Emile Benizri

Impact of growth stage on the bacterial community structure along maize roots, as determined by metabolic and genetic fingerprinting

Metabolic and genetic profiles were used to determine whether spatial and temporal variations in rhizodeposition along maize roots coincide with different bacterial community structures. Bacterial communities were extracted from the bulk soil and adhering soil of three maize rhizosphere zones (ramification, root hair-elongation, apex) 2 and 4 weeks after planting. Biolog® substrate utilization profiles, and rDNA internal spacer analysis of bacterial communities, were compared. Biolog® data showed that the functional abilities of bacterial communities from bulk and adhering soils were distinct after 2 weeks (Day 15). Moreover, these abilities were dissimilar between ramification zone on the one hand, and the root hair-elongation zone and apex on the other hand. The differences between bulk and rhizosphere soil responses were more pronounced after 4 weeks (Day 30), but rhizosphere samples were clearly aggregated. These results argue in favor of a greater influence of the maize rhizosphere environment on bacterial metabolic potentialities, mainly based on the developmental state of the plant. Different genetic fingerprints, and thus different genetic structures, were observed between bacterial communities at distinct sampling dates and, to a lesser extent, between rhizospheric and non-rhizospheric samples at both sampling dates. The latter difference was reinforced at Day 30. However, no clear groupings of samples could be identified on the basis of root-zone origin (ramification, root hair, apex). These results suggest a marked influence of time on microbial pools, irrespective of their root zone origins.

Metabolic fingerprint of microbial communities from distinct maize rhizosphere compartments

The metabolic abilities of microbial communities extracted from maize rhizosphere and non-rhizosphere compartments were compared by using the Biolog® system. A double sampling protocol (from bulk soil to root surface and along the root) was used for testing the hypothesis that nature and quantity of rhizodeposits could be key factors governing microbial community structure. Rhizoplane microbes are closer to rhizodeposition sites than microbes living in the adhering soil. This proximity generates distinct pools of rhizodeposits between adhering soil and rhizoplane; the carbon pool available for adhering soil microbes is qualitatively and quantitatively different from the original one because of rhizoplane microbial activity. The first sampling was designed to extract microbes from these two compartments. Moreover the different parts of a root are known to release distinct pools of rhizodeposits; the second sampling along the root was designed to extract microbes from the main morphological parts of a root (ramification zone, root hair-elongation zone and apex). Bulk soil without plant (control soil) was also investigated to obtain microbes deprived from rhizodeposits. The comparative metabolic profiling revealed clear shifts between the microbial populations of the rhizoplane and the control soil ones. The main discriminating substrates were carbohydrates, amino acids and amides. Microbial populations from the ramification zone and the control soil were clearly separated from those of the root hair-elongation zone. This discrimination was mainly based on some carboxylic acids, carbohydrates, amino acids and amides. The sampling procedure and the community structure differences revealed in this study confirm a possible structural effect of maize rhizodeposits on telluric microbial populations.

Effect of maize rhizodeposits on soil microbial community structure

In order to characterise the relationships between a plant and its rhizosphere microflora, we studied the effect of rhizodeposits released by roots on the phenotypic and genetic structures of soil bacterial populations. Sterile cultures of maize (cv. F66) were cultivated for 30 days in order to obtain sterile rhizodeposits. These rhizodeposits were added daily to a test soil during the last 15 days of maize cultivation. Afterwards, the structure of bacterial communities was analysed using a phenotypic test (Biolog®) and several dominant isolates were analysed by genetic test (ARDRA). Bacteria isolated from the soil watered only with plant nutrient solution used certain carboxylic acids more intensely than bacteria isolated from the soil conditioned with F66 rhizodeposits. Among the most representative ribotypes, Bacillus mycoides dominated in soil enriched with rhizodeposits, whereas Bacillus macroides was isolated from soil without carbon amendment.

