Arbuscular mycorrhizal fungi associated with the rhizosphere of an endemic terrestrial bromeliad and a grass in the Brazilian neotropical dry forest.

Arbuscular mycorrhizal fungi form symbiotic associations with 80-90% of all known plants, allowing the fungi to acquire plant-synthesized carbon, and confer an increased capacity for nutrient uptake by plants, improving tolerance to abiotic and biotic stresses. We aimed at characterizing the mycorrhizal community in the rhizosphere of Neoglaziovia variegata (so-called `caroa`) and Tripogonella spicata (so-called resurrection plant), using high-throughput sequencing of the partial 18S rRNA gene. Both plants are currently undergoing a bioprospecting program to find microbes with the potential of helping plants tolerate water stress. Sampling was carried out in the Caatinga biome, a neotropical dry forest, located in northeastern Brazil. Illumina MiSeq sequencing of 37 rhizosphere samples (19 for N. variegata and 18 for T. spicata) revealed a distinct mycorrhizal community between the studied plants. According to alpha diversity analyses, T. spicata showed the highest richness and diversity based on the Observed ASVs and the Shannon index, respectively. On the other hand, N. variegata showed higher modularity of the mycorrhizal network compared to T. spicata. The four most abundant genera found (higher than 10%) were Glomus, Gigaspora, Acaulospora, and Scutellospora, with Glomus being the most abundant in both plants. Nonetheless, Gigaspora, Diversispora, and Ambispora were found only in the rhizosphere of N. variegata, whilst Scutellospora, Paraglomus, and Archaeospora were exclusive to the rhizosphere of T. spicata. Therefore, the community of arbuscular mycorrhizal fungi of the rhizosphere of each plant encompasses a unique composition, structure and modularity, which can differentially assist them in the hostile environment.

Impact of Water Shortage on Soil and Plant Attributes in the Presence of Arbuscular Mycorrhizal Fungi from a Harsh Environment.

Arbuscular mycorrhizal fungi (AMF) play a crucial role in plant health due to their ability to improve tolerance to biotic and abiotic stresses. Our aim was to evaluate the effectiveness of a pool of native AMF from a harsh environment on plant performance and changes in soil attributes under different levels of drought. An experiment using maize was established, varying the soil water content to simulate severe drought (30% of the water-holding capacity [WHC]), moderate (50% of the WHC) and no drought (80% of the WHC, control treatment). Soil and plant attributes were measured (enzyme activity, microbial biomass, AMF root colonisation and plant biomass and nutrient uptake). There was a two-fold increase in plant biomass under moderate drought when compared to no drought treatment, but there was no difference in nutrient uptake. Under severe drought, there were the highest enzyme activities related to phosphorus (P) cycling and P microbial biomass, indicating higher P microbial immobilization. The increase in AMF root colonisation was observed in plants under moderate and no drought. Our findings demonstrated that the better use of the AMF inoculum varied according to drought levels, with better performance under moderate drought due to the increase in plant biomass.

Potential of growth-promoting bacteria in maize (Zea mays L.) varies according to soil moisture.

Climate change has caused irregularities in water distribution, which affect the soil drying-wetting cycle and the development of economically important agricultural crops. Therefore, the use of plant growth-promoting bacteria (PGPB) emerges as an efficient strategy to mitigate negative impacts on crop yield. We hypothesized that the use of PGPB (in consortium or not) had potential to promote maize (Zea mays L.) growth under a soil moisture gradient in both non-sterile and sterile soils. Thirty PGPB strains were characterized for direct plant growth-promotion and drought tolerance induction mechanisms and were used in two independent experiments. Four soil water contents were used to simulate a severe drought (30% of field capacity [FC]), moderate drought (50% of FC), no drought (80% of FC) and, finally, a water gradient comprising the three mentioned soil water contents (80%, 50%, and 30% of FC). Two bacteria strains (BS28-7 Arthrobacter sp. and BS43 Streptomyces alboflavus), in addition to three consortia (BC2, BC4 and BCV) stood out in maize growth performance in experiment 1 and were used in experiment 2. Overall, under moderate drought, inoculation with BS43 surpassed the control treatment in root dry mass and nutrient uptake. Considering the water gradient treatment (80-50-30% of FC), the greatest total biomass was found in the uninoculated treatment when compared to BS28-7, BC2, and BCV. The greatest development of Z. mays L. was only observed under constant water stress conditions in the presence of PGPB. This is the first report that demonstrated the negative effect of individual inoculation of Arthrobacter sp. and the consortium of this strain with Streptomyces alboflavus on the growth of Z. mays L. based on a soil moisture gradient; however, future studies are needed for further validation.

