Use of a lux-marked rhizobacterium as a biosensor to assess changes in rhizosphere C flow due to pollutant stress

The flow of carbon from plant roots into soil supports a range of microbial processes and is therefore critical to ecosystem function and health. Pollution-induced stress, which influences rhizosphere C flow is of considerable potential importance, and therefore needs to be evaluated. This paper reports on a method, based on reporter gene technology, for quantifying pollutant effects on rhizosphere C flow. The method uses the lux-marked rhizobacterium Pseudomonas fluorescens, where bioluminescence output of this biosensor is directly correlated with the metabolic activity and reports on C flow in root exudate. Plantago lanceolata was treated with paraquat (representing a model pollutant stress) in a simple microcosm system. The lux-biosensor response correlated closely with C concentrations in the exudate and demonstrated that the pollutant stress increased the C flow from the plantago roots, 24 h after application of the herbicide. The lux-reporter system therefore potentially offers a technique for use in assessing the impact of pollutant stress on rhizosphere C flow through the soil microbial biomass.

A critical review of labelling techniques used to quantify rhizosphere carbon-flow

The rhizosphere is a major sink for photo-assimilated carbon and quantifying inputs into this sink is one of the main goals of rhizosphere biology as organic carbon lost from plant roots supports a higher microbial population in the rhizosphere compared to bulk soil. Two fundamentally different¹⁴CO₂ labelling strategies have been developed to estimate carbon fluxes through the rhizosphere - continuous feeding of shoots with labelled carbon dioxide and pulse-chase experiments. The biological interpretation that can be placed on the results of labelling experiments is greatly biased by the technique used. It is the purpose of this paper to assess the advantages, disadvantages and the biological interpretation of both continuous and pulse labelling and to consider how to partition carbon fluxes within the rhizosphere. © 1994 Kluwer Academic Publishers.

Die mikrobielle Besiedlung von Wurzeloberfläche und Rhizosphäre in ihrer Bedeutung für Stoffumsätze in Böden

