Effect of temperature and free ammonia on nitrification and nitrite accumulation in landfill leachate and analysis of its nitrifying bacterial community by FISH

The cause of seasonal failure of a nitrifying municipal landfill leachate treatment plant utilizing a fixed biofilm was investigated by wastewater analyses and batch respirometric tests at every treatment stage. Nitrification of the leachate treatment plant was severely affected by the seasonal temperature variation. High free ammonia (NH3-N) inhibited not only nitrite oxidizing bacteria (NOB) but also ammonia oxidizing bacteria (AOB). In addition, high pH also increased free ammonia concentration to inhibit nitrifying activity especially when the NH4-N level was high. The effects of temperature and free ammonia of landfill leachate on nitrification and nitrite accumulation were investigated with a semi-pilot scale biofilm airlift reactor. Nitrification rate of landfill leachate increased with temperature when free ammonia in the reactor was below the inhibition level for nitrifiers. Leachate was completely nitrified up to a load of 1.5 kg NH4-N m(-3) d(-1) at 28 degrees C. The activity of NOB was inhibited by NH3-N resulting in accumulation of nitrite. NOB activity decreased more than 50% at 0.7 mg NH3-N L-1. Fluorescence in situ hybridization (FISH) was carried out to analyze the population of AOB and NOB in the nitrite accumulating nitrifying biofilm. NOB were located close to AOB by forming small clusters. A significant fraction of AOB identified by probe Nso1225 specifically also hybridized with the Nitrosonlonas specific probe Nsm156. The main NOB were Nitrobacter and Nitrospira which were present in almost equal amounts in the biofilm as identified by simultaneous hybridization with Nitrobacter specific probe Nit3 and Nitrospira specific probe Ntspa662. (c) 2005 Elsevier Ltd. All rights reserved.

Hydrochemistry measurements and diazotroph abundances

Microbial dinitrogen (N2) fixation, the nitrogenase enzyme-catalysed reduction of N2 gas into biologically available ammonia, is the main source of new nitrogen (N) in the ocean. For more than 50 years, oceanic N2 fixation has mainly been attributed to the activity of the colonial cyanobacterium Trichodesmium. Other smaller N2-fixing microorganisms (diazotrophs)-in particular the unicellular cyanobacteria group A (UCYN-A)-are, however, abundant enough to potentially contribute significantly to N2 fixation in the surface waters of the oceans. Despite their abundance, the contribution of UCYN-A to oceanic N2 fixation has so far not been directly quantified. Here, we show that in one of the main areas of oceanic N2 fixation, the tropical North Atlantic7, the symbiotic cyanobacterium UCYN-A contributed to N2 fixation similarly to Trichodesmium. Two types of UCYN-A, UCYN-A1 and -A2, were observed to live in symbioses with specific eukaryotic algae. Single-cell analyses showed that both algae-UCYN-A symbioses actively fixed N2, contributing ~20% to N2 fixation in the tropical North Atlantic, revealing their significance in this region. These symbioses had growth rates five to ten times higher than Trichodesmium, implying a rapid transfer of UCYN-A-fixed N into the food web that might significantly raise their actual contribution to N2 fixation. Our analysis of global 16S rRNA gene databases showed that UCYN-A occurs in surface waters from the Arctic to the Antarctic Circle and thus probably contributes to N2 fixation in a much larger oceanic area than previously thought. Based on their high rates of N2 fixation and cosmopolitan distribution, we hypothesize that UCYN-A plays a major, but currently overlooked role in the oceanic N cycle.

Global distributions of diazotrophs abundance and biomass - Depth integrated values computed from a collection of source datasets - Contribution to the MAREDAT World Ocean Atlas of Plankton Functional Types

The MAREDAT atlas covers 11 types of plankton, ranging in size from bacteria to jellyfish. Together, these plankton groups determine the health and productivity of the global ocean and play a vital role in the global carbon cycle. Working within a uniform and consistent spatial and depth grid (map) of the global ocean, the researchers compiled thousands and tens of thousands of data points to identify regions of plankton abundance and scarcity as well as areas of data abundance and scarcity. At many of the grid points, the MAREDAT team accomplished the difficult conversion from abundance (numbers of organisms) to biomass (carbon mass of organisms). The MAREDAT atlas provides an unprecedented global data set for ecological and biochemical analysis and modeling as well as a clear mandate for compiling additional existing data and for focusing future data gathering efforts on key groups in key areas of the ocean. The present data set presents depth integrated values of diazotrophs abundance and biomass, computed from a collection of source data sets.

