In this study, we provide experimental evidences that in calcareous soils microbial degradation/decomposition of citrate can promote Al-(hydr)oxide precipitation concurrently decreasing copper (Cu) solubility by a coprecipitation process. Citrate is an organic acid anion commonly released by roots to increase nutrient availability or to limit Al toxicity. However, under specific environmental conditions (i.e. high microbial activity of Al-citrate-degrading bacteria, alkaline pH), this organic acid may become ineffective in mobilizing Cu for the plant acquisition process. To demonstrate this, a calcareous soil and an artificial soil system have been treated with citrate solutions; then, changes in Al and Cu solubility and the formation of Cu-containing Al-(hydr)oxides were monitored. Both in experiments with the artificial soil and in those where the soil was inoculated with microbial strains, the formation of Cu-Al coprecipitates not only occurred but was also concurrent with the decrease of Cu and Al solubility. The role of bacteria in metal-citrate complex degradation has been assessed, and the 16S rDNA of bacteria related with these processes has been sequenced for genus identification. Bacteria belonging to Pseudomonas, Sphingomonas, Bradyrhizobium and Sphingopixis have been identified as possible candidates to degrade Al- and Cu-citrate complexes thus triggering the metal precipitation phenomena.

Background: Epidemiology of celiac disease (CD) is increasing. CD mainly presents in early childhood with small intestinal villous atrophy and signs of malabsorption. Compared to healthy individuals, CD patients seemed to be characterized by higher numbers of Gram-negative bacteria and lower numbers Gram-positive bacteria. Results: This study aimed at investigating the microbiota and metabolome of 19 celiac disease children under gluten-free diet (treated celiac disease, T-CD) and 15 non-celiac children (HC). PCR-denaturing gradient gel electrophoresis (DGGE) analyses by universal and group-specific primers were carried out in duodenal biopsies and faecal samples. Based on the number of PCR-DGGE bands, the diversity of Eubacteria was the higher in duodenal biopsies of T-CD than HC children. Bifidobacteria were only found in faecal samples. With a few exceptions, PCRDGGE profiles of faecal samples for Lactobacillus and Bifidobacteria differed between T-CD and HC. As shown by culture-dependent methods, the levels of Lactobacillus, Enterococcus and Bifidobacteria were confirmed to be significantly higher (P = 0.028; P = 0.019; and P = 0.023, respectively) in fecal samples of HC than in T-CD children. On the contrary, cell counts (CFU/ml) of presumptive Bacteroides, Staphylococcus, Salmonella, Shighella and Klebsiella were significantly higher (P = 0.014) in T-CD compared to HC children. Enterococcus faecium and Lactobacillus plantarum were the species most diffusely identified. This latter species was also found in all duodenal biopsies of T-CD and HC children. Other bacterial species were identified only in T-CD or HC faecal samples. As shown by Randomly Amplified Polymorphic DNA-PCR analysis, the percentage of strains identified as lactobacilli significantly (P = 0.011) differed between T-CD (ca. 26.5%) and HC (ca. 34.6%) groups. The metabolome of T-CD and HC children was studied using faecal and urine samples which were analyzed by gas-chromatography mass spectrometry-solid-phase microextraction and 1H-Nuclear Magnetic Resonance. As shown by Canonical Discriminant Analysis of Principal Coordinates, the levels of volatile organic compounds and free amino acids in faecal and/or urine samples were markedly affected by CD. Conclusion: As shown by the parallel microbiology and metabolome approach, the gluten-free diet lasting at least two years did not completely restore the microbiota and, consequently, the metabolome of CD children. Some molecules (e.g., ethyl-acetate and octyl-acetate, some short chain fatty acids and free amino acids, and glutamine) seems to be metabolic signatures of CD.

oil management techniques can definitely influence soil quality, and particularly soil organic matter content, biological complexity, structure, and water holding capacity. Tillage may also have a negative effect by increasing erosion and organic matter oxidation processes, which have unavoidable repercussions on fertility. The objective of the current research was to test the effects of five different management techniques applied for 35 years on a rain-fed almond grove (Prunus amygdalus Batsch) in a hot-dry environment on some physicochemical, hydrological, and biological parameters. The following soil management techniques were compared: no-till (NT), with weed control by preemergence herbicides; NT, with chemical weed control by foliar herbicides; NT, with weed control by mowing; tillage, with sowing and field bean green manuring; and conventional tillage. The current survey supplied interesting results, considering the typical soil and climate conditions of the tested area (southern Italy), characterized by high summer temperatures, low rainfall, clay loam soil, and an arable layer of 0.40 m. The most influenced values are those concerning the organic matter due to the supply of biomass resulting from weed mowing or field bean green manuring. The NT system with a single mowing in the spring seems to induce a higher water holding capacity (–15,000 hPa) as compared with the traditionally plowed soil. The biomass incorporation through field bean green manure resulted in a higher available water content (11.82%). All practices favoring an increase in organic matter induced a subsequent increase of microbial biomass content. The number of existing families and species of weed flora was largely influenced by different soil management techniques, as shown by the greater adaptation of grasses to the management practices involving weed control by foliar herbicide or mowing, and of several species associated with the technique involving the application of preemergence herbicides. In general, the almond orchard management involving minimum soil disturbance and the supply of biomass resulting from specially sown cover crops or weed development have shown substantial benefits to the physicochemical, hydrologic, and biologic soil properties.