Impact of auxin-compounds produced by the antagonistic fungus Pythium oligandrum or the minor pathogen Pythium group F on plant growth

Plant growth promotion induced by the antagonistic fungus, Pythium oligandrum, is the result of a complex interaction which includes an indirect effect through control of pathogens in the rhizosphere and/or a direct one mediated by plant-induced resistance. The present study shows an increased plant growth associated with direct interaction between P. oligandrum and roots, which is mediated by a fungus-produced auxin compound, tryptamine (TNH2). In vitro experiments provided evidence that P. oligandrum metabolised specifically indole derivatives, such as tryptophan and indole-3-acetaldehyde, to produce THN2 through the tryptamine pathway. When P. oligandrum grew in sterile root exudates, it also produced an auxin-like compound. Additional experiments on P. oligandrum–root interaction showed that, in amended nutrient solution of plants, the antagonist metabolised Trp into TNH2 and that root absorption of this newly formed auxin-compound in appropriate concentrations was associated with enhancement of plant growth. This phenomenon was observed only when nutrient solution was amended with low tryptophan (Trp) concentrations, i.e. 0.05 and 0.1 mM; higher concentration (0.5 and 1 mM Trp) induced abnormal root development. Similar experiments were performed with Pythium group F, a minor pathogen known for its ability to produce auxin-compounds through the tryptamine pathway. In this case, irregular root development was always noticed with all Trp concentrations added to the nutrient solution of plants. Moreover, Pythium group F colonization of roots was associated with leakage of auxin-compounds in the nutrient solution. Our results, therefore, highlight that the production of similar auxin-compounds by two Pythium species has contrary effects on plant development.

Soil microbial diversity as affected by the rhizosphere of the hyperaccumulator Thlaspi caerulescens under natural conditions.

It is hypothesized that metal hyperaccumulator plants have specific rhizosphere conditions, potentially modifying the bioavailability of soil metals. This article aims to further the knowledge about the rhizosphere of the hyperaccumulator Thlaspi caerulescens, focusing on its microflora isolated from metalliferous soils collected in situ where the plants grow naturally. We characterized the cultivable microbial communities isolated from the rhizosphere of one population of this Ni hyperaccumulator species grown on a serpentine soil. The rhizosphere soil harbored a wide variety of microorganisms, predominantly bacteria, confirming the stimulatory effect of the T. caerulescens rhizosphere on microbial growth and proliferation. We tested the hypothesis that the rhizosphere of T. caerulescens influences 1) the metabolic diversity of the bacterial community and 2) the bacterial resistance to metals. The principal component analysis of the Biolog® plates data confirmed a structural effect of the rhizosphere of T. caerulescens on bacterial communities. The percentage of Ni-resistant bacteria was higher in the rhizosphere than in the bulk soil, suggesting a direct effect of the rhizosphere on Ni tolerance, reflecting a greater bacterial tolerance to Ni in the rhizosphere.

Fate of two microorganisms in maize simulated rhizosphere under hydroponic and sterile conditions

Abstract The interaction, between a rhizobacterium ( Pseudomonas sp.) of maize and the agent of root rot of maize ( Fusarium graminearum ), was studied in vitro under hydroponic and sterile conditions. The Pseudomonas showed an antagonistic activity, in vitro , towards F. graminearum . The bacteria inhibited fungal growth, but the inhibition depended on the agar medium used. The antifungal activity of antagonistic bacteria was due to the production of antibiotics, volatile compounds and siderophores. The fate of the two microorganisms was investigated in a simulated maize rhizosphere. Under hydroponic and sterile conditions, both microorganisms grew well in the presence of root exudates used as nutrients. As a result of competition between both microorganisms for the C source, the presence of F. graminearum affected the survival of the bacterium and, conversely, the Pseudomonas sp. reduced germination of F. graminearum .

The effect of plant density in nickel-phytomining field experiments with Alyssum murale in Albania

Ultramafic vertisols cover large areas in Albania and offer opportunities for phytomining. We undertook a field experiment with native Alyssum murale on two representative Vertisols at a distance of 20 km from each other (Pojske and Domosdove, Albania), to test the effect of planting density (transplanted seedlings) on a phytomining cropping system. Both areas were cleared in late summer 2012 and then ploughed and the soils were characterised. At Domosdove, an area of 0.5 ha was planted with local native seedlings at a density of six plants per square metre in September 2012. Spontaneous plants that had germinated in Spring 2012 were left to grow without any competition from other plants on a second 0.1-ha plot at Domosdove. All plots were weeded manually in the autumn of 2012 and spring of 2013. Individual plants occupied ~1 m2 at maturity. At Pojske, an area of 0.3 ha was also planted in September 2012 with local native seedlings of A. murale at a density of four plants per square metre. Plants grown at initial densities of four and six plants per square metre did not fully cover the ground; gaps were filled in naturally by a second spontaneous generation of A. murale seedlings (recruits) that had germinated in Autumn 2012. Other weeds were eliminated with herbicides. At Domosdove, at densities of one and six plants and at Pojske of four plants per square metre, the biomass yield was 10, 5 and 10 t ha–1, respectively. Concentration of phytoextracted nickel was 77, 41 and 112 kg ha–1. We suggest that a density of four plants per square metre is suitable for phytoextraction with native populations of A. murale. A. murale can be a weed itself and lower the nickel phytoextraction yield. Plants responded differently in their native environment than in previous field trials in North America.