Influence of temperature and soil moisture on the toxic potential of clothianidin to collembolan Folsomia candida in a tropical field soil.

Climate change can alter the toxic effects of pesticides on soil invertebrates. However, the nature and magnitude of the influence of climatic factors on clothianidin impacts in tropical soils are still unknown. The influence of increasing atmospheric temperature and the reduction in soil moisture on the toxicity and risk of clothianidin (seed dressing formulation Inside FS®) were assessed through chronic toxicity tests with collembolans Folsomia candida in a tropical field soil (Entisol). The risk of clothianidin for collembolans was estimated using the Toxicity-Exposure Ratio (TER) approach. Organisms were exposed to increasing clothianidin concentrations at 20, 25 and 27 °C in combination with two soil moisture conditions (30 and 60% of the maximum water holding capacity-WHC). The effect of temperature and soil moisture content on clothianidin toxicity was verified through the number of F. candida juveniles generated after 28 days of exposure to the spiked soil. The toxicities estimated at 25 °C (EC50_30%WHC = 0.014 mg kg-1; EC50_60%WHC = 0.010 mg kg-1) and 27 °C (EC50_30%WHC = 0.006 mg kg-1; EC50_60%WHC = 0.007 mg kg-1) were 2.9-3.0-fold (25 °C) and 4.3-6.7-fold (27 °C) higher than those found at 20 °C (EC50_30%WHC = 0.040 mg kg-1; EC50_60%WHC = 0.030 mg kg-1), indicating that clothianidin toxicity increases with temperature. No clear influence of soil moisture content on clothianidin toxicity could be observed once the EC50 values estimated at 30% and 60% WHC, within the same temperature, did not significantly differ. A significant risk was detected in all temperatures and soil moisture scenarios studied, and the TER values indicate that the risk can increase with increasing temperatures. Our results revealed that temperature could overlap with soil moisture in regulating clothianidin toxicity and reinforce the importance of including climatic factors in the prospective risk assessment of pesticides.

Are the Brazilian prevention values for copper and zinc in soils suitable for protecting earthworms against metal toxicity?

The current Brazilian copper (Cu) and zinc (Zn) prevention values (PV) for soil quality do not take into account the ecotoxicological impacts on soil organisms, which suggests these guiding values may not be protective of soil ecological trophic levels. This study assessed the acute (mortality) and chronic toxicity (reproduction), as well as the cumulative (bioaccumulation) potential of Cu and Zn (pseudo-total and available fractions) for earthworms Eisenia andrei in a Tropical Artificial Soil (TAS) and two tropical field soils (Oxisol and Alfisol). Toxicity data based on pseudo-total fractions were compared to PV. The Lowest Observed Effect Concentrations (LOEC) for the mortality endpoint were found at Cu and Zn concentrations higher than their PV (60 and 300 mg kg-1, respectively), regardless of the soil type. However, concentrations lower than PV reduced the reproduction of E. andrei by 20% (compared to the controls) for Cu in all tested soils (EC20s from 31.7 to 51.2 mg kg-1) and by 50% for Zn in Oxisol and Alfisol (EC50s = 225 and 283 mg kg-1, respectively). In TAS, only the EC20 (273 mg kg-1) for Zn was lower than PV. Increases of Cu in earthworm tissues occurred at concentrations higher than PV in all tested soils (LOEC values from 70 to 107 mg kg-1). The same was observed for Zn in TAS (LOEC = 497 mg kg-1), while in the field soils, the increases of Zn in earthworm tissues were lower than PV (LOEC = 131 and 259 mg kg-1 in Alfisol and Oxisol, respectively). We suggest the following: (1) The current Brazilian PV for Cu and Zn are not protective for earthworms (E. andrei) in the field soils tested; (2) PV derived from ecotoxicological assays in artificial soil cannot be representative for Brazilian field soils; (3) Using PV based on the pseudo-total fraction, without a soil-type normalizing factor, may limit the representativeness of this threshold for different soil types.