Geochemical composition of ODP Site 210-1276 shales

During Ocean Drilling Program Leg 210, a greatly expanded sedimentary sequence of continuous Cretaceous black shales was recovered at Site 1276. This section corresponds to the Hatteras Formation, which has been documented widely in the North Atlantic Ocean. The cored sequence extends from the lowermost Albian, or possibly uppermost Aptian, to the Cenomanian/Turonian boundary and is characterized by numerous gravity-flow deposits and sporadic, finely laminated black shales. The sequence also includes several sedimentary intervals with high total organic carbon (TOC) contents, in several instances of probable marine origin that may record oceanic anoxic events (OAE). These layers might correspond to the Cenomanian-Turonian OAE 2; the mid-Cenomanian event; and OAE 1b, 1c, and 1d in the Albian. In addition, another interval with geochemical characteristics similar to OAE-type layers was recognized in the Albian, although it does not correspond to any of the known OAEs. This study investigates the origin of the organic matter contained within these black shale intervals using TOC and CaCO3 contents, Corg/Ntot ratios, organic carbon and nitrogen isotopes, trace metal composition, and rock-eval analyses. Most of these black shale intervals, especially OAE 2 and 1b, are characterized by low 15N values (<0) commonly observed in mid-Cretaceous black shales, which seem to reflect the presence of an altered nitrogen cycle with rates of nitrogen fixation significantly higher than in the modern ocean.

(Supplement 1) Selected chemical composition of Icelandic river waters and bedrock age

In this study we map the spatial distribution of selected dissolved constituents in Icelandic river waters using GIS methods to study and interpret the connection between river chemistry, bedrock, hydrology, vegetation and aquatic ecology. Five parameters were selected: alkalinity, SiO2, Mo, F and the dissolved inorganic nitrogen and dissolved inorganic phosphorus mole ratio (DIN/DIP). The highest concentrations were found in rivers draining young rocks within the volcanic rift zone and especially those draining active central volcanoes. However, several catchments on the margins of the rift zone also had high values for these parameters, due to geothermal influence or wetlands within their catchment area. The DIN/DIP mole ratio was higher than 16 in rivers draining old rocks, but lowest in rivers within the volcanic rift zone. Thus primary production in the rivers is limited by fixed dissolved nitrogen within the rift zone, but dissolved phosphorus in the old Tertiary catchments. Nitrogen fixation within the rift zone can be enhanced by high dissolved molybdenum concentrations in the vicinity of volcanoes. The river catchments in this study were subdivided into several hydrological categories. Importantly, the variation in the hydrology of the catchments cannot alone explain the variation in dissolved constituents. The presence or absence of central volcanoes, young reactive rocks, geothermal systems and wetlands is important for the chemistry of the river waters. We used too many categories within several of the river catchments to be able to determine a statistically significant connection between the chem¬istry of the river waters and the hydrological categories. More data are needed from rivers draining one single hydrological category. The spatial dissolved constituent distribution clearly revealed the difference between the two extremes, the young rocks of the volcanic rift zone and the old Tertiary terrain.

Seawater carbonate chemistry and combined physiological effects of CO2 and light on the N2-fixing cyanobacterium Trichodesmium IMS101 during experiments, 2010

Recent studies on the diazotrophic cyanobacterium Trichodesmium erythraeum(IMS101) showed that increasing CO2 partial pressure (pCO2) enhances N2 fixation and growth. Significant uncertainties remain as to the degree of the sensitivity to pCO2, its modification by other environmental factors, and underlying processes causing these responses. To address these questions, we examined the responses ofTrichodesmium IMS101 grown under a matrix of low and high levels of pCO2 (150 and 900 µatm) and irradiance (50 and 200 µmol photons m-2 s-1). Growth rates as well as cellular carbon and nitrogen contents increased with increasing pCO2 and light levels in the cultures. The pCO2-dependent stimulation in organic carbon and nitrogen production was highest under low light. High pCO2 stimulated rates of N2fixation and prolonged the duration, while high light affected maximum rates only. Gross photosynthesis increased with light but did not change with pCO2. HCO3- was identified as the predominant carbon source taken up in all treatments. Inorganic carbon uptake increased with light, but only gross CO2 uptake was enhanced under high pCO2. A comparison between carbon fluxes in vivo and those derived from 13C fractionation indicates high internal carbon cycling, especially in the low-pCO2treatment under high light. Light-dependent oxygen uptake was only detected underlow pCO2 combined with high light or when low-light-acclimated cells were exposed to high light, indicating that the Mehler reaction functions also as a photoprotective mechanism in Trichodesmium. Our data confirm the pronounced pCO2 effect on N2fixation and growth in Trichodesmium and further show a strong modulation of these effects by light intensity. We attribute these responses to changes in the allocation of photosynthetic energy between carbon acquisition and the assimilation of carbon and nitrogen under elevated pCO2. These findings are supported by a complementarystudy looking at photosynthetic fluorescence parameters of photosystem II, photosynthetic unit stoichiometry (photosystem I:photosystem II), and pool sizes of key proteins in carbon and nitrogen acquisition.