Start-up phase of an anaerobic full-scale farm reactor - Appearance of mesophilic anaerobic conditions and establishment of the methanogenic microbial community.

The goal of this study was to investigate how the microbial community structure establishes during the start-up phase of a full-scale farm anaerobic reactor inoculated with stale and cold cattle slurry. The 16S/18S high-throughput amplicon sequencing results showed an increase of the bacterial, archaeal and eukaryotic diversity, evenness and richness during the settlement of the mesophilic anaerobic conditions. When a steady performing digestion process was reached, the microbial diversity, evenness and richness decreased, indicating the establishment of a few dominant microbial populations, best adapted to biogas production. Interestingly, among the environmental parameters, the temperature, alkalinity, free-NH3, total solids and O2 content were found to be the main drivers of microbial dynamics. Interactions between eukaryotes, characterized by a high number of unknown organisms, and the bacterial and archaeal communities were also evidenced, suggesting that eukaryotes might play important roles in the anaerobic digestion process.

Compartmentalization and regulation of arylsulfatase activities in Streptomyces sp., Microbacterium sp. and Rhodococcus sp. soil isolates in response to inorganic sulfate limitation

Arylsulfatases allow microorganisms to satisfy their sulfur (S) requirements as inorganic sulfate after sulfate ester hydrolysis. Our objectives were to investigate the arylsulfatase activities among soil isolates, especially Streptomyces sp., Microbacterium sp. and Rhodococcus sp., because such investigations are limited for these bacteria, which often live in sulfate-limited conditions. Physiological and biochemical analyses indicated that these isolates possessed strong specific arylsulfatase activities ranging from 6 to 8 U. Moreover, for Streptomyces sp., an arylsulfatase localization study revealed 2 forms of arylsulfatases. A first form was located in the membrane, and a second form was located in the intracellular compartment. Both arylsulfatases had different patterns of induction. Indeed, the intracellular arylsulfatase was strictly induced by inorganic sulfate limitation, whereas the membrane arylsulfatase was induced both by substrate presence or S demand independently. For Microbacterium and Rhodococcus isolates, only a membrane arylsulfatase was found. Consequently, our results suggest the presence of a previously undescribed arylsulfatase in these microorganisms that allows them to develop an alternative strategy to fulfill their S requirements compared to bacteria previously studied in the literature.

External and Internal Root Colonization of Maize by Two Pseudomonas Strains: Enumeration by Enzyme-Linked Immunosorbent Assay (ELISA)

Abstract. Maize root colonization by two fluorescent Pseudomonas strains M.3.1. and TR335, isolated respectively from maize and tomato roots, were studied in hydroponic conditions. Each bacterium was inoculated separately, and three different colonization areas were studied: nutrient solution, rhizoplane, and endorhizosphere. The two Pseudomonas strains established large rhizosphere populations, and rhizoplane colonization of the entire root system was similar for both strains. However, strain M.3.1. colonized the endorhizosphere more efficiently than strain TR335. Seminal root cuttings from the tip to the seed allowed the assessment of colonization of three different root areas (i.e., root cap and elongation area, root-hair zone, and mature zone). Rhizoplane colonizations of all these three areas by M.3.1. were significantly the same, whereas strain TR335 colonized the rhizoplane of the root cap and elongation area more actively than the root-hair zone and mature zone. Population size of the strain M.3.1. in the internal tissue of these areas was greater than that of strain TR335. Co-inoculations of the two strains indicated a stimulation of the population size of strain M.3.1. regardless of root area studied, whereas population size of strain TR335 remained unchanged. These results demonstrated that external and internal maize root tissues were colonized to a greater extent by a strain isolated from a maize rhizosphere than by one isolated from another rhizosphere.