The use of sewage sludge as remediation for imidacloprid toxicity in soils.

This study investigated the influence of the sewage sludge (SS) soil amendment on the chronic toxicity of imidacloprid (through the seed dressing formulation MUCH 600 FS®-600 g active ingredient L-1) to collembolans Folsomia candida. Individuals 10-12 days old were exposed to two contrasting tropical soils (Oxisol and Entisol) amended with SS doses (0, 20, 40, 80, 160, and 320 g SS kg-1 soil; the SS doses have low intrinsic toxicity, which was checked before its application) in a full factorial combination with five imidacloprid concentrations (varying from 0.25 to 4 mg kg-1 in Oxisol and 0.03-0.5 mg kg-1 in Entisol) plus a control. None of the SS doses (without imidacloprid) in both soils reduced the number of generated juvenile collembolans. The imidacloprid concentrations reducing the collembolan reproduction in 50% (EC50) in Oxisol and Entisol without SS were 0.49 and 0.08 mg kg-1, respectively. However, the EC50 values generally increased with increasing SS doses in soils, varying from 1.03 to 1.41 in Oxisol and 0.07 to 0.21 in Entisol. The SS-amended soils showed 2.1- to 2.9-fold lower imidacloprid toxicity (EC50-based) in Oxisol and 1.8- to 2.7-fold lower toxicity in Entisol. Our results suggest the most effective SS doses alleviating the imidacloprid toxicity (EC50-based) to collembolans are 20 g kg-1 in Oxisol and 80 g kg-1 in Entisol. These results indicate that the tested SS has the potential to be employed as a soil amendment agent by reducing the toxicity of imidacloprid to the reproduction of F. candida.

Arbuscular mycorrhizal fungi community in soils under desertification and restoration in the Brazilian semiarid.

Soil desertification has a significant social, economic, and environmental impact worldwide. Mycorrhizal diversity remains poorly understood in semiarid regions impacted by desertification, especially in Brazilian drylands. More importantly, positive impacts of grazing exclusion on mycorrhizal communities are still incipient. Here, we hypothesized that overgrazing changes the structure of Arbuscular Mycorrhizal Fungi (AMF) community compared to native areas and, grazing exclusion is effective to restore the AMF community. Thus, we analyzed the status of AMF community in soils under desertification (overgrazing) and restoration (twenty-years of grazing exclusion) in the Brazilian semiarid. AMF-spores were extracted via humid decantation methodology, morphologically classified, and alpha diversity metrics were calculated. Soil samples were chemically, and physically characterized and multivariate statistical analyses were applied to verify the impact of soil degradation and restoration on AMF-community. Briefly, native, and restored areas presented higher contents of organic matter, phosphorus, microbial carbon, and ß-glucosidase activity. However, degraded soil showed higher Al3+, Na+, and bulk soil density values. The abundance of AMF spores was higher in restored soil, followed by degraded and native vegetation, and Shannon's diversity index was significantly higher in restored soils, followed by native vegetation. AMF-spores were classified into four families (Gigasporaceae > Acaulosporaceae > Glomeraceae > Ambisporaceae). Ambisporaceae was closed correlated with degraded soil, mainly with Al3+, Na+, and bulk soil density properties. On the other hand, Acaulosporaceae and Glomeraceae were positively correlated with native vegetation and restored soil, respectively, thereby improving Shannon index, richness, enzyme activity, and soil respiration. Thus, grazing exclusion, in long term, can be a good strategy to restore AMF-diversity in soils in the Brazilian semiarid.

Does Organomineral Fertilizer Combined with Phosphate-Solubilizing Bacteria in Sugarcane Modulate Soil Microbial Community and Functions?

