Increased degradation of phenanthrene in soil by Pseudomonas sp. GF3 in the presence of wheat

A phenanthrene-degrading bacterial strain Pseudomonas sp. GF3 was examined for plant-growth promoting effects and phenanthrene removal in soil artificially contaminated with low and high levels of phenanthrene (0, 100 and 200 mg kg⁻¹) in pot experiments. Low and high phenanthrene treatments significantly decreased the growth of wheat. Inoculation with bacterial strain Pseudomonas sp. GF3 was found to increase root and shoot growth of wheat. Strain GF3 was able to degrade phenanthrene effectively in the unplanted and planted soils. Over a period of 80 days the concentration of phenanthrene in soil in which wheat was grown was significantly lower than in unplanted soil (p<0.05). At the end of the 80-d experiments, 62.2% and 42.3% of phenanthrene had disappeared from planted soils without Pseudomonas sp. GF3 when the phenanthrene was added at 100 and 200 mg kg⁻¹ soil, respectively, but 84.8% and 70.2% of phenanthrene had disappeared from planted soils with the bacterial inoculation. The presence of vegetation significantly enhances the dissipation of phenanthrene in the soil. There was no significant difference in soil polyphenol oxidase activities among the applications of 0, 100 and 200 mg kg⁻¹ of phenanthrene. However, the enzyme activities in planted and unplanted soils inoculated with the strain Pseudomonas sp. GF3 were significantly higher than those of non-inoculation controls. The bacterial isolate was also able to colonize and develop in the rhizosphere soil of wheat after inoculation.     

Seed coating with inocula of arbuscular mycorrhizal fungi and plant growth promoting rhizobacteria for nutritional enhancement of maize under different fertilisation regimes

Arbuscular mycorrhizal (AM) fungi and plant growth-promoting rhizobacteria, responsible for enhancing plant nutrition, vigour and growth, may be used to reduce dosages of chemical fertilisers. Technologies that allow an economically viable and efficient application of these beneficial microbes in large scale agriculture must be studied. Seed coating is a potential delivery system for efficiently introducing minor amounts of bioinoculants. Despite the dramatic reduction on inoculum dose per plant, inoculation of AM fungi via seed coating was as effective as conventional soil inoculation. Fertilisation and inoculation had a significant impact on maize shoots nutrient concentrations. Different fertilisation regimes did not influence mycorrhizal colonisation. Plants without fertilisation and singly inoculated with R. irregularis showed shoot nutrient concentration increments of 110, 93, 88 and 175% for nitrogen, phosphorus, potassium and zinc, respectively, comparing with non-inoculated controls. Plants singly inoculated with P. fluorescens via seed coating under full fertilisation, presented enhancements of 100, 75 and 141% for magnesium, zinc and manganese, respectively, comparing with non-inoculated controls. Seed coating is a promising tool for delivering microbial inoculants into the soil, while promoting sustainable production of maize. This technology is particularly pertinent in low input agriculture, with potential environmental profits and food quality improvements.     

Suppression of root-knot disease by Pseudomonas fluorescens CHA0 in tomato: importance of bacterial secondary metabolite, 2,4-diacetylpholoroglucinol