Im Vordergrund der Arbeit stand die Erfassung der mikrobiellen Biomasse bzw. Residualmasse an der Wurzeloberfläche, im Rhizosphärenboden und im umgebenden Boden. Durch den Vergleich von verschiedenen Methoden zur Erfassung der mikrobiellen Biomasse wurden die Gehalte von pilzlichem und bakteriellem Kohlenstoff an der Rhizoplane und in der Rhizosphäre quantifiziert. Dabei wurde die Fumigations-Extraktions-Methode zur Erfassung der mikrobiellen Biomasse eingesetzt. Ergosterol diente als Indikator für die pilzliche Biomasse und die Aminozucker Glucosamin und Muraminsäure sollten Aufschluss geben über die bakterielle und pilzliche Biomasse bzw. Residualmasse in den drei Probenfraktionen. Dazu wurden Umrechnungsfaktoren erstellt, die zur Berechnung des bakteriellen und pilzlichen Kohlenstoffs aus den Gehalten von Muraminsäure und Pilz-Glucosamin dienten. Die Bestimmung von Aminozuckern wurde insoweit modifiziert, dass sowohl in Boden- als auch in Wurzelhydrolysaten die Messung von Glucosamin, Galactosamin, Muraminsäure und Mannosamin gleichzeitig als automatisiertes Standardverfahren mit Hilfe der HPLC erfolgen konnte. Es wurden drei Gefäßversuche durchgeführt: Im ersten Versuch wurde der Einfluss der Pflanzenart auf die mikrobielle Besiedlung der Wurzeloberflächen untersucht. Dabei wurden Wurzeln und Rhizosphärenboden von 15 verschiedenen Pflanzenarten miteinander verglichen. Im zweiten Versuch stand der Einfluss der mikrobiellen Biomasse eines Bodens auf die mikrobielle Besiedlung von Wurzeloberflächen im Vordergrund. Deutsches Weidelgras (Lolium perenne L.) wurde auf sieben verschiedenen Böden angezogen. Bei den Böden handelte es sich um sechs Oberböden, die sich hinsichtlich des Bodentyps und der Bewirtschaftungsform voneinander unterschieden, und einen Unterboden. Im dritten Versuch wurde die mikrobielle Besiedlung von Wurzeln nach teilweiser und vollständiger Entfernung der oberirdischen Biomasse beobachtet. Welsches Weidelgras (Lolium multiflorum Lam.) wurde 24 Tage nach der Aussaat beschnitten. Anschließend wurde über einen Versuchszeitraum von acht Tagen die mikrobielle Besiedlung an den Wurzeln und in den Bodenfraktionen bestimmt. Es bestätigte sich, dass der Einfluss der einzelnen Pflanzenart von entscheidender Bedeutung für die mikrobielle Besiedlung von Wurzeln ist. Bei fast allen Pflanzen wurde die mikrobielle Biomasse an den Wurzeln von Pilzen dominiert. Das Verhältnis von pilzlichem zu bakteriellem Kohlenstoff an den Wurzeln der 15 Pflanzenarten lag im Mittel bei 2,6. Bei der Betrachtung verschiedener Böden zeigte sich, dass die mikrobielle Besiedlung in tieferen Bodenschichten signifikant niedriger ist als in den Oberböden. Dabei war der Pilzanteil an der mikrobiellen Biomasse im Unterboden deutlich erhöht. Der Vergleich der Oberböden untereinander ergab, dass sowohl der Bodentyp als auch die Bewirtschaftungsform einen signifikanten Einfluss auf mikrobielle Besiedlung ausüben. Durch die teilweise oder vollständige Entfernung der oberirdischen Biomasse wurde eine Veränderung der mikrobiellen Besiedlung an den Wurzeln beobachtet. Das Verhältnis von pilzlichem zu bakteriellem Kohlenstoff sank in dem Versuchszeitraum von 2,5 auf 1,4. Dabei war die Förderung der Pilze in der Variante mit teilweise entfernter oberirdischer Biomasse relativ größer als in der Variante mit vollständig entfernter oberirdischer Biomasse. Entgegen der weit verbreiteten Annahme, dass bei den wurzelbesiedelnden Mikroorganismen die Bakterien gegenüber den Pilzen dominieren, zeigten die Ergebnisse ein gegensätzliches Bild. In allen drei Versuchen ergab sich gleichermaßen, dass sowohl im Boden als auch an den Wurzeln die Pilze gegenüber den Bakterien dominieren.

Bedeutung der C- und N-Rhizodeposition für das mikrobielle Wachstum und den mikrobiellen Umsatz von C und N in der Rhizosphäre in Abhängigkeit von der Entfernung zur Wurzel

Ziel dieser Arbeit war es, die Bedeutung des C und N mit Herkunft aus der Rhizodeposition für das mikrobielle Wachstum und den C- und N-Umsatz in der Rhizosphäre in Abhängigkeit von der Entfernung zur Wurzel zu untersuchen. Dazu wurde als wesentliche methodische Voraussetzung ein künstliches Rhizosphärensystem entwickelt, um die durch die Rhizodeposite induzierten Prozesse an der Grenzfläche zwischen Wurzeloberfläche und Boden zu untersuchen. Dieses eingesetzte Rhizosphärensystem wurde nach einem Vorbild eines in schon vielfältigen Untersuchungen der Rhizosphäre eingesetzten Systems von GAHOONIA und NIELSEN (1991) konzipiert. Zur Verfolgung der pflanzlichen C- und N-Rhizodeposition im Boden wurden 13C- und 15N-Tracer-Isotopen-Techniken zur Isotopenmarkierung der Testpflanzen eingesetzt. Zur Probenahme des Rhizosphärenbodens in räumlichen Abstand zur künstlichen Rhizoplane wurde eine Schneidvorrichtung nach FITZ et al. (2003) eingesetzt, die es ermöglicht frische Bodenproben in definiertem Abstand zur künstlichen Rhizoplane zu schneiden. Das Rhizosphärensystem wurde in 13C- und 15N-Doppelmarkierungsexperimenten mit Lolium perenne, Triticum aestivum und Avena sativa eingesetzt.Das unterschiedliche Ansprechen der mikrobiellen Gemeinschaft auf den Substrateintrag in unterschiedlicher Entfernung zur Wurzel zeigte komplexe Wechselwirkungen in Bezug auf das mikrobielle Wachstum, den mikrobiellen Umsatz, den Substrateintrag aus der Rhizodeposition und den stimulierten Abbau der nativen organischen Bodensubstanz. Der Eintrag von Rhizodepositen stellt daher eine bedeutende Funktion in der Regulation der mikrobiellen Biomasse und der Prozesse des Umsatzes der organischen Bodensubstanz während und wahrscheinlich bis nach der Vegetationsperiode dar. Die simultane Verfolgung von Rhizodepositions-C und -N im Boden und deren Funktionen innerhalb der Rhizosphäre, gerade im Hinblick auf die mikrobielle Biomasse und den Umsatz der organischen Substanz im Boden, führt zu einem besseren Verständnis der komplexen wechselseitigen Prozesse zwischen Pflanze und Boden.