Soil bacterial and fungal communities are suitable soil ecosystem health indicators due to their sensitivity to management practices and their role in soil ecosystem processes. Here, information on composition and functions of bacterial and fungal communities were evaluated at two phenological stages of sugarcane (six and twelve months, equivalent to the most intensive vegetative stage and to final maturation, respectively) when organomineral fertilizer, combined with phosphate-solubilizing bacteria (PSB), was added into the soil. Organic compost enriched with apatite (C + A) or phosphorite (C + P) and compost without phosphate enrichment (C) were used in the presence or absence of PSB. In addition, we used a control fertilized with soluble triple superphosphate. The differences were more related to the sampling period than to the type of organomineral fertilizer, being observed higher available phosphorus at six months than at twelve months. Only in the C treatment we observed the presence of Bacillaceae and Planococcaceae, while Pseudomonadaceae were only prevalent in inoculated C + A. As for fungi, the genera Chaetomium and Achroiostachys were only present in inoculated C + P, while the genus Naganishia was most evident in inoculated C + A and in uninoculated C + P. Soliccocozyma represented 75% of the total fungal abundance in uninoculated C while in inoculated C, it represented 45%. The bacterial community was more related to the degradation of easily decomposable organic compounds, while the fungal community was more related to degradation of complex organic compounds. Although the microbial community showed a resilient trait, subtle changes were detected in microbial community composition and function, and this may be related to the increase in yield observed.

Arbuscular Mycorrhizal Fungi and Soil Quality Indicators in Eucalyptus genotypes With Different Drought Tolerance Levels.

Silviculture has great importance worldwide, and the use of Eucalyptus species, which account for 75% of the local planted forest in Brazil, is one of the factors that contributes to the success of this activity in the country. Despite its adaptability, the yield of Eucalyptus is often affected by climate change, particularly water deficiency. Plants have developed strategies to mitigate water stress, for example, through their association with mycorrhizal fungi. The genus Eucalyptus, particularly in the plant domain, establishes symbioses with arbuscular mycorrhizal fungi (AMF) and ectomycorrhizal fungi (ECMF). The influence of Eucalyptus species on AMF and soil quality indicators is not well understood. Our aim was to conduct a preliminary evaluation of the various responses of soil AMF communities and soil nutrient dynamics in the presence of Eucalyptus species with different degrees of drought tolerance. A field experiment was established containing six Eucalyptus species, E. brassiana, E. camaldulensis, E. citriodora, E. cloeziana, E. grandis, and E. urophylla, all of which were planted in large plots. Soil and root samples were taken when the plants were 1.7 and 2.2 years old. We found that Eucalyptus species with low (E. grandis and E. urophylla) and intermediate drought tolerance (E. citriodora and E. cloeziana) showed stronger correlations with the AMF community than Eucalyptus species with high drought tolerance (E. brassiana and E. camaldulensis). Differences were also found between Eucalyptus species for AMF spore numbers and root colonization percentages, which was most evident for E. urophylla. The microbiological attributes found to be most responsive to Eucalyptus species were soil enzyme activities, AMF spore numbers, root colonization percentages, and fungal abundance. Soil organic carbon, phosphorus, potassium, zinc, copper, and iron were the main chemical drivers related to the soil AMF community structure in the presence of E. brassiana.

Toxicity of imidacloprid to collembolans in two tropical soils under different soil moisture.

Shifts in precipitation regimes due to the predicted climate changes can alter the water content in tropical soils and, consequently, may influence the toxicity of pesticides to soil fauna. This study assessed the influence of soil moisture content on the toxicity of the insecticide imidacloprid to the collembolans Folsomia candida in two tropical soils and evaluated the risk of this active ingredient for this species in the soils tested through the toxicity exposure ratio approach. Acute and chronic toxicity tests with F. candida were performed using an Entisol and an Oxisol. The soils were spiked with increasing imidacloprid concentrations while simulating normal water availability (60% of the water holding capacity [WHC]) and water restriction (30 or 45% WHC) for the tests. In the Oxisol, the reduction of soil moisture content significantly increased the toxic effects of imidacloprid on F. candida's survival (LC50 at 45% WHC = 23.8 vs. LC50 at 60% WHC >64 mg kg-1) and reproduction (effective concentration causing reductions in species reproduction of 50% [EC50] at 45% WHC = 0.32 vs. EC50 at 60% WHC = 2.83 mg kg-1), but in the Entisol no clear influence of the soil moisture on the toxicity of imidacloprid for collembolans was found. A significant risk for F. candida was observed in the Oxisol only when in water restriction, whereas in the Entisol it occurred regardless of soil moisture, suggesting that the imidacloprid hazard and risk for F. candida may be increased if soil moisture decreases due to climate changes, depending on the soil type.