The antimicrobial metabolites 2,4-diacetylphloroglucinol (2,4-DAPG) and pyoluteorin contribute to the ability of Pseudomonas fluorescens strain CHA0 to control plant diseases caused by soil-borne pathogens. P. fluorescens strain CHA0 and its derivatives CHA89 (antibiotics-deficient) and CHA0/pME3424 (antibiotics overproducing) were investigated as potential biocontrol agents against Meloidogyne javanica the root-knot nematode. Exposure of root-knot nematode to culture filtrates of P. fluorescens under in vitro conditions significantly reduced egg hatch and caused substantial mortality of M. javanica juveniles. Nutrient broth yeast extract (NBY) medium amended with 2% (w/v) glucose or 1 mM EDTA markedly repressed hatch inhibition activity of the strain CHA0 but not that of CHA0/pME3424 or CHA89. On the other hand, NBY medium amended with glucose significantly enhanced nematicidal activity of the strain CHA0/pME3424. Neither glucose nor EDTA had an influence on the nematicidal activity of the strains CHA0 and CHA89. Under in vitro conditions, antibiotic overproducing strain CHA0/pME3424 and CHA0 expressed phl‘-’lacZ reporter gene but strain CHA89 did not. Expression of the reporter gene reflects actual production of DAPG. In general, CHA0/pME3424 expressed reporter gene to a greater extent compared to its wild type counterpart CHA0. Regardless of the bacterial strains, reporter gene expression was markedly enhanced when NBY medium was amended with glucose but EDTA had no such effect. A positive correlation between the degree of juvenile mortality and extent of phl‘-’lacZ reporter gene expression was also observed in vitro. Strain CHA0 produced zones of 4-6 mm on MM medium containing gelatin while strain CHA0/pME3424 and CHA89 did not. When MM medium containing gelatin was amended with 2% glucose of 1 mM EDTA size of haloes produced by the strain CHA0 reduced to 2 mm. Under glasshouse conditions aqueous cell suspension of the strains CHA0 or CHA0/pME3424 at various inoculum levels (10⁷, 10⁸ or 10⁹ cfu ml⁻¹) significantly reduced root-knot development. CHA89 caused significant reduction in galling when applied at 10⁹ cfu ml⁻¹. To better understand the mechanism of nematode suppression, split root bioassay was performed. Split-root experiments, that guarantee a spatial separation of inducing agent and a challenging pathogen, showed that soil treatment of one half of the root system with cell suspension of CHA0 or CHA0/pME3424 resulted in a significant systemic induced resistance leading to reduction of M. javanica infection of tomato roots in the non-baterized nematode treated half. The results clearly suggest that the antibiotic 2,4-DAPG from P. fluorescens CHA0 act as the inducing agents of systemic resistance in tomato roots. Populations of CHA0 and its derivatives declined progressively by 10-fold between first and fourth harvests (0-21 days after inoculation). However, bacterial populations increased at final harvest (28 days after application).     

The abundance and efficacy of Rhizobium leguminosarum bv. viciae in cultivated soils of the eastern Canadian prairie

For optimum production, the use of commercial rhizobial inoculant on pea (Pisum sativum L.) at seeding is necessary in the absence of compatible rhizobial strains or when rhizobial soil populations are low or symbiotically ineffective. Multiple site experiments were conducted to characterize the abundance and effectiveness of resident populations of Rhizobium leguminosarum bv. viciae (Rlv) in eastern Canadian prairie soils. A survey of 20 sites across a broad geographical range of southern Manitoba was carried out in 1998 and was followed by more intensive study of five of the sites in 1999 and 2000. Appreciable nodulation of uninoculated pea was observed at all sites which had previously grown inoculated pea. However, uninoculated pea grown at two sites, which had not previously grown pea, had negligible nodulation. Likewise, wild Lathyrus sp. and Vicia sp. plants collected from uncultivated areas adjacent to agricultural sites were poorly nodulated. In the more intensively studied sites, there was a tendency towards higher nodulation in pea plants receiving commercial inoculant containing Rlv strain PBC108 across all site-years (e.g., 4.7% in nodulation and 22% in nodule mass), but the effect was significant at only 2 of 10 site-years. Despite a relatively high range of soil pH (6-8), regression analysis indicated that decreasing soil pH resulted in lower nodulation rates. Likewise, electrical conductivity (EC) was correlated to nodulation levels, however the effect of EC was likely more indicative of the influence of soil texture and organic matter than salinity. As with nodulation, commercial inoculation tended to increase above-ground dry matter (DM) and fixed-N (estimated by the difference method) at the early pod-filling stage, but again the effects were significant at only 2 of 10 site-years. Specifically, above-ground DM and fixed-N levels were up to 29 and 51% greater, respectively, in inoculated compared to non-inoculated treatments at these sites. Addition of N-fertilizer at a rate of 100 kg N ha⁻¹ decreased nodulation at almost all site-years (by as much as 70% at one site), but rarely resulted in increases in above-ground DM compared to inoculated plots. The study indicates for the first time that populations of infective, and generally effective strains of Rlv occur broadly in agricultural soils across the eastern Canadian prairie, but that there is a tendency for increased symbiotic efficiency with the use of commercial inoculant.     

