Fungal succession on the decomposition of three plant species from a Brazilian mangrove.

Leaf decomposition is the primary process in release of nutrients in the dynamic mangrove habitat, supporting the ecosystem food webs. On most environments, fungi are an essential part of this process. However, due to the peculiarities of mangrove forests, this group is currently neglected. Thus, this study tests the hypothesis that fungal communities display a specific succession pattern in different mangrove species and this due to differences in their ecological role. A molecular approach was employed to investigate the dynamics of the fungal community during the decomposition of three common plant species (Rhizophora mangle, Laguncularia racemosa, and Avicennia schaueriana) from a mangrove habitat located at the southeast of Brazil. Plant material was the primary driver of fungi communities, but time also was marginally significant for the process, and evident changes in the fungal community during the decomposition process were observed. The five most abundant classes common to all the three plant species were Saccharomycetes, Sordariomycetes, Tremellomycetes, Eurotiomycetes, and Dothideomycetes, all belonging to the Phylum Ascomycota. Microbotryomycetes class were shared only by A. schaueriana and L. racemosa, while Agaricomycetes class were shared by L. racemosa and R. mangle. The class Glomeromycetes were shared by A. schaueriana and R. mangle. The analysis of the core microbiome showed that Saccharomycetes was the most abundant class. In the variable community, Sordariomycetes was the most abundant one, mainly in the Laguncularia racemosa plant. The results presented in this work shows a specialization of the fungal community regarding plant material during litter decomposition which might be related to the different chemical composition and rate of degradation.

Ultrastructural and metabolic disorders induced by short-term cadmium exposure in Avicennia schaueriana plants and its excretion through leaf salt glands.

Environmental cadmium (Cd) sources have increased in mangrove sediments in recent decades, inducing cellular damage to many plants. Avicennia schaueriana is abundant in mangrove sites and has been subject to Cd contamination. The possible effects of Cd toxicity and the structural and physiological disturbances to this plant were studied. Can this plant express early cellular tolerance mechanisms to such metal contamination? Seedlings of A. schaueriana were collected from sites of their natural occurrence, placed in plastic pots containing nutrient solution for 60 days, and subsequently exposed to increasing Cd concentrations for 5 days under experimental conditions. The anatomical, ultrastructural and physiological changes induced by Cd were analysed. Cd accumulated mainly in the root system and in pneumatophores, stems and leaves, induced differential accumulation of mineral nutrients, but did not induce necrosis or changes in leaf anatomy. However, there was a decrease in starch grains and an increase in deposited electron-dense material in the cortex and vascular bundles. Cd induced both increases in calcium (Ca) content in shoots and Ca oxalate crystal precipitation in leaf mesophyll and