Nitrogen and carbon dynamics in grassland soils and plants after application of digestate

A better understanding of effects after digestate application on plant community, soil microbial community as well as nutrient and carbon dynamics is crucial for a sustainable grassland management and the prevention of species and functional diversity loss. The specific research objectives of the thesis were: (i) to investigate effects after digestate application on grass species and soil microbial community, especially focussing on nitrogen dynamic in the plant-soil system and to examine the suitability of the digestate from the “integrated generation of solid fuel and biogas from biomass” (IFBB) system as fertilizer (Chapter 3). (ii) to investigate the relationship between plant community and functionality of soil microbial community of extensively managed meadows, taking into account temporal variations during the vegetation period and abiotic soil conditions (Chapter 4). (iii) to investigate the suitability of IFBB-concept implementation as grassland conservation measure for meadows and possible associated effects of IFBB digestate application on plant and soil microbial community as well as soil microbial substrate utilization and catabolic evenness (Chapter 5). Taken together the results indicate that the digestate generated during the IFBB process stands out from digestates of conventional whole crop digestion on the basis of higher nitrogen use efficiency and that it is useful for increasing harvestable biomass and the nitrogen content of the biomass, especially of L. perenne, which is a common species of intensively used grasslands. Further, a medium application rate of IFBB digestate (50% of nitrogen removed with harvested biomass, corresponding to 30 50 kg N ha-1 a-1) may be a possibility for conservation management of different meadows without changing the functional above- and belowground characteristic of the grasslands, thereby offering an ecologically worthwhile alternative to mulching. Overall, the soil microbial biomass and catabolic performance under planted soil was marginally affected by digestate application but rather by soil properties and partly by grassland species and legume occurrence. The investigated extensively managed meadows revealed a high soil catabolic evenness, which was resilient to medium IFBB application rate after a three-year period of application.

Cadaver decomposition in terrestrial ecosystems

A dead mammal (i.e. cadaver) is a high quality resource (narrow carbon:nitrogen ratio, high water content) that releases an intense, localised pulse of carbon and nutrients into the soil upon decomposition. Despite the fact that as much as 5,000 kg of cadaver can be introduced to a square kilometre of terrestrial ecosystem each year, cadaver decomposition remains a neglected microsere. Here we review the processes associated with the introduction of cadaver-derived carbon and nutrients into soil from forensic and ecological settings to show that cadaver decomposition can have a greater, albeit localised, effect on belowground ecology than plant and faecal resources. Cadaveric materials are rapidly introduced to belowground floral and faunal communities, which results in the formation of a highly concentrated island of fertility, or cadaver decomposition island (CDI). CDIs are associated with increased soil microbial biomass, microbial activity (C mineralisation) and nematode abundance. Each CDI is an ephemeral natural disturbance that, in addition to releasing energy and nutrients to the wider ecosystem, acts as a hub by receiving these materials in the form of dead insects, exuvia and puparia, faecal matter (from scavengers, grazers and predators) and feathers (from avian scavengers and predators). As such, CDIs contribute to landscape heterogeneity. Furthermore, CDIs are a specialised habitat for a number of flies, beetles and pioneer vegetation, which enhances biodiversity in terrestrial ecosystems.

