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Aragno, Michel
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Aragno, Michel
Affiliation principale
Email
Michel.Aragno@unine.ch
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Voici les éléments 1 - 7 sur 7
- PublicationMétadonnées seulementUse of the frc gene as a molecular marker to characterize oxalate-oxidizing bacterial abundance and diversity structure in soil(2009)
; ; Oxalate catabolism, which can have both medical and environmental implications, is performed by phylogenetically diverse bacteria. The formyl-CoA-transferase gene was chosen as a molecular marker of the oxalotrophic function. Degenerated primers were deduced from an alignment of frc gene sequences available in databases. The specificity of primers was tested on a variety of frc-containing and frc-lacking bacteria. The frc-primers were then used to develop PCR-DGGE and real-time SybrGreen PCR assays in soils containing various amounts of oxalate. Some PCR products from pure cultures and from soil samples were cloned and sequenced. Data were used to generate a phylogenetic tree showing that environmental PCR products belonged to the target physiological group. The extent of diversity visualised on DGGE pattern was higher for soil samples containing carbonate resulting from oxalate catabolism. Moreover, the amount of frc gene copies in the investigated soils was detected in the range of 1.64x10(7) to 1.75x10(8)/g of dry soil under oxalogenic tree (representing 0.5 to 1.2% of total 16S rRNA gene copies), whereas the number of frc gene copies in the reference soil was 6.4x10(6) (or 0.2% of 16S rRNA gene copies). This indicates that oxalotrophic bacteria are numerous and widespread in soils and that a relationship exists between the presence of the oxalogenic trees Milicia excelsa and Afzelia africana and the relative abundance of oxalotrophic guilds in the total bacterial communities. This is obviously related to the accomplishment of the oxalate-carbonate pathway, which explains the alkalinization and calcium carbonate accumulation occurring below these trees in an otherwise acidic soil. The molecular tools developed in this study will allow in-depth understanding of the functional implication of these bacteria on carbonate accumulation as a way of atmospheric CO2 sequestration. (c) 2008 Elsevier B.V. All rights reserved. - PublicationAccès libreUse of the frc gene as a molecular marker to characterize oxalate-oxidizing bacterial abundance and diversity structure in soil(2009)
; ; Oxalate catabolism, which can have both medical and environmental implications, is performed by phylogenetically diverse bacteria. The formyl-CoA-transferase gene was chosen as a molecular marker of the oxalotrophic function. Degenerated primers were deduced from an alignment of frc gene sequences available in databases. The specificity of primers was tested on a variety of frc-containing and frc-lacking bacteria. The frc-primers were then used to develop PCR-DGGE and real-time SybrGreen PCR assays in soils containing various amounts of oxalate. Some PCR products from pure cultures and from soil samples were cloned and sequenced. Data were used to generate a phylogenetic tree showing that environmental PCR products belonged to the target physiological group. The extent of diversity visualised on DGGE pattern was higher for soil samples containing carbonate resulting from oxalate catabolism. Moreover, the amount of frc gene copies in the investigated soils was detected in the range of 1.64 × 107 to 1.75 × 108/g of dry soil under oxalogenic tree (representing 0.5 to 1.2% of total 16S rRNA gene copies), whereas the number of frc gene copies in the reference soil was 6.4 × 106 (or 0.2% of 16S rRNA gene copies). This indicates that oxalotrophic bacteria are numerous and widespread in soils and that a relationship exists between the presence of the oxalogenic trees Milicia excelsa and Afzelia africana and the relative abundance of oxalotrophic guilds in the total bacterial communities. This is obviously related to the accomplishment of the oxalate–carbonate pathway, which explains the alkalinization and calcium carbonate accumulation occurring below these trees in an otherwise acidic soil. The molecular tools developed in this study will allow in-depth understanding of the functional implication of these bacteria on carbonate accumulation as a way of atmospheric CO2 sequestration. - PublicationAccès libreBiologically induced mineralization in the tree Milicia excelsa (Moraceae) : its causes and consequences to the environment(2004)
; ; Iroko trees (Milicia excelsa) in Ivory Coast and Cameroon are unusual because of their highly biomineralized tissues, which can virtually transform the trunk into stone. Oxalic acid (C2O4H2) and metal-oxalate play important roles in their ecosystems. In this study, the various forms of oxalate and carbonate mineralization reactions are investigated by using scanning electron microscopy and X-ray diffraction. Calcium oxalate monohydrate is associated with stem, bark and root tissues, whereas calcium oxalate dihydrate is found with wood rot fungi in soils, as well as in decaying wood. Laboratory cultures show that many soil bacteria are able to oxidize calcium oxalate rapidly, resulting in an increase in solution pH. In terms of M. excelsa, these transformations lead to the precipitation of calcium carbonate, not only within the wood tissue, but also within the litter and soil. We calculate that c. 500 kg of inorganic carbon is accumulated inside an 80-year-old tree, and c. 1000 kg is associated with its surrounding soil. Crucially, the fixation of atmospheric CO2 during tree photosynthesis, and its ultimate transformation into calcite, potentially represents a long-term carbon sink, because inorganic carbon has a longer residence time than organic carbon. Considering that calcium oxalate biosynthesis is widespread in the plant and fungal kingdoms, the biomineralization displayed by M. excelsa may be an extremely common phenomena. - PublicationAccès libreGeochemical influences on H40/1 bacteriophage inactivation in glaciofluvial sands(2004)