How deep can ectomycorrhizas go? A case study on Pisolithus down to 4 meters in a Brazilian eucalypt plantation.

Despite the strong ecological importance of ectomycorrhizal (ECM) fungi, their vertical distribution remains poorly understood. To our knowledge, ECM structures associated with trees have never been reported in depths below 2 meters. In this study, fine roots and ECM root tips were sampled down to 4-m depth during the digging of two independent pits differing by their water availability. A meta-barcoding approach based on Illumina sequencing of internal transcribed spacers (ITS1 and ITS2) was carried out on DNA extracted from root samples (fine roots and ECM root tips separately). ECM fungi dominated the root-associated fungal community, with more than 90% of sequences assigned to the genus Pisolithus. The morphological and barcoding results demonstrated, for the first time, the presence of ECM symbiosis down to 4-m. The molecular diversity of Pisolithus spp. was strongly dependent on depth, with soil pH and soil water content as primary drivers of the Pisolithus spp. structure. Altogether, our results highlight the importance to consider the ECM symbiosis in deep soil layers to improve our understanding of fine roots functioning in tropical soils.

Ecotoxicological assessment of Fluazuron: effects on Folsomia candida and Eisenia andrei.

The cattle production in Brazil has increased considerably in the last years, mainly due to the control of parasite infestation of the animals, which cause loss of productivity to the sector. Fluazuron is an active ingredient (a.i.) of the benzoylurea class used to control ticks in cattle. As this a.i. has been found unchanged in animal feces, which may present a risk to edaphic organisms, this study aimed to assess the effects of fluazuron on survival, reproduction, and behavior of the soil invertebrates Folsomia candida and Eisenia andrei, through ecotoxicological assays. We carried out bioassays in a tropical artificial soil (TAS) spiked with increasing doses of the insecticide. Earthworm mortality was found only at the highest tested fluazuron concentration (LOEC = 160 mg a.i. kg-1 dry soil and NOEC = 80 mg kg-1), while the reproduction of F. candida and E. andrei was reduced at lower fluazuron concentrations (EC50 = 4.48 mg kg-1 and EC50 = 20.8 mg kg-1, respectively). Avoidance behavior was detected for both species at lower concentrations than those that caused impacts on reproduction, indicating that the substance may affect the soil habitat function. Since the possible adverse effects of fluazuron on edaphic fauna are still unknown or neglected, this study also warns about the possible harmful effect of veterinary pharmaceutical products on edaphic fauna.

Ecotoxicological impact of arsenic on earthworms and collembolans as affected by attributes of a highly weathered tropical soil.

High levels of heavy metals in soils may impose serious impacts on terrestrial organisms. In Brazil, the prevention values for evaluating the ecological risk of these elements are based only on soil chemical analyses and/or on data from ecotoxicological assays performed in soils of temperate regions. However, the attributes of the Brazilian highly-weathered tropical soils can influence the availability of heavy metals for soil fauna, resulting in different toxic values. To provide more accurate ecotoxicological risk values for arsenic (As) in tropical soils, we assessed the impacts of sodium arsenate (Na2HAsO4·7H2O) on the reproduction of earthworms (Eisenia andrei) and collembolans (Folsomia candida), as well as on As bioaccumulation and growth (weight loss) of E. andrei in a tropical artificial soil (TAS) and in an Oxisol. In TAS, As doses reduced the reproduction of the species and promoted weight loss of earthworms. On the other hand, the reproductions of the species as well as the earthworm growth were not altered by As in the Oxisol. The effective concentrations that reduce the reproduction of E. andrei and F. candida by 50 % (EC50) obtained in TAS (22.7 and 26.1 mg of As kg-1 of dry soil, respectively) were lower than those in the Oxisol (>135 mg kg-1, for both species). Although there was As bioaccumulation in earthworms in both soils, the internal concentrations in the earthworms were much higher in the oligochaetes exposed to arsenic in TAS. All these differences were attributed to the higher availability of As in the TAS, compared to the Oxisol, which increased the exposure of the species to the metal. The lower availability in the Oxisol was related to higher contents of type 1:1 silicate minerals and Fe and Al oxides and hydroxides, which strongly bind to As. These results highlight the importance of using tropical soils of humid regions to derive the Brazilian ecological risk prevention values for heavy metals, since the toxicity values are specific for these soils.