Inhibition of glutamine synthetase by phosphinothricin leads to transcriptome reprograming in root nodules of Medicago truncatula

Glutamine synthetase (GS) is a vital enzyme for the assimilation of ammonia into amino acids in higher plants. In legumes, GS plays a crucial role in the assimilation of the ammonium released by nitrogen-fixing bacteria in root nodules, constituting an important metabolic knob controlling the nitrogen (N) assimilatory pathways. To identify new regulators of nodule metabolism, we profiled the transcriptome of Medicago truncatula nodules impaired in N assimilation by specifically inhibiting GS activity using phosphinothricin (PPT). Global transcript expression of nodules collected before and after PPT addition (4, 8, and 24 h) was assessed using Affymetrix M. truncatula GeneChip arrays. Hundreds of genes were regulated at the three time points, illustrating the dramatic alterations in cell metabolism that are imposed on the nodules upon GS inhibition. The data indicate that GS inhibition triggers a fast plant defense response, induces premature nodule senescence, and promotes loss of root nodule identity. Consecutive metabolic changes were identified at the three time points analyzed. The results point to a fast repression of asparagine synthesis and of the glycolytic pathway and to the synthesis of glutamate via reactions alternative to the GS/GOGAT cycle. Several genes potentially involved in the molecular surveillance for internal organic N availability are identified and a number of transporters potentially important for nodule functioning are pinpointed. The data provided by this study contributes to the mapping of regulatory and metabolic networks involved in root nodule functioning and highlight candidate modulators for functional analysis.

Aplicação foliar de molibdênio em adubos verdes e o desempenho do feijoeiro em sucessão

The knowledge of molybdenum application in legumes on the availability of N, by BNF, increased enzymatic activity and the residual effect caused on crops growth and yield can contribute to the greater scientific understanding involved in green manure processes. The aim of this study was to evaluate the Mo application and the N from Crotalaria juncea and Canavalia ensiformis green manures on common bean performance. Were conducted field experiments for the crops succession system (green manures - common bean) and laboratory essays for the enzymatic activities. Green manure production was installed in a factorial arrangement 2 x 4, with two green manure legumes species, sunnhemp (Crotalaria juncea) and jack beans (Canavalia ensiformis), and four Mo doses (0, 40, 80, 120 g ha-1) in the form of sodium molybdate (Na2MoO4), foliar applied, in a randomized block design with four replicates. For succession crop (common bean) additional treatment was added, beans grown without any fertilization, following the same experimental design from the previous crop. The dry matter decomposition and the N mineralization of green manure were monitored through collection of residues over time, by using the litter bags method. In laboratory were carried out tests of nitrate reductase activity in green manures and common beans at 90 and 66 days after sowing, respectively. The sunnhemp responded linearly positively to the application of Mo as the dry matter and N accumulation. While the jack beans presented a negative quadratic response for dry matter and there was no adjustment of regression models to N. The jack beans showed a higher decomposition rate and N mineralization compared to sunnhemp. The half lives for decomposing 50% of dry matter on the soil was 123 and 104 days to sunnhemp and jack beans, respectively, and 50% of N present in the residues was mineralized at 93 and 85 days. In common bean, differed from the control for number of pods the dose of 40 g ha-1 of Mo in both species of green manures and the dose 80 g ha-1 of Mo in jack beans. For number of grains only in sunnhemp on the dose of 40 g ha-1 of Mo differ from the control. The nitrate reductase activity was influenced by developmental stage of green manure species. In common bean, the activity of nitrate reductase was up to three times higher than the dose 0 g ha-1 of Mo compared to treatment with application of Mo in both species. There was no effect of Mo doses or species of green manure on common bean yield.

