortium, which degrades phenol via its carboxylation to benzoic acid, was progressively adapted to the effluent. Despite the very high effluent toxicity (EC50 of 2% with Microtox), the adapted consortium degraded 97% of 156 mg/L phenol in the supplemented effluent after 13 days in batch cultures (serum bottle). The addition of proteose peptone to the effluent is essential for phenol degradation. o-cresol was also transformed but not meta- or para-cresols. A continuous flow fixed-film anaerobic bioreactor was developed with the consortium. Treating the effluent with the bioreactor reduced phenol and phenolic compounds concentrations by 97 and 83%, respectively, for a hydraulic residence time of 6 h. This treatment also reduced by about half the effluent toxicity. Oils and greases and ammoniacal nitrogen were not affected. Similar microbiological forms were observed in serum bottles and in the bioreactors with or without the petrochemical effluent. These results indicate that this methanogenic consortium can treat efficiently the phenolic compounds in this specific petrochemical effluent.

Bacteria, Anaerobic↗

PubMed1999

Simultaneous removal of phenol, ortho- and para-cresol by mixed anaerobic consortia.

Two different anerobic consortia, one removing phenol and ortho (o-) cresol and other removing para(p-) cresol, were cultivated in serum bottles using whey as cosubstrate substitute for proteose peptone. Phenol and p-cresol removal with the phenol-removing consortium were the same with 0.0125% (w/v) whey as with 0.05% proteose peptone. For the other consortium, 8 days were required to decrease the p-cresol concentration from 35 to 2 mg/L with 0.025% whey, while 35 days were required to achieve a similar removal with 0.5% proteose peptone. The two consortia were mixed and cultivated with 0.025% whey. Phenolic compound removal with the mixed consortia was as good as that achieved by each of the two initial consortia against their respective substrates. This removal activity was maintained after several transfers. In a continuous upflow fixed-film reactor, the mixed consortia removed over 98% of 150 mg/L of phenol and 35 mg/L of each o- and p-cresol in the influent at 29 degrees C, with 0.025% whey as cosubstrate. The hydraulic retention time (HRT) was 0.25 day, corresponding to a phenolic compound volumic loading rate of 880 mg/(L of reactor x day). Once the continuous flow reactor achieved constant phenolic compound removal, no intermediates were found in the effluent, while in serum bottles, m-toluic acid, an o-cresol intermediate, accumulated. Measurements of the specific activity for the uptake of different substrates demonstrated the presence of all trophic groups involved in methanogenic fermentation. These activities were, in mg of substrate/(g of volatile suspended solids x day), as follows: 849 +/- 25 for the acidogens; 554 +/- 15 for the acetogens; 934 +/- 37 for the aceticlastic methanogens; and 135 +/- 15 for the hydrogenophilic methanogens. Electron micrographs of the mixed consortia showed seven different morphological bacterial types, including Methanotrix-like bacteria.

Anaerobiosis↗

PubMed1998

Anaerobic biodegradation of pentachlorophenol in a contaminated soil inoculated with a methanogenic consortium or with Desulfitobacterium frappieri strain PCP-1.

Anaerobic biodegradation of pentachlorophenol (PCP) in a contaminated soil from a wood-treating industrial site was studied in soil slurry microcosms inoculated with a PCP-degrading methanogenic consortium. When the microcosms containing 10%-40% (w/v) soil were inoculated with the consortium, more than 90% of the PCP was removed in less than 30 days at 29 degrees C. Less-chlorinated phenols, mainly 3-chlorophenol were slowly degraded and accumulated in the cultures. Addition of glucose and sodium formate to the microcosms was not necessary, suggesting that the organic compounds in the soil can sustain the dechlorinating activity. Inoculation of Desulfitobacterium frappieri strain PCP-1 along with a 3-chlorophenol-degrading consortium in the microcosms also resulted in the rapid dechlorination of PCP and the slow degradation of 3-chlorophenol. Competitive polymerase chain reaction experiments showed that PCP-1 was present at the same level throughout the 21-day biotreatment. D. frappieri, strain PCP-1, inoculated into the soil microcosms, was able to remove PCP from soil containing up to 200 mg PCP/kg soil. However, reinoculation of the strain was necessary to achieve more than 95% PCP removal with a concentration of 300 mg and 500 mg PCP/kg soil. These results demonstrate that D. frappieri strain PCP-1 can be used effectively to dechlorinate PCP to 3-chlorophenol in contaminated soils.

Bacteria, Anaerobic↗