Soil macrofauna in organic and conventional coffee plantations in Brazil / Macrofauna do solo em plantações de café orgânico e convencional no Brasil

Abstract: Brazil has always been one of the most important coffee producing countries. Lately, there has equally been a renewed interest in alternative coffee production systems. The state of Espírito Santo is the second greatest coffee producer in Brazil; so, we used local coffee plantations to evaluate the relations between soil macrofauna and chemical and microbiological soil properties to identify which of these properties discriminate more effectively between the organic management system (OS) and the conventional management system (CS) of coffee plantations. For each of these two cultivation systems we chose three coffee farms who employed both cultivation systems and picked out the most similar fields from each property. At each site, first we sampled the litter at the soil surface. Afterwards, we sampled nine soil monoliths to evaluate the macrofauna, in summer and winter. We also collected nine supplemental soil samples, taken at a few centimeters from the soil monoliths, for chemical and microbiological analyses. Macrofauna density was evaluated by ANOVA and multivariate analysis. The chemical and microbiological properties are environmental variables, while the data on macrofauna are the explanatory variables. The total number of individuals recovered in this study was 3,354, and the climate, identified by the sampling season, was a great modulator of macrofauna, with higher numbers in winter. The principal components analysis showed that soil moisture, organic matter, nitrogen, phosphorus, boron, copper, pH, acid and alkaline phosphatases and microbial biomass carbon, were the most outstanding ones to discriminate both cultivation systems. We found no statistical significant differences in macrofauna density between OS and CS, probably due to a general great variability, since there was a tendency for much greater values in OS. We detected the interference of chemical and microbiological soil properties on the macrofauna community in both systems of coffee cultivation, and some results clearly correlated much better with climate data than with other factors. To our knowledge, this is the first time in which the data point to a clear separation between the more numerous and diversified soil macrofauna in coffee with organic cultivation from that with a conventional cultivation system.

Resumo: O Brasil sempre foi um dos mais importantes países produtores de café. Ultimamente, houve um renovado interesse pelos sistemas alternativos de produção de café. O Estado do Espírito Santo é o segundo maior produtor de café do Brasil; então, utilizaram-se plantações locais de café para avaliar as relações entre a macrofauna do solo e as propriedades químicas e microbiológicas do solo para identificar quais dessas propriedades discriminam mais efetivamente entre o sistema de manejo orgânico (SO) e o sistema de manejo convencional (SC) das plantações de café. Para cada um desses dois sistemas de cultivo, escolheu-se três fazendas de café que empregaram ambos os sistemas de cultivo e selecionaram os campos mais similares de cada propriedade. Em cada local, primeiro amostrou-se a serrapilheira do solo. Posteriormente, amostraram-se nove monólitos de solo para avaliar a macrofauna, tanto no verão como no inverno. Também, foram coletadas nove amostras suplementares de solo, a alguns centímetros dos monólitos de solo, para análises químicas e microbiológicas. A densidade da macrofauna foi avaliada por ANOVA e análise multivariada. As propriedades químicas e microbiológicas são variáveis ambientais, enquanto os dados sobre a macrofauna são as variáveis explicativas. O número total de indivíduos recuperados neste estudo foi de 3.354 e o clima, identificado pela época de amostragem, foi um ótimo modulador da macrofauna, com maiores números no inverno. A análise dos componentes principais mostrou que a umidade do solo, matéria orgânica, nitrogênio, fósforo, boro, cobre, pH, fosfatases ácido e alcalina e carbono da biomassa microbiana, foram os mais destacados para discriminar ambos os sistemas de cultivo. Encontraram-se diferenças significativas na densidade da macrofauna entre SO e SC, provavelmente devido a uma grande variabilidade geral, uma vez que houve uma tendência para valores muito maiores no SO. Detectou-se a interferência das propriedades químicas e microbiológicas do solo na comunidade de macrofauna em ambos os sistemas de cultivo de café, e alguns resultados claramente correlacionados muito melhor com os dados climáticos do que com outros fatores. A nosso conhecimento, esta é a primeira vez em que os dados apontam para uma clara separação entre a macrofauna do solo mais numerosa e diversificada em café com cultivo orgânico de que com um sistema de cultivo convencional.