Evaluación del efecto de la inoculación de Bacillus spp. con potencial fijador de nitrógeno en cultivos de Chlorella sorokiniana

El uso de microalgas y PGPBs como principio activo de problemas ambientales ha generado interés científico en los últimos años -- Entre las soluciones propuestas se encuentra el uso de los cocultivos de estos 2 tipos de microrganismos para la formulación de bioinsumos y la biorremedación -- El objetivo de este estudio se centró en evaluar el efecto de la inoculación de Bacillus spp. con potencial fijador de nitrógeno ambiental en cultivos de Chlorella sorokiniana a nivel de matraz -- Se encontró que 10 cepas PGPB, del banco de cepas del grupo CIBIOP de la Universidad EAFIT, probablemente fijaron nitrógeno debido a que crecen en medio NFb -- De las cuales, Bacillus subtilis EA-CB0575 promueve el crecimiento de la microalga Chlorella sorokiniana UTEX 1230, aumentando en un 94% la densidad celular y en 4.5 veces el tamaño de la microalga comparada su crecimiento individual -- Finalmente se usó el método ARA para evaluar fijación de nitrógeno, y se encontró que la bacteria fue capaz de reducir el acetileno en etileno cuando se siembra en medio NFb y BBM libre de nitrógeno, además cuando se cocultivo con la microalga, lo que indicaría que posiblemente la fijación de nitrógeno sea el método de promoción de crecimiento microalgar

Proteomic analysis of Rhizobium freirei PRF 81 responses to low pH.

Rhizobium freirei PRF 81 is employed in common bean commercial inoculants in Brazil, due to its outstanding efficiency in fixing nitrogen, competitiveness and tolerance to abiotic stresses. Among the environmental conditions faced by rhizobia in soils, acidity is perhaps the encountered most, especially in Brazil. So, we used proteomics based approaches to study the responses of PRF 81 to a low pH condition. R. freirei PRF 81 was grown in TY medium until exponential phase in two treatments: pH 6,8 and pH 4,8. Whole-cell proteins were extracted and separated by two-dimensional gel electrophoresis, using IPG-strips with pH range 4-7 and 12% polyacrilamide gels. The experiment was performed in triplicate. Protein spots were detected in the high-resolution digitized gel images and analyzed by Image Master 2D Platinum v 5.0 software. Relative volumes (%vol) of compared between the two conditions tested and were statistically evaluated (p ≤ 0.05). Even knowing that R. freirei PRF 81 can still grow in more acid conditions, pH 4.8 was chosen because didn´t affect significantly the bacterial growth kinetics, a factor that could compromise the analysis. Using a narrow pH range, the gel profiles displayed a better resolution and reprodutibility than using broader pH range. Spots were mostly concentrated between pH 5-7 and molecular masses between 17-95 kDa. From the six hundred well-defined spots analyzed, one hundred and sixty-three spots presented a significant change in % vol, indicating that the pH led to expressive changes in the proteome of R. freirei PRF 81. Of these, sixty-one were up-regulated and one hundred two was downregulated in pH 4.8 condition. Also, fourteen spots were only identified in the acid condition, while seven spots was exclusively detected in pH 6.8. Ninety-five differentially expressed spots and two exclusively detected in pH 4,8 were selected for Maldi-Tof identification. Together with the genome sequencing and the proteome analysis of heat stress, we will search for molecular determinants of PRF 81 related to capacity to adapt to stressful tropical conditions.

A novel aerobic-anoxic biological filter for nitrogen removal from UASB effluent using biogas compounds as electron donors for denitrification

The performance of a new trickling filter (TF) configuration composed of an upper compartment for nitrification and a lower compartment for denitrification of effluent from a UASB reactor treating domestic sewage was evaluated. The TF was packed with new plastic material characterized by its durability and high percentage of void spaces. The feasibility of using the reduced compounds present in the biogas produced by a UASB reactor as electron donor for denitrification was also evaluated. Efficient nitrification and denitrification was achieved for the mean hydraulic (5.6 m(3) m(-2) d(-1)) organic (0.26 kg COD m(-3) d(-1)) and ammonia-N (0.08 kg m(-3) d(-1)) loading rates applied, resulting in ammonia-N removal ranging from 60 to 74%. The final effluent presented ammonia-N lower than 13 mg L(-1). Despite the presence of dissolved oxygen (DO) in the denitrification compartment, its performance was considered quite satisfactory and final nitrate concentrations were lower than 10 mg L(-1). The results indicate that methane was the main electron donor used for denitrification. Additionally, denitrification can probably be improved by avoiding high DO concentration in the denitrification compartment and by enhancing biogas transfer in the anoxic zone.