Nickel-tolerant Brevibacillus brevis and arbuscular mycorrhizal fungus can reduce metal acquisition and nickel toxicity effects in plant growing in nickel supplemented soil

The growth of clover (Trifolium repens ) and its uptake of N, P and Ni were studied following inoculation of soil with Rhizobium trifolii, and combinations of two Ni-adapted indigenous bacterial isolates (one of them was Brevibacillus brevis) and an arbuscular mycorrhizal (AM) fungus (Glomus mosseae). Plant growth was measured in a pot experiment containing soil spiked with 30 (Ni I), 90 (Ni II) or 270 (Ni III) mg kg⁻¹ Ni-sulphate (corresponding to 11.7, 27.6 and 65.8 mg kg⁻¹ available Ni on a dry soil basis). Single inoculation with the most Ni-tolerant bacterial isolate (Brevibacillus brevis) was particularly effective in increasing shoot and root biomass at the three levels of Ni contamination in comparison with the other indigenous bacterial inoculated or control plants. Single colonisation of G. mosseae enhanced by 3 fold (Ni I), by 2.4 fold (Ni II) and by 2.2 fold (Ni III) T. repens dry weight and P-content of the shoots increased by 9.8 fold (Ni I), by 9.9 fold (Ni II) and by 5.1 fold (Ni III) concomitantly with a reduction in Ni concentration in the shoot compared with non-treated plants. Coinoculation of G. mosseae and the Ni-tolerant bacterial strain (B. brevis) achieved the highest plant dry biomass (shoot and root) and N and P content and the lowest Ni shoot concentration. Dual inoculation with the most Ni-tolerant autochthonous microorganisms (B. brevis and G. mosseae) increased shoot and root plant biomass and subtantially reduced the specific absorption rate (defined as the amount of metal absorbed per unit of root biomass) for nickel in comparison with plants grown in soil inoculated only with G. mosseae. B. brevis increased nodule number that was highly depressed in Ni I added soil or supressed in Ni II and Ni III supplemented soil. These results suggest that selected bacterial inoculation improved the mycorrhizal benefit in nutrients uptake and in decreasing Ni toxicity. Inoculation of adapted beneficial microorganisms (as autochthonous B. brevis and G. mosseae) may be used as a tool to enhance plant performance in soil contaminated with Ni.     

Biodegradation of benzoate with nitrate as electron acceptor at different redox potentials in sand column microcosms

 The purpose of this work was to assess the influence of redox conditions on benzoate biodegradation coupled with oxygen and nitrate as electron acceptors. A benzoate-degrading, facultatively denitrifying bacterium was isolated from a sediment sample and was tentatively identified as Comamonas terrigena (strain J92-6). The experimental system was based on sand columns that were filled with liquid medium containing benzoate and nitrate. The columns were inoculated to provide a fixed biofilm on the sand. Conditions were created by aeration, dinitrogen-purging, and sodium sulfide amendment that comprised oxic, anoxic, and reduced (–375 mV) zones, respectively, at different depths of the columns. Anaerobic biodegradation of benzoate was nitrate-dependent and proceeded at all redox potential values ranging from +400 to –375 mV. Thus, benzoate degradation coupled with denitrification was not inhibited at low redox potentials characteristic of sulfate reduction and methanogenesis. The results demonstrate that the fixed biofilm column system can be successfully used to evaluate the influence of environmental factors on the biodegradation of benzoate, a central decomposition product of anaerobic, aromatic hydrocarbon biodegradation. Received: 16 April 1997     

Fate, tree growth effect and potential impact on soil microbial communities of mycorrhizal and bacterial inoculation in a forest plantation

The knowledge of the survival of inoculated beneficial fungal and bacterial strains in the field and the effects of their release on the indigenous microbial communities has been of great interest since the practical use of selected natural or genetically modified microorganisms has been developing. The aim of this study was to monitor, 4 years after plantation into the field site, the effects of Douglas fir (Pseudotsuga menziesii) co-inoculation with the mycorrhiza helper bacterial strain Pseudomonas fluorescens BBc6R8 and/or the fungal strain Laccaria bicolor S238N on seedling growth and on the indigenous bacterial and ectomycorrhizal communities using quantitative and qualitative approaches. The field persistence of the inoculated strains was also monitored. The seedling shoot volume estimate was statistically significantly higher in the fungal inoculated plots in comparison to the non-inoculated plots but no treatment-related changes in the quantitave or qualitative microbial measurements were observed and the inoculated strains could not be detected after 4 years.     