Shifts in the bacterial community composition along deep soil profiles in monospecific and mixed stands of Eucalyptus grandis and Acacia mangium.

Our knowledge of the rhizosphere bacterial communities in deep soils and the role of Eucalyptus and Acacia on the structure of these communities remains very limited. In this study, we targeted the bacterial community along a depth profile (0 to 800 cm) and compared community structure in monospecific or mixed plantations of Acacia mangium and Eucalyptus grandis. We applied quantitative PCR (qPCR) and sequence the V6 region of the 16S rRNA gene to characterize composition of bacterial communities. We identified a decrease in bacterial abundance with soil depth, and differences in community patterns between monospecific and mixed cultivations. Sequence analysis indicated a prevalent effect of soil depth on bacterial communities in the mixed plant cultivation system, and a remarkable differentiation of bacterial communities in areas solely cultivated with Eucalyptus. The groups most influenced by soil depth were Proteobacteria and Acidobacteria (more frequent in samples between 0 and 300 cm). The predominant bacterial groups differentially displayed in the monospecific stands of Eucalyptus were Firmicutes and Proteobacteria. Our results suggest that the addition of an N2-fixing tree in a monospecific cultivation system modulates bacterial community composition even at a great depth. We conclude that co-cultivation systems may represent a key strategy to improve soil resources and to establish more sustainable cultivation of Eucalyptus in Brazil.

Ecotoxicological evaluation of swine manure disposal on tropical soils in Brazil.

Swine production in Brazil results in a great volume of manure that normally is disposed of as agricultural fertilizer. However, this form of soil disposal, generally on small farms, causes the accumulation of large amounts of manure and this results in contaminated soil and water tables. To evaluate the effects of increasing concentrations of swine manure on earthworms, several ecotoxicological tests were performed using Eisenia andrei as test organism in different tropical soils, classified respectively as Ultisol, Oxisol, and Entisol, as well as Tropical Artificial Soil (TAS). The survival, reproduction and behavior of the earthworms were evaluated in experiments using a completely randomized design, with five replications. In the Ultisol, Oxisol and TAS the swine manure showed no lethality, but in the Entisol it caused earthworm mortality (LOEC=45 m(3)ha(-1)). In the Entisol, the waste reduced the reproductive rate and caused avoidance behavior in E. andrei (LOEC=30 m(3)ha(-1)) even in lower concentrations. The Entisol is extremely sandy, with low cation exchange capacity (CEC), and this may be the reason for the higher toxicity on soil fauna, with the soil not being able to hold large amounts of pollutants (e.g. toxic metals), but leaving them in bioavailable forms. These results should be a warning of the necessity to consider soil parameters (e.g. texture and CEC) when evaluating soil contamination by means of ecotoxicological assays, as there still are no standards for natural soils in tropical regions. E. andrei earthworms act as indicators for a soil to support disposal of swine manure without generating harm to agriculture and ecosystems.

Soil properties discriminating Araucaria forests with different disturbance levels.

Soil biological, chemical, and physical properties can be important for monitoring soil quality under one of the most spectacular vegetation formation on Atlantic Forest Biome, the Araucaria Forest. Our aim was to identify a set of soil variables capable of discriminating between disturbed, reforested, and native Araucaria forest soils such that these variables could be used to monitor forest recovery and maintenance. Soil samples were collected at dry and rainy season under the three forest types in two state parks at São Paulo State, Brazil. Soil biological, chemical, and physical properties were evaluated to verify their potential to differentiate the forest types, and discriminant analysis was performed to identify the variables that most contribute to the differentiation. Most of physical and chemical variables were sensitive to forest disturbance level, but few biological variables were significantly different when comparing native, reforested, and disturbed forests. Despite more than 20 years following reforestation, the reforested soils were chemically and biologically distinct from native and disturbed forest soils, mainly because of the greater acidity and Al3+ content of reforested soil. Disturbed soils, in contrast, were coarser in texture and contained greater concentrations of extractable P. Although biological properties are generally highly sensitive to disturbance and amelioration efforts, the most important soil variables to discriminate forest types in both seasons included Al3+, Mg2+, P, and sand, and only one microbial attribute: the NO2- oxidizers. Therefore, these five variables were the best candidates, of the variables we employed, for monitoring Araucaria forest disturbance and recovery.