Il ruolo della componente biotica del suolo nei processi rizosferici di mobilizzazione e assorbimento dei nutrienti è ancora oggetto di ampia discussione. Tuttavia sono sempre più numerose le evidenze a supporto di un ruolo determinante svolto dai Plant growth-Promoting Rhizobacteria (PGPR) sull’accumulo dei nutrienti nei tessuti vegetali che è il risultato anche di ciò che accade alla rizosfera nei cicli biogeochimici dei diversi elementi/nutrienti. Pertanto, lo scopo di questo lavoro è stato quello di valutare gli effetti fisiologici e biochimici indotti dal PGPR Azospirillum brasilense su piante di cetriolo cresciute in un suolo calcareo. Inoltre, sono stati valutati anche gli effetti indotti dal rilascio di essudati radicali sulle componenti mineralogiche del suolo. A tale scopo, le piante di cetriolo sono state cresciute per 14 giorni in coltura idroponica in condizione di carenza di Fe e successivamente poste in contatto per 7 giorni con un suolo calcareo inoculato con A. brasilense. Al momento del campionamento, sono stati rilevati i parametri di crescita delle piante, i profili quali-quantitativi degli essudati rilasciati e il contenuto dei nutrienti nei tessuti. Inoltre, le variazione nella componente mineralogica del suolo sono state studiate mediante la tecnica XRPD (X-ray powder diffraction). I risultati ottenuti mostrano che la presenza di A. brasilense nel suolo facilita il recupero delle piante dalla clorosi ferrica, molto probabilmente tramite la modulazione dell’attività di essudazione della pianta, sia da un punto di vista qualitativo che quantitativo. In particolare, è stato evidenziato che due amminoacidi, la glicina e il glutammato, potrebbero essere coinvolti nell’interazione tra la pianta e il microrganismo nel processo di mobilizzazione del Fe dal suolo. L’analisi XRPD del suolo rizosferico di piante di cetriolo cresciute in carenza di Fe ha evidenziato una drastica riduzione della componente amorfa, probabilmente causata dal rilascio di amminoacidi, composti fenolici e acidi organici. In ogni caso, tali analisi hanno evidenziato che gli essudati rilasciati da piante di cetriolo carenti di Fe sono in grado di indurre modificazioni nella componente mineralogica del suolo calcareo in un breve lasso di tempo (7 giorni).

Plant growth-promoting rhizobacteria (PGPR) are soil bacteria that are able to colonize rhizosphere and to enhance plant growth by means of a wide variety of mechanisms like organic matter mineralization, biological control against soil-borne pathogens, biological nitrogen fixation, and root growth promotion. A very interesting feature of PGPR is their ability of enhancing nutrient bioavailability. Several bacterial species have been characterized as P-solubilizing microorganisms while other species have been shown to increase the solubility of micronutrients, like those that produce siderophores for Fe chelation. The enhanced amount of soluble macro- and micronutrients in the close proximity of the soil-root interface has indeed a positive effect on plant nutrition. Furthermore, several pieces of evidence highlight that the inoculation of plants with PGPR can have considerable effects on plant at both physiological and molecular levels (e.g., induction of rhizosphere acidification, up- and downregulation of genes involved in ion uptake, and translocation), suggesting the possibility that soil biota could stimulate plants being more efficient in retrieving nutrients from soil and coping with abiotic stresses. However, the molecular mechanisms underlying these phenomena, the signals involved as well as the potential applications in a sustainable agriculture approach, and the biotechnological aspects for possible rhizosphere engineering are still matters of discussion.