; - PublicationAccès libreAdvances in biological tracer techniques for hydrology and hydrogeology using bacteriophages: Optimization of the methods and investigation of the behavior of bacterial viruses in surface waters and in porous and fractured aquifers(1994)
; This work was undertaken in the Laboratoire de Microbiologie of the University of Neuchâtel, in close collaboration with the Center of Hydrogeology (CHYN). This multi-disciplinary research project was instigated because of the growing need both for environmentally harmless hydrogeological investigation tools, and for new tracers that can be used in multitracing experiments. This work treats of the use of bacteriophages as biological tracers in underground and surface waters. The size of bacteriophages is of the order of a few hundred nm. These viruses only attack specific bacteria. They are absolutely harmless for any other living organism. Previous experiments have shown the value of bacteriophages as tracers in fissured environments. The aims of the present work were: to select several bacteriophages particularly well adapted to the use as hydrogeological tracers ; to optimize the production of these bacteriophages, and the analytical methods for detecting them; to study the behavior of these bacteriophages experimentally, under different physico-chemical conditions; to test the phages in karstic fissured environments and to extend their use to saturated porous environments and surface hydrology; to compare the migration of these phages to that of the best conventional hydrological tracers. The first part of this work treats of the isolation and characterization of new bacteriophages. Over twenty bacteriophage/host bacterium (BHB) systems were studied. Seven bacteriophages were selected for the second part of the project. The selection was based on the physiological properties of the host bacteria and the physico-chemical characteristics of the bacteriophages. These seven phages exhibit an interesting variety of shapes and physical characteristics. They include three phages of strictly marine bacteria. None of these BHB systems are naturally found in aquifer waters. The rest of this work includes the development of an analytical and enumeration technique. We chose to work with the double agar-layer technique, using Petri dishes. The growth media and the different steps of this technique were optimized for each BHB system. This increased the reliability and reproducibility of the technique, without adding to the cost or workload. The behavior of the phages was studied in laboratory experiments. As this behavior is mostly determined by the inactivation and adsorption of the phages, we investigated the influence on these two phenomena of various parameters, either physical (temperature, pH, agitation), or chemical (ionic concentrations, presence of proteins, of sand or colloidal clay particles). Our results show that in water, phages react very rapidly and massively to the presence of colloidal particles, even to very low concentrations. Agitation causes the viruses to be rapidly inactivated. Raising the temperature increases this inactivation. Colloidal clay particles (Montmorillonite and Attapulgite) as well as organic macromolecules efficiently protect the phages from inactivation. Only one phage was inactivated faster in the presence of mineral colloids. Each phage reacted individually to the presence of these mineral colloids. However, no correlation could be established with the physico-chemical characteristics of the bacteriophages. The reactions can be classified into three distinct types, described by Grant et al. [1993]. According to the type of reaction of a phage, it is possible to qualitatively predict its behavior during a tracing experiment. The tracing experiments performed in karstic fissured environments showed that the migration of bacteriophages is similar to that of the best fluorescent tracers (Uranine, Sulphorhodamine). The restitutions are generally high. The phages always reappear faster, which shows that they can be diluted more, before reaching their inferior detection limit. The trials performed in a porous saturated environment, on the Wilerwald (CH) test site, showed that bacteriophages are also adapted to this type of conditions. Their speed of migration is higher than that of dissolved chemical compounds (Uranine, Naphtionate). Only one trial was performed in a river. It nevertheless showed that phages are also perfectly suitable to these types of tracings. By the simultaneous injection of several bacteriophages at different points along the river, it was possible to scan its course and discover an infiltration zone. This work confirms that phages are of undeniable value to hydrology as biological tracers. Their impact on the environment is practically nil and they can therefore be used also in delicate situations (springs used for drinking water, tracings in inhabited areas). This method is not limited to karstic aquifers anymore. Our modifications