Soil texture and irrigation influence the transport and the development of Pasteuria penetrans, a bacterial parasite of root-knot nematodes

The transport of the spores of Pasteuria penetrans was studied in three contrasted textured soils (a sandy, a sandy-clay and a clay soils), cultivated with tomato, inoculated with juveniles of Meloidogyne javanica and watered with 25 or 150 mm day⁻¹. One month after inoculation of the nematodes, 53% of the spores inoculated were leached by water flow in the sandy soil but only 14% in the sandy-clay soil and 0.1% in the clay soil. No nematodes survived in the clay soil, while the population was multiplied both in the sandy and in the sandy-clay soils. But juveniles of M. javanica were more infected by P. penetrans in the sandy-clay soil than in the sandy soil. Comparing different combinations of bare soils containing 1.1-57% of clay showed that the best spore percolation and retention balance occurred in soils amended with 10-30% clay. However, the spore recoveries decreased when the soil was enriched with more than 30% clay. The role of clay particles on the extractability of spores and on their availability to attach to the nematode cuticle in the soil is discussed.     

Nematode community structure as a sensitive indicator of microbial perturbations induced by a genetically modified Pseudomonas fluorescens strain

The effects of seed inoculation with the Pseudomonas fluorescens strains F113lacZY [a genetically marked biocontrol agent producing the anti-fungal agent 2,4-diacetylphloroglucinol (DAPG)] and F113G22 [a genetically modified (GM) derivative strain of F113lacZY incapable of producing DAPG] on associated nematode communities were investigated over 17 days of plant growth. Plant growth measurements and colony forming unit counts (CFU) derived from rhizosphere soil indicated only small and transient perturbations as a result of introductions of the GM bacteria. Total nematode numbers were increased significantly in the rhizosphere of inoculated plants compared with the non-inoculated control treatments. These increases were mainly due to increases in bacterial feeding nematodes. This indicates that inoculation with the GM P. fluorescens strains induced high bacterial growth rates in the rhizosphere of plants inoculated with these strains. No indication of greater root colonisation by fluorescent Pseudomonas spp. could be found using CFU counts on Pseudomonas-selective media. Numbers of fungal feeding nematodes decreased initially, probably as a result of lack of intact hyphae in the soil. However, inoculation with the two different GM P. fluorescens strains resulted in a rapid recovery of fungal feeding nematode populations, whereas in the non-inoculated control populations of fungal feeding nematodes remained small. This result is surprising as one of the strains (F113lacZY) produces the anti-fungal agent DAPG and it would be expected that this agent would result in a decrease in fungal activity.     

Microbial responses to fluctuation of soil aeration status and redox conditions

 The response of the microbial community to changes in aeration status, from oxic to anoxic and from anoxic to oxic, was determined in arable soil incubated in a continuous flow incubation apparatus. Soil incubated in permanently oxic (air) and/or anoxic (O₂-free N₂) conditions was used as the control. Before experiments soil was preincubated for 6 days, then aeration status was changed and glucose added. Glucose concentration, extractable C, CO₂ production, microbial biomass, pH and redox potential were determined 0, 4, 8, 12, 16, 24, 36 and 48 h after change of aeration status. If oxic conditions were changed to anoxic, the amount of glucose consumed was reduced by about 60%, and CO₂ production was 10 times lower at the end of incubation compared to the control (permanently oxic conditions). Microbial biomass increased by 114% in glucose-amended soil but did not change in unamended soil. C immobilization prevailed over C mineralization. Redox potential decreased from +627 mV to –306 mV. If anoxic conditions were changed to oxic, consumption of glucose and CO₂ evolution significantly increased, compared to permanently anoxic conditions. Microbial biomass did not change in glucose-amended soil, but decreased by 78% in unamended soil. C mineralization was accelerated. Redox potential increased from +238 to +541 mV. The rate of glucose consumption was low in anoxic conditions if soil was incubated in pure N₂ but increased significantly when incubation was carried out in a CO₂/N₂ mixture. Received: 6 January 1999     

Use of electronic nose technology to measure soil microbial activity through biogenic volatile organic compounds and gases release