A revegetation process of a closed urban solid waste landfill was initiated in 2009 for the environmental restoration of the site. The landfill is located in Foggia district in the North part of the Apulia region. Growth experiments in pots and in open field were carried out on eight plant species in order to select those that could be more appropriate in the particular extreme field conditions (climate, high slope of the walls, bad leachate control). In particular, plant growth was assessed in two different soil types (agricultural and leachate contaminated soil). Among the different species assessed, Pistacia lentiscus L. and Puccinellia borreri. L. were selected because of their performance in the contaminated soil and mostly since they are typical native Mediterranean plant species. Rhizosphere and bulk soil of these two plant species, sampled from the pots, has been characterized by biochemical and molecular methods in order to highlight shift in the composition of the microbial community caused by the presence of leachate. Enzymatic activities (dehydrogenase, fluorescein diacetate hydrolysis, ?-glucosidase and protease) and microbial biomass C were analyzed to evaluate the activity of microbial population of soils. The molecular characterization was carried out by the direct extraction of DNA, gene amplification of the 16S rRNA of the Eubacteria by PCR and specific groups such as the Actinomycetes and Ammonia-oxidizing bacteria. The amplification products were separated by DGGE under appropriate conditions of denaturation and the resulting genetic fingerprinting were analyzed using the software Bionumerics.

The influence of humic substances (HS) on crops and soil microbial communities still lacks a complete understanding. The influence of HS on chemical composition of maize rhizodeposits and diversity of rhizosphere microbial communities was studied here in rhizobox systems. Eleven different HS were characterized by elemental analyses and 13C-CPMAS NMR, and applied to the upper compartment of rhizobox systems cultivated with maize. Once a full rhizoplane was formed, the lower compartment of the rhizoboxes was sampled with a slicing device, in order to obtain rhizopshere (0–2 mm) and bulk (>10 mm) soil samples, which were subjected to chemical analyses of organic acids and sugars (pentoses and hexoses), bioassays with a luminescent biosensor for the detection and quantification of bioavailable C forms, and analyzed for bacterial diversity by means of PCR-DGGE fingerprint. Multivariate techniques were applied to assess correlations between the parameters involved and to assess the most relevant chemical drivers of rhizosphere microbial diversity. Results showed that the hydrophilic and poorly aromatic HS changed the pattern of sugars and organic acids released in the rhizosphere by maize roots, and this, in turn, affected the composition of rhizosphere microbial communities. Differences in chemical and microbiological composition were also found, but to a lesser extent, among bulk soil samples of different treatments, thus indicating that rhizodeposits can influence soil composition at several mm distance from the root plane. Multiple regression analyses suggested a prominent role of succinic and malic acid in shaping the diversity of microbial communities. The approach and results described here proved to be useful to test bioactivity of HS towards an agricultural crop, and identify correlations between chemical and microbiological constituents at the tri-phase soil–roots–microbes system.

Soil is an extremely heterogeneous matrix where organic and inorganic phases coexist with a variety of living organisms. In particular, the rhizosphere is a very dynamic environment where plants and microorganisms compete (or cooperate) for mineral nutrients. In such a dynamic system, thermodynamic previsions about chemical processes can be controverted by peculiar conditions, which may kinetically favor other unexpected reactions. It is widely accepted that low-molecular-weight organic acids released into the rhizosphere have complexing properties thereby enhancing the solubility of several mineral elements and increasing their availability for plant uptake. Among these, it is well known that citrate can increase mineral nutrients mobility (particularly Fe), especially in alkaline soils. However, experimental evidence showed that, in a calcareous soil (pH 8.2, CaCO3 61.8% w/w, Corg 0.86% w/w), citrate (0.1 mM and 1 mM) was not as effective in solubilizing Cu as Al, Fe and Mn from soil. In addition, Automated Particle Analyses (APA) using Scanning Electron Microscopy coupled to Energy Dispersive X-ray spectroscopy (SEM-EDX) and Cluster Analysis evidenced the formation of a large number of aluminum oxide (AlOx) particles, about 50% of which containing a significant amount of Cu. In this research we demonstrated that in particular conditions of alkalinity and high microbial activity, which can be typically found in the rhizosphere of plants grown in calcareous soils, citrate efficacy in mobilizing Cu is strongly reduced by co-precipitation of the metal within Al (hydr)oxides. To prove this hypothesis, conditions similar to those occurring in the real soil (100% calcite, pH 8.2, [Al]=17 mg/l, [Cu]=7 mg/l) have been simulated, at first, by gradually reducing the concentration of citrate and thus mimicking the action of soil microorganisms feeding on citrate (as also observed experimentally) and, in a second step, by introducing soil microorganisms into the system. As a result, in both cases we observed a significant and concomitant decrease of Al and Cu solubility; then APA analyses on the solid phase revealed the formation of Cu-containing Al (hydr)oxides almost identical (in size, shape and composition) to those observed in the real sample. In addition, 16S rDNA PCR-DGGE of DNA extracted from the samples revealed changes in microbial communities suggesting that specific bacteria may be responsible for the degradation of citrate-metal-complexes. Sequencing of the main DNA gel bands allowed identification of these bacteria as Sphingomonax sp., Sphingopyxis sp. and Propionibacterium sp. In conclusion, in highly calcareous soils, citrate can be ineffective in mobilizing Cu from soil because specific soil microbes can degrade Al-citrate complexes thus causing the co-precipitation of soluble Cu within Al-(hydr)oxides and making it unavailable for plants.