and adaptations of the technique make it suitable also for saturated porous environments and rivers. It advantages are numerous: The phages are non-pathogenic, non-toxic and invisible. So this method can also be used for studying drinking water (springs, reservoirs, etc). Each phage generally attacks only one bacterial species. By a careful selection of the BHB systems, any effect on the aquifers microflora can be avoided. No background noise exists in the aquifers, since they phages are not found there naturally. None will be generated or will persist over time, because of their short life span. The analysis of the samples is fast and cheap. Only a few milliliters of water are necessary for the enumeration of the phages. The detection level of the routine-analysis technique is about 1 phage per 2 ml of water. This sensitivity is comparable, and most often superior, to that of the best fluorescent tracers. If need be, the sensitivity can be lowered to 1 phage per 10 ml. It is also possible to differentiate and count a mixture of phages in a single sample. Theoretically, bacteriophages offer unlimited possibilities of multitracings. Ten to twenty liters of phage culture are necessary for one tracing experiment. Such an amount is easily transportable, even to inaccessible injection sites. It will contain 1014 - 1015 phages, which represents in all about 1 gram of protein and a few grams of mineral salts and of various organic substances (amino acids, growth medium components). The influence on the aquifer will be negligible, even if the flow is small. - PublicationAccès libreFungi, bacteria and soil pH: the oxalate–carbonate pathway as a model for metabolic interaction
; ; ; ; ; ; The oxalate–carbonate pathway involves the oxidation of calcium oxalate to low-magnesium calcite and represents a potential long-term terrestrial sink for atmospheric CO2. In this pathway, bacterial oxalate degradation is associated with a strong local alkalinization and subsequent carbonate precipitation. In order to test whether this process occurs in soil, the role of bacteria, fungi and calcium oxalate amendments was studied using microcosms. In a model system with sterile soil amended with laboratory cultures of oxalotrophic bacteria and fungi, the addition of calcium oxalate induced a distinct pH shift and led to the final precipitation of calcite. However, the simultaneous presence of bacteria and fungi was essential to drive this pH shift. Growth of both oxalotrophic bacteria and fungi was confirmed by qPCR on the frc (oxalotrophic bacteria) and 16S rRNA genes, and the quantification of ergosterol (active fungal biomass) respectively. The experiment was replicated in microcosms with non-sterilized soil. In this case, the bacterial and fungal contribution to oxalate degradation was evaluated by treatments with specific biocides (cycloheximide and bronopol). Results showed that the autochthonous microflora oxidized calcium oxalate and induced a significant soil alkalinization. Moreover, data confirmed the results from the model soil showing that bacteria are essentially responsible for the pH shift, but require the presence of fungi for their oxalotrophic activity. The combined results highlight that the interaction between bacteria and fungi is essential to drive metabolic processes in complex environments such as soil. - PublicationAccès libreExperimental calcium-oxalate crystal production and dissolution by selected wood-rot fungi
; ; ; Twenty-six species of white-rotting Agaricomycotina fungi (Basidiomycota) were screened for their ability to produce calcium-oxalate (CaOx) crystals in vitro. Most were able to produce CaOx crystals in malt agar medium in the absence of additional calcium. In the same medium enriched with Ca2+, all the species produced CaOx crystals (weddellite or whewellite). Hyphae of four species (Ganoderma lucidum, Polyporus ciliatus, Pycnoporus cinnabarinus, and Trametes versicolor) were found coated with crystals (weddellite/whewellite). The production of CaOx crystals during the growth phase was confirmed by an investigation of the production kinetics for six of the species considered in the initial screening (Pleurotus citrinopileatus, Pleurotus eryngii, Pleurotus ostreatus, P. cinnabarinus, Trametes suaveolens, and T. versicolor). However, the crystals produced during the growth phase disappeared from the medium over time in four of the six species (P. citrinopileatus, P. eryngii, P. cinnabarinus, and T. suaveolens). For P. cinnabarinus, the disappearance of the crystals was correlated with a decrease in the total oxalate concentration measured in the medium from 0.65 μg mm-2 (at the maximum accumulation rate) to 0.30 μg mm-2. The decrease in the CaOx concentration was correlated with a change in mycelia morphology. The oxalate dissolution capability of all the species was also tested in a medium containing calcium oxalate as the sole source of carbon (modified Schlegel medium). Three species (Agaricus blazei, Pleurotus tuberregium, and P. ciliatus) presented a dissolution halo around the growth zone. This study shows that CaOx crystal production is a widespread phenomenon in white-rot fungi, and that an excess of Ca2+ can enhance CaOx crystal production. In addition, it shows that some white-rot fungal species are capable of dissolving CaOx crystals after growth has ceased. These results highlight a diversity of responses around the production or dissolution of calcium oxalate in white-rot fungi and reveal an unexpected potential importance of fungi on the oxalate cycle in the environment.