Gas and volatile organic compounds (VOCs) release in soil is known to be linked to microbial activity and can differently affect the life of organisms in soil. Electronic noses (E-noses) are sensing devices composed of sensor arrays able to measure and monitor gases and VOCs in air. This is the first report on the use of such a sensing device to measure specifically microbial activity in soil. In the present study, γ-irradiated sterilised soil was inoculated with Pseudomonas fluorescens. To be sure for a rapid microbial growth and activity, two pulses of nutrient solution with organic and inorganic C, N, P and S sources were added to soil and the resulting microcosms were incubated for 23 d. During the incubation, respiration and enzyme activities of acid phosphatase, β-glucosidase, fluorescein diacetate hydrolase and protease, were measured, and microbial growth as global biomass of vital cells based on substrate-induced respiration (SIR-C_mic) and enumeration of viable and culturable cells by means of dilution plate counts (CFU) were also monitored. Concurrently, VOCs and/or gas evolution in the headspace of the soil microcosms were measured through the E-nose, upon their adsorption on quartz crystal microbalances (QCMs) comprising the sensory device. The E-nose typically generated an odorant image (olfactory fingerprint) representative of the analysed samples (soils) and resulting from the concurrent perception of all or most of the analytes in headspace, as it commonly happens when several selective but not specific sensors are used together (array). The basic hypothesis of this study was that different soil ecosystems expressing distinct microbial metabolic activities, tested through respiration and enzyme activities, might generate different olfactory fingerprints in headspace. Furthermore, the possibility to detect several substances at the same time, released from the soil ecosystems, possibly deriving from both abiotic and biotic (microbial metabolism) processes provides an “odorant image” representative of the whole ecosystem under study. The E-nose here used succeeded in discriminating between inoculated and non-inoculated ecosystems and in distinguishing different metabolic and growth phases of the inoculated bacteria during incubation. Specifically, E-nose responses were proved highly and significantly correlated with all hydrolytic activities linked to the mobilisation of nutrients from soil organic matter and their cycling, with CO₂ fluxes (respiration and presumed heterotrophic fixation) and with P. fluorescens population dynamics during exponential, stationary and starvation phases measured by SIR-C_mic and CFUs. Interestingly, the E-nose successfully detected soil microbial activity stimulated by nutrient supply, even though none of the catalytic activities tested directly produced VOCs and/or gases. The E-nose technology was then proved able to supply a real holistic image of microbial activity in the entire gnotobiotic and axenic soil ecosystems.     

Quantification of the biocontrol agent Pseudomonas fluorescens Pf153 in soil using a quantitative competitive PCR assay unaffected by variability in cell lysis- and DNA-extraction efficiency

Although often neglected, variability in cell lysis efficiency and DNA extraction yield represents the major hurdles of any polymerase chain reaction (PCR)-based quantification protocol in soil and other natural environments. In this study we developed a technique that minimizes the effects of these constraints, providing at the same time a reliable internal control to distinguish between PCR-inhibition and negative results. We used Pseudomonas fluorescens Pf153, a root-colonizing bacterium that shows biocontrol activity against tobacco and cucumber black root rot, as the target organism for PCR quantification. Prior to DNA extraction, the genetically engineered, cognate reference strain P. fluorescens CHA0/c2 was inoculated in a reference soil. CHA0/c2 in the reference soil and Pf153 in the soil sample were lysed in parallel and afterward the lysates were mixed in known proportions. CHA0/c2 carries the plasmid pME6031-cmp2 that contains an allelic variant (competitor) of the Pf153 specific sequence Pf153_2. In a quantitative competitive PCR (QC-PCR) assay the competitor allows the quantification of the target strain down to 0.66 Pf153 CFU/mg soil. Processing the reference strain in the same way as Pf153 enables the exact quantification of the target strain in biocontrol assays performed in natural soil, overcoming differences in DNA extraction efficiency and PCR amplification from different soil environments. This technique is easily adaptable to other Pseudomonas strains simply by replacing the competitor used here with one derived from a SCAR-marker which is specific for the strain of choice.     

Soil organic N dynamics and stand quality in Pinus radiata pinewoods of the temperate humid region