Indole-3-acetic acid producing root-associated bacteria on growth of Brazil Pine (Araucaria angustifolia) and Slash Pine (Pinus elliottii).

Araucaria forests in Brazil today correspond to only 0.7 % of the original 200 km(2) of natural forest that covered a great part of the southern and southeastern area of the Atlantic Forest and, although Araucaria angustifolia is an endangered species, illegal exploitation is still going on. As an alternative to the use of hardwoods, Pinus elliottii presents rapid growth and high tolerance to climatic stress and low soil fertility or degraded areas. Thus, the objective of this study was to evaluate the effect of IAA-producing bacteria on the development of A. angustifolia and P. elliottii. We used five bacterial strains previously isolated from the rhizosphere of A. angustifolia, which produce quantities of IAA ranging from 3 to 126 µg mL(-1). Microbiolized seeds were sown in a new gnotobiotic system developed for this work, that allowed the quantification of the plant hormone IAA produced by bacteria, and the evaluation of its effect on seedling development. Also, it was shown that P. elliottii roots were almost as satisfactory as hosts for these IAA producers as A. angustifolia, while different magnitudes of mass increases were found for each species. Thus, we suggest that these microbial groups can be helpful for the development and reestablishment of already degraded forests and that PGPR isolated from Araucaria rhizosphere have the potential to be beneficial in seedling production of P. elliottii. Another finding is that our newly developed gnotobiotic system is highly satisfactory for the evaluation of this effect.

Plant growth promoting bacteria in Brachiaria brizantha.

Brachiaria brizantha is considered one of the preferred fodders among farmers for having high forage yield and large production of root mass. The association of beneficial bacteria with these grasses can be very valuable in the recovery of the pasture areas with nutritional deficiency. With the aim of studying this possibility, we carried out the sampling of soil and roots of B. brizantha in three areas (Nova Odessa-SP, São Carlos-SP and Campo Verde-MT, Brazil). Seventy-two bacterial strains were isolated and used in tests to evaluate their biotechnological potential. Almost all isolates presented at least one positive feature. Sixty-eight isolates produced analogues of indole-3-acetic acid, ten showed nitrogenase activity when subjected to the method of increasing the concentration of total nitrogen (total N) in the culture medium and sixty-five isolates showed nitrogenase activity when subjected to acetylene reduction technique. The partial sequencing of 16S rRNA of these isolates allowed the identification of seven main groups, with the prevalence of those affiliated to the genus Stenotrophomonas (69 %). At the end, this work elected the strains C4 (Pseudomonadaceae) and C7 (Rhodospirillaceae) as promising organisms for the development of inoculants due to their higher nitrogenase activity.

Isolation, selection and characterization of root-associated growth promoting bacteria in Brazil Pine (Araucaria angustifolia).

Araucaria angustifolia, a unique species of this genus that occurs naturally in Brazil, has a high socio-economic and environmental value and is critically endangered of extinction, since it has been submitted to intense predatory exploitation during the last century. Root-associated bacteria from A. angustifolia were isolated, selected and characterized for their biotechnological potential of growth promotion and biocontrol of plant pathogenic fungi. Ninety-seven strains were isolated and subjected to chemical tests. All isolates presented at least one positive feature, characterizing them as potential PGPR. Eighteen isolates produced indole-3-acetic acid (IAA), 27 were able to solubilize inorganic phosphate, 21 isolates were presumable diazotrophs, with pellicle formation in nitrogen-free culture medium, 83 were phosphatases producers, 37 were positive for siderophores and 45 endospore-forming isolates were antagonistic to Fusarium oxysporum, a pathogen of conifers. We also observed the presence of bacterial strains with multiple beneficial mechanisms of action. Analyzing the fatty acid methyl ester (FAME) and partial sequencing of the 16S rRNA gene of these isolates, it was possible to characterize the most effective isolates as belonging to Bacillaceae (9 isolates), Enterobacteriaceae (11) and Pseudomonadaceae (1). As far as we know, this is the first study to include the species Ewingella americana as a PGPR.