Determinação de fósforo microbiano: comparação entre métodos e coberturas florestais

Muitos trabalhos mostram a importância da biomassa microbiana do solo (BMS), principalmente como fonte/dreno de C e de N em plantações florestais; contudo, são escassos os trabalhos relacionados ao fósforo microbiano (PBM), sobretudo aqueles relativos aos métodos de determinação do PBM nesses ecossistemas. O presente trabalho foi realizado com o objetivo de avaliar métodos de determinação do PBM em solo com diferentes coberturas vegetais. O trabalho consistiu da análise de amostras de Latossolo Vermelho-Amarelo distrófico muito argiloso (LVAd) localizado no município de Viçosa (MG), coletadas nas profundidades de 0 a 5 e 5 a 10 cm, em áreas com as seguintes coberturas vegetais: pínus (Pinus taeda), eucalipto (Eucalyptus grandis) e floresta nativa. Para determinação do P microbiano, foram empregados os métodos fumigação-extração (FE), irradiação com micro-ondas-extração (IE) e irradiação com micro-ondas-extração com membrana de troca aniônica (EMTA). em termos gerais, menores teores de PBM foram obtidos com o método irradiação-extração. Considerando a cobertura vegetal, foi detectada diferença significativa entre os três métodos sob floresta de eucalipto e floresta nativa, principalmente na camada superficial. Sob pínus, apenas o método IE diferiu dos demais, na camada subsuperficial. Menores coeficientes de variação (CV) foram obtidos com o FE, retratando maior precisão do método. Entretanto, o método IE mostrou-se, em termos operacionais, o mais adequado à determinação do PBM quando se tem maior número de amostras. Com relação às coberturas vegetais, a grande variabilidade observada nos CVs obtidos para cada cobertura, nos três métodos testados, inviabiliza a escolha de um único método que apresente maior precisão na avaliação do PBM.

Biomassa microbiana e atividade enzimática em solos do Estado de São Paulo sob vegetação nativa e cultivados

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Parâmetros microbiológicos no sistema de plantio direto e convencional em solos com diferentes teores de argila

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Influência da variação sazonal de atributos de fertilidade do solo sobre a biomassa microbiana em uma cronosseqüencia de plantio de palma de óleo (Elaeis oleifera (Kunth) Cortés) no Nordeste Paraense

O Brasil possui cerca de 32 milhões de hectares de áreas com aptidão para a expansão da cultura da palma de óleo e mais de 90% destas estão na Amazônia. Há necessidade de compreender a interação de novos plantios e de seu desenvolvimento com o meio ambiente. Este estudo foi conduzido em plantios comerciais da empresa Dendê do Pará SA (DENPASA) em Castanhal – Pará e avaliou o efeito da variação sazonal da precipitação sobre a fertilidade do solo e desta sobre a biomassa microbiana do solo em uma cronosseqüencia de plantios de palma de óleo com 5, 8 e 12 anos e floresta secundária. Foram mensurados/estimados e correlacionados os atributos do solo carbono da biomassa microbiana (C_BMS), carbono total (C_TOTAL), nitrogênio (N_TOTAL), respiração basal (RBS), quociente metabólico (qCO₂), relação carbono da biomassa microbiana/ carbono total (C_BMS:C_TOTAL), relação carbono/nitrogênio (C/N), umidade gravimétrica (Ug), fósforo (P), potássio (K), cálcio (Ca), magnésio (Mg), alumínio (Al), capacidade de troca efetiva de cátions (CTC) e saturação por alumínio (m). O C_BMS foi o atributo mais sensível para diferenciar as áreas do estudo e os períodos seco e chuvoso. Os atributos químicos de fertilidade do solo e a biomassa microbiana do solo apresentaram correlações significativas mais fortes e em maior número no período chuvoso. Os índices microbianos qCO₂ e C_BMS:C_TOTAL comprovaram que áreas de plantio convencional podem ser relativamente eficientes em relação à dinâmica do C em comparação a área de floresta secundária.