Soil organic-N dynamics, its controlling factors and its relationships with stand quality were studied in the 0-15 cm soil layer of 24 pinewoods with contrasting age, productivity and parent material (granite; acid schists), searching for N variables useful to predict stand growth and site quality. No significant differences were found between young and old stands for any of the N variables considered, nor two- or three-order interactions among stand age, site quality and parent material. The soil total-N content, which was correlated positively with the Al oxides content (a soil organic matter (SOM) stabilizing agent), did not vary significantly according to parent material, but it was lower (P≤0.02) in stands with high than with low site index (2.68±1.11 and 3.97±1.13 g N kg⁻¹ soil, respectively). The soil δ¹⁵N ranged from +3.5 to +6.5 δ, without significant differences among stand groups, and it was negatively correlated with water holding capacity, exchangeable bases, Al oxides and N content, suggesting that: (i) N losses by NO₃⁻ leaching are the most important controlling factor of δ¹⁵N in these temperate humid region soils; and (ii) soil N richness is related with limited N losses, which discriminate against ¹⁵N. At any incubation time, no significant differences were found in soil inorganic-N content among stand groups (7.78±4.57, 39.33±16.20 and 67.80±26.50 mg N kg⁻¹ soil at 0, 42 and 84 d, respectively). During the incubation, the relative importance of ammonification decreased and that of the nitrification increased. The net N mineralization rate (NNMR, in percentage of organic N) was significantly higher in granite than in schists soils at both 42 d (1.24±0.34 and 0.75±0.37%, respectively) and 84 d (2.18±0.56 and 1.53±0.66%, respectively). In high quality pinewoods, the NNMR at 42 and 84 d (1.16±0.45 and 2.12±0.79%, respectively) were significantly higher than in low quality stands (0.83±0.35 and 1.59±0.45%, respectively). This result, together with those on soil total-N and inorganic-N supply, suggests that soil N dynamics in low and high quality stands are different: in the former there is a bigger N pool with a slower turnover, whereas in the latter there is a smaller N pool with a faster turnover, both factors being nearly compensated, making the soil available N supply in both types of stand similar. After 42 and 84 d of incubation, the NNMR and the nitrification rates were higher in the coarse textured soils, likely due to the low physical and chemical protection of their SOM; both rates were positively correlated with available P, exchangeable K⁺ and CEC base saturation, suggesting strong relationships among the availabilities of the main plant nutrients, and they increased with SOM quality (low C-to-N ratio). The strong negative correlation of site index with soil total-N (r=−0.707; P≤0.005), and its positive correlations with NNMR after 42 and 84 d of incubation, suggested that site quality and potential productivity are closely related to soil organic-N dynamics. Half of the site index variation in the stands studied could be predicted with a cheap and easy analysis of soil N content, the prediction being slightly improved if soil δ¹⁵N is included and, more significantly, by including N mineralization measurements.     

Hydrolysis of cellobiose by β-glucosidase in the presence of soil minerals - Interactions at solid-liquid interfaces and effects on enzyme activity levels

Extracellular enzymatic activities in soils are essential for the cycling of organic matter. These activities take place in multiphase environments where solid phases profoundly affect biocatalytic activities. Aspergillus niger is ubiquitous in soils; its β-glucosidase plays an important role in the degradation of cellulose, and therefore in the global carbon cycle and in the turnover of soil organic matter. However, the information on the interactions of this protein with soil minerals is very limited, and even less is known about their consequences for the hydrolysis of the natural substrate cellobiose. We therefore characterised the sorptive interactions of this enzyme with the soil minerals montmorillonite, kaolinite and goethite and quantified the resulting changes in the hydrolysis rate of cellobiose. Fractions of adsorbed protein, and the resulting catalytic activity loss, were lower for montmorillonite than for kaolinite and goethite at given experimental conditions; adsorption was 9.7 ± 7.3% for montmorillonite, 70.3 ± 3.1% for kaolinite and 71.4 ± 1.8% for goethite, respectively. Adsorption of the protein to the minerals caused a total decrease in the catalytic activity of 18.8 ± 3.4% for kaolinite and 17.9 ± 4.7% for goethite whereas it was not significant for montmorillonite. The average catalytic activity lost by the pool of adsorbed molecules was 26.8% for kaolinite and 25.0% for goethite. Both the amount of adsorbed protein and the resulting loss of catalytic activity were found to be independent of the specific surface areas yet were influenced by the electrical properties of the mineral surfaces. Under the experimental conditions, montmorillonite and kaolinite are negatively charged whereas goethite is positively charged. However, because of the adsorption of phosphate anions from the buffer, a charge reversal took place at the surface of goethite. This was confirmed by zeta (ζ)-potential measurements in phosphate buffer, revealing negative values for all the tested minerals. Indeed goethite interacted with the enzyme as a negatively charged surface: the amount of adsorbed protein and the resulting catalytic activity loss were very similar to those of kaolinite. Our results show that, even if an important fraction of β-glucosidase is adsorbed to the minerals, the catalytic activity is largely retained. We suggest that this strong activity retention in presence of soil minerals results from a selective pressure on A. niger, which benefits from the activity of the adsorbed, and thus stabilized, enzyme pool.     