Sustainable soil management of orchards can have positive effects on both soils and crop yields due to increases in microbial biomass, activity and complexity. The aim of this study was to investigate medium-term effects (12 yr) of two different management practices termed ‘sustainable’ (ST) and ‘conventional’ (CT) on soil microbial composition and metabolic diversity of a rainfed mature olive orchard in Southern Italy. ST included no-till, self-seeding weeds (mainly graminaceous and leguminosae), and mulch derived from olive tree prunings, whilst CT was managed by frequent tillage and included severe pruning with residues removed from the orchard. Microbial analyses were carried out by culture-dependent methods (microbial cultures and Biolog). Molecular methods were used to confirm the identification by light microscopy of the isolates of fungi and Streptomyces. Significantly more culturable fungi and bacteria were found in ST than in CT. The number of fungal groups in ST was also significantly greater than in CT. Overall and substrate-specific Biolog metabolic diversity indices of microbial communities and soil enzyme activities were greater in ST. The results demonstrate that soil micro-organisms respond positively to sustainable orchard management characterized by periodic applications of locally derived organic matter. This study confirms the need to encourage farmers with orchards in the Mediterranean basin to practise soil management based on organic matter inputs associated with zero tillage to improve soil functionality.

In this research, the optimum growth conditions for two zinc solubilizing bacteria (ZSB) have been studied for their potential application as bioinoculants to overcome Zn unavailability in soils. For this purpose, a laboratory-scale experiment was carried out to evaluate the zinc solubilizing ability of 80 plant growth promoting bacteria (PGPB) strains isolated from the rhizosphere of barley and tomato plants. To select effective ZSB, isolates were evaluated on Tris-mineral medium supplemented separately with zinc oxide, zinc carbonate, and zinc phosphate at a concentration of 0.1%. Two strains (Agrobacterium tumefaciens and Rhizobium sp.) were selected, based on a clear halo zone around their colonies in the solid medium supplemented with zinc oxide after 10 days of incubation at 29 °C. Results of solubilization at different pH values showed that these strains had solubilization activity in the range of pH 8-10 while no solubilization was observed at pH 6 and 7. The maximum Zn solubilization values were noted at pH 9: 51.4 mg L-1 (Agrobacterium tumefaciens) and 72.1 mg L-1 (Rhizobium sp). According to findings, bacterial growth was affected by different NaCl concentrations under in vitro condition. The salt concentration required for 50% inhibition of absorbance was 2.11 and 2.27% NaCl for Agrobacterium tumefaciens and Rhizobium sp., respectively. The maximum bacterial growth was observed at about 0.8% NaCl concentration.

Plants have evolved two different strategies (Strategy I and II) to cope with Fe shortage, based on the exudation of organic and inorganic compounds to favor its mobilization and the root uptake. The role of the soil biotic component in the nutritional processes in the rhiszophere needs to be elucidated, since plants inoculated with PGPR showed an increased content of nutrients and a stronger resistance to abiotic stresses. The aim of the present work is the evaluation of the physiological effects, induced by Azospirillum brasilense in a calcareous soil on cucumber plants. Plants were grown in hydroponic Fe deficient solution followed by a 7-day period of contact with the A. brasilense-inoculated calcareous soil. At sampling, biometrics measurements, quali-quantitative analyses of root exudates and analyses of the nutrients content in plant tissues were carried out. Variations in soil mineralogy were assessed by X-ray powder diffraction (XRPD). Our results showed that A. brasilense facilitates plant growth in calcareous soils due to an enhanced recovery from the micronutrient deficiency. A. brasilense increases most likely the Fe availability within the rhizosphere by a) affecting the solubilisation of Fe thanks to the siderophore release and b) up and down-regulating the exudation activity of plants with an effect also on its molecular complexity. Further studies are needed to better understand and highlight the interactions between these two mechanisms and microorganisms. In particular, the present study shed light for the first time on two AAs, namely Gly and Glu, which could be involved in the plant-microorganism-soil interaction for the retrieval of Fe within a calcareous soil. XRPD analysis revealed a slight decrease of calcite and an increase of smectite under Fe-deficiency conditions.