Matéria orgânica do solo em rotações de culturas sob sistema semeadura direta

Pós-graduação em Agronomia (Agricultura) - FCA

Qualidade física e química do solo em função do sistema de produção e da aplicação superficial de silicato e calcário em experimento de longa duração

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Nutrient additions affecting matter turnover in forest and pasture ecosystems

Nutrient inputs into ecosystems of the tropical mountain rainforest region are projected to further increase in the next decades. To investigate whether important ecosystem services such as nutrient cycling and matter turnover in native forests and pasture ecosystems show different patterns of response, two nutrient addition experiments have been established: NUMEX in the forest and FERPAST at the pasture. Both ecosystems already responded 1.5 years after the start of nutrient application (N, P, NP, Ca). Interestingly, most nutrients remained in the respective systems. While the pasture grass was co-limited by N and P, most tree species responded to P addition. Soil microbial biomass in the forest litter layer increased after NP fertilization pointing to nutrient co-limitation. In pasture soils, microorganisms were neither limited by N nor P. The results support the hypothesis that multiple and temporally variable nutrient limitations can coexist in tropical ecosystems.

Do improved pastures affect enzymatic activity and C and N dynamics in soils of the montado system ?

Vast montado areas are threatened by degradation, as the result of a long history of land use changes. Since improved pastures have been installed aiming soil quality improvement and system sustainability, it is crucial to evaluate the effects of these management changes on soil organic matter status and soil biological activity, as soil quality indicators. Therefore, a 35-yr old improved pasture and a natural pasture were studied, considering areas beneath tree canopy and in the open. Total organic C, total N, hot water soluble (HWS) and particulate (POM) C, microbial biomass C (MBC) and N (MBN), C mineralization rate (CMR) and net N mineralization rate (NMR) were determined. In addition, for a 1-yr period, soil β-glucosidase, urease, proteases and acid phosphomonoesterase were periodically determined. Improved pasture promoted the increase of soil C and N through POM-C increment, particularly beneath the trees canopies. The two study pastures did not show differences regarding soil microbial biomass, but variations in CMR, HWS-C and N availability (proteases and urease activities) suggest divergent soil microbial communities. Tree regulator role on C, N and P transformation processes in soil was confirmed

Below-ground processes in meadow soil under elevated ozone and carbon dioxide : - Greenhouse gas fluxes, N cycling and microbial communities

This thesis focuses on how elevated CO2 and/or O3 affect the below-ground processes in semi-natural vegetation, with an emphasis on greenhouse gases, N cycling and microbial communities. Meadow mesocosms mimicking lowland hay meadows in Jokioinen, SW Finland, were enclosed in open-top chambers and exposed to ambient and elevated levels of O3 (40-50 ppb) and/or CO2 (+100 ppm) for three consecutive growing season, while chamberless plots were used as chamber controls. Chemical and microbiological analyses as well as laboratory incubations of the mesocosm soils under different treatments were used to study the effects of O3 and/or CO2. Artificially constructed mesocosms were also compared with natural meadows with regards to GHG fluxes and soil characteristics. In addition to research conducted at the ecosystem level (i.e. the mesocosm study), soil microbial communities were also examined in a pot experiment with monocultures of individual species. By comparing mesocosms with similar natural plant assemblage, it was possible to demonstrate that artificial mesocosms simulated natural habitats, even though some differences were found in the CH4 oxidation rate, soil mineral N, and total C and N concentrations in the soil. After three growing seasons of fumigations, the fluxes of N2O, CH4, and CO2 were decreased in the NF+O3 treatment, and the soil NH4+-N and mineral N concentrations were lower in the NF+O3 treatment than in the NF control treatment. The mesocosm soil microbial communities were affected negatively by the NF+O3 treatment, as the total, bacterial, actinobacterial, and fungal PLFA biomasses as well as the fungal:bacterial biomass ratio decreased under elevated O3. In the pot survey, O3 decreased the total, bacterial, actinobacterial, and mycorrhizal PLFA biomasses in the bulk soil and affected the microbial community structure in the rhizosphere of L. pratensis, whereas the bulk soil and rhizosphere of the other monoculture, A. capillaris, remained unaffected by O3. Elevated CO2 caused only minor and insignificant changes in the GHG fluxes, N cycling, and the microbial community structure. In the present study, the below-ground processes were modified after three years of moderate O3 enhancement. A tentative conclusion is that a decrease in N availability may have feedback effects on plant growth and competition and affect the N cycling of the whole meadow ecosystem. Ecosystem level changes occur slowly, and multiplication of the responses might be expected in the long run.