Fraxinus-Glomus-Pisolithus symbiosis: Plant growth and soil aggregation effects

It has been established that arbuscular mycorrhizal (AM) fungi are involved in the conservation of soil structure. However, the effect of ectomycorrhizal (EM) fungi alone or in interaction with AM fungi in soil structure has been much less studied. This experiment evaluated EM and AM fungi effects on soil aggregation and plant growth. Ash plants (Fraxinus uhdei) were grown in pots, and were inoculated with Glomus intraradices and Pisolithus tinctorius separately but also in combination. Our results showed that F. uhdei established a symbiotic association with EM and AM fungi, and that these organisms, when interacting, showed synergistic and additive effects on plant growth compared to singly inoculated treatments. EM and AM fungi prompted changes in root morphology and increased water-stable aggregates. AM fungi affect mainly small-sized macroaggregates, while EM and EM-AM fungi interaction mainly affected aggregates bigger than 0.5 mm diameter. These results suggest that ectomyccorrhizal as well as arbuscular mycorrhizal fungi should be considered in restoration programs with Fraxinus plants.     

Performance of Pseudomonas spp. containing ACC-deaminase for improving growth and yield of maize (Zea mays L.) in the presence of nitrogenous fertilizer

Some plant-growth-promoting rhizobacteria (PGPR) promote plant growth by lowering the endogenous ethylene synthesis in the roots through their 1-aminocylopropane-1-carboxylate (ACC)-deaminase activity. However, in the vicinity of the roots may decrease the efficiency of these PGPR by stimulating ACC-oxidase activity resulting in greater ethylene production by the roots. This study was designed to assess the performance of PGPR containing ACC-deaminase for improving growth and yield of maize grown in N-amended soil. Several strains of rhizobacteria containing ACC-deaminase were screened for their growth-promoting activity in maize roots under gnotobiotic conditions. Six strains were selected and their effectiveness in soil amended with N at a concentration of 175 kg ha⁻¹ (1050 mg pot⁻¹) was investigated by conducting a pot trial on maize. Significant increases in plant height, root weight and total biomass were observed in response to inoculation. Based upon the results of pot trials, the three most efficient strains were selected and tested in the field for their effectiveness in the presence and absence of N fertilizer. Results of the field trial revealed that the inoculum performed relatively better in the absence of N-fertilizer application. Pseudomonas fluorescens biotype G (N₃) was the most effective strain both in the presence and absence of N fertilizer. Results may imply that even in the presence of optimum levels of nitrogenous fertilizers, inoculation with rhizobacteria containing ACC-deaminase activity could be effective to improve the growth and yield of inoculated plants.     

Mycorrhizal influence on storage metabolites and mineral nutrition in seed propagated potato (Solanum tuberosum L.) plant

Abstract

Potato (Solanum tuberosum) showed a well-established mutualistic association with arbuscular mycorrhizal fungi. In the present study, earthen pots containing autoclaved soil were taken in which surface sterilized potato seeds were sown. The seed sowed earthen pots were inoculated by sterilized spores of arbuscular mycorrhizal fungi along with sterilized inoculated maize-root fragments, while the rest half of the seed filled pots, without any inoculation depicting control but were provided with non-inoculated maize root fragments. The inoculation was performed twice; first inoculation was done 3 days prior to sowing of potato seeds and second at the time of seedling emergence. Sampling of the inoculated as well as of control plants was performed at 20-day intervals till 80?days after tuber initiation. An increment in the level of metabolites as well as mineral nutrient was found in mycorrhizal inoculated potato tubers in contrast to non-inoculated. Our work demonstrates that inoculation has a great potential in enriching storage metabolites and nutrients in potato plant in low yielding soils.     

Increase of secondary metabolite content in marigold by inoculation with plant growth-promoting rhizobacteria

The effects of single inoculation and co-inoculation of two plant growth-promoting rhizobacteria (PGPR) (Pseudomonas fluorescens, Azospirillum brasilense) on growth and essential oil (EO) composition and phenolic content were evaluated in marigold (Tagetes minuta). Plant growth parameters (shoot fresh weight, root dry weight, leaf number, node number) were measured. EO yield increased 70% in P. fluorescens-inoculated and co-inoculated plants in comparison with control (non-inoculated) plants, without altering EO composition. The biosynthesis of the major EO components was increased in the inoculated plants. Shoot fresh weight and EO yield were significantly higher in P. fluorescens-inoculated and in co-inoculated plants than in control plants. The total phenolic content was 2-fold higher in singly-inoculated or co-inoculated treatments than in controls. In view of the economic importance of monoterpenes and phenolic compounds for a variety of applications in the food and cosmetic industries, P. fluorescens and other PGPR have clear potential for improving the productivity of cultivated aromatic plants. Better understanding of the processes that affect the accumulation of secondary metabolites will lead to increased yields of these commercially valuable natural products.     