delta N-15 values of tropical savanna and monsoon forest species reflect root specialisations and soil nitrogen status

A large number of herbaceous and woody plants from tropical woodland, savanna, and monsoon forest were analysed to determine the impact of environmental factors (nutrient and water availability, fire) and biological factors (microbial associations, systematics) on plant delta(15)N values. Foliar delta(15)N values of herbaceous and woody species were not related to growth form or phenology, but a strong relationship existed between mycorrhizal status and plant delta(15)N. In woodland and savanna, woody species with ectomycorrhizal (ECM) associations and putative N-2-fixing species with ECM/arbuscular (AM) associations had lowest foliar delta(15)N values (1.0-0.6parts per thousand), AM species had mostly intermediate delta(15)N values (average +0.6parts per thousand), while non-mycorrhizal Proteaceae had highest delta(15)N values (+2.9 to +4.1parts per thousand). Similar differences in foliar delta(15)N were observed between AM (average 0.1 and 0.2parts per thousand) and non-mycorrhizal (average +0.8 and +0.3parts per thousand) herbaceous species in woodland and savanna. Leguminous savanna species had significantly higher leaf N contents (1.8-2.5% N) than non-fixing species (0.9-1.2% N) indicating substantial N acquisition via N-2 fixation. Monsoon forest species had similar leaf N contents (average 2.4% N) and positive delta(15)N values (+0.9 to +2.4parts per thousand). Soil nitrification and plant NO3- use was substantially higher in monsoon forest than in woodland or savanna. In the studied communities, higher soil N content and nitrification rates were associated with more positive soil delta(15)N and plant delta(15)N. In support of this notion, Ficus, a high NO3- using taxa associated with NO3- rich sites in the savanna, had the highest delta(15)N values of all AM species in the savanna. delta(15)N of xylem sap was examined as a tool for studying plant delta(15)N relations. delta(15)N of xylem sap varied seasonally and between differently aged Acacia and other savanna species. Plants from annually burnt savanna had significantly higher delta(15)N values compared to plants from less frequently burnt savanna, suggesting that foliar N-15 natural abundance could be used as marker for assessing historic fire regimes. Australian woodland and savanna species had low leaf delta(15)N and N content compared to species from equivalent African communities indicating that Australian biota are the more N depauperate. The largest differences in leaf delta(15)N occurred between the dominant ECM Australian and African savanna (miombo) species, which were depleted and enriched in N-15, respectively. While the depleted delta(15)N of Australian ECM species are similar to those of previous reports on ECM species in natural plant communities, the N-15-enriched delta(15)N of African ECM species represent an anomaly.

Photoelectron spectroscopy of nitrogen containing molecules of biological and industrial interest

A thesis submitted for the degree of Ph. D. in Physics

Fertilization and cover crop effects on soil nitrogen and plant nutrition in a young guarana plantation

Fruit tree production is gaining an increasing importance in the central Amazon and elsewhere in the humid tropics, but very little is known about the nutrient dynamics in the soil-plant system. The present study quantified the effects of fertilization and cover cropping with a legume (Pueraria phaseoloides (Roxb.) Benth.) on soil nitrogen (N) dynamics and plant nutrition in a young guarana plantation (Paullinia cupana Kunth. (H.B. and K.) var. sorbilis (Mart.) Ducke) on a highly weathered Xanthic Ferralsol. Large subsoil nitrate (NO3-) accumulation at 0.3-3 m below the guarana plantation indicated N leaching from the topsoil. The NO3- contents to a depth of 2 m were 2.4 times greater between the trees than underneath unfertilized trees (P<0.05). The legume cover crop between the trees increased soil N availability as shown by elevated aerobic N mineralization and lower N immobilization in microbial biomass. The guarana N nutrition and yield did not benefit from the N input by biological fixation of atmospheric N2 by the legume cover (P>0.05). Even without a legume intercrop, large amounts of NO3- were found in the subsoil between unfertilized trees. Subsoil NO3- between the trees could be utilized, however, by fertilized guarana. This can be explained by a more vigorous growth of fertilized trees which had a larger nutrient demand and exploited a larger soil volume. With a legume cover crop, however, more mineral N was available at the topsoil which was leached into the subsoil and consequently accumulated at 0.3-3 m depth. Fertilizer additions of P and K were needed to increase subsoil NO3- use between trees.