The mycorrhizal fungus Glomus intraradices and rock phosphate amendment influence plant growth and microbial activity in the rhizosphere of Acacia holosericea

Plants inoculated with arbuscular mycorrhizal (AM) fungi utilize more soluble phosphorus from soil mineral phosphate than non-inoculated plants. However, there is no information on the response of soil microflora to mineral phosphate weathering by AM fungi and, in particular, on the catabolic diversity of soil microbial communities.The AM fungus, Glomus intraradices was examined for (i) its effect on the growth of Acacia holosericea, (ii) plant-available phosphate and (iii) soil microbial activity with and without added rock phosphate.After 4-months culture, AM fungal inoculation significantly increased the plant biomasses (by 1.78× and 2.23× for shoot and root biomasses, respectively), while mineral phosphate amendment had no effect in a sterilized soil. After 12-months culture, the biomasses of A. holosericea plants growing in a non-sterilized soil amended with mineral phosphate were significantly higher than those recorded in the control treatment (by 2.5× and 5× for shoot and root biomasses, respectively). The fungal inoculation also significantly stimulated plant growth, which was significantly higher than that measured in the mineral phosphate treatment. When G. intraradices and mineral phosphate were added together to the soil, shoot growth were significantly stimulated over the single treatments (inoculation or amendment) (1.45×). The P leaf mineral content was also higher in the G. intraradices+mineral phosphate treatment than in G. intraradices or rock phosphate amendment. Moreover, the number of fluorescent pseudomonads has been significantly increased when G. intraradices and/or mineral phosphate were added to the soil. By using a specific type of multivariate analysis (co-inertia analysis), it has been shown that plant growth was positively correlated to the metabolization of ketoglutaric acid, and negatively linked to the metabolisation of phenylalanine and other substrates, which shows that microbial activity is also affected.G. intraradices inoculation is highly beneficial to the growth of A. holosericea plants in controlled conditions. This AM symbiosis optimises the P solubilization from the mineral phosphate and affects microbial activity in the hyphosphere of A. holosericea plants.     

Hydrological controls on soil redox dynamics in a peat-based,restored wetland

Increasing areas of altered wetland are being restored by re-flooding the soil. Evidence in the literature indicates that this practice can induce the redox-mediated release of soil nutrients, thereby increasing the risk of diffuse water pollution. However, for the sake of improving wetland management decisions, there is a need for more detailed studies of the underlying relationship between the hydrological and redox dynamics that explain this risk; this is particularly the case in agricultural peatlands that are commonly targeted for the creation of lowland wet grassland. A 12-month field study was conducted to evaluate the relationship between hydrological fluctuations and soil redox potential (Eh) in a nutrient-rich peat field (32 g N kg^− 1 and 1100 mg P kg^− 1 in the surface 0–30 cm soil) that had been restored as lowland wet grassland from intensive arable production. Field tensiometers were installed at the 30-, 60- and 90-cm soil depths, and Pt electrodes at the 10-, 30-, 60- and 90-cm depths, for daily logging of soil water tension and Eh, respectively. The values for soil water tension displayed a strong negative relationship (P < 0.001) with monthly dip well observations of water table height. Calculations of soil water potential from the logged tension values were used, therefore, to provide a detailed profile of field water level and, together with precipitation data, explained some of the variation in Eh. For example, during the summer, alternating periods of aerobism (Eh > 330 mV) in the surface, 0–10 cm layer of peat coincided with intense precipitation events. Redox potential throughout the 30–100 cm profile also fluctuated seasonally; indeed, at all depths Eh displayed a strong, negative relationship (P < 0.001) with water table height over the 12-month study period. However, Eh throughout the 30–100 cm profile remained relatively low (< 230 mV), indicating permanently reduced conditions that are associated with denitrification and reductive dissolution of Fe-bound P. The implications of these processes in the N- and P-rich peat for wetland plant diversity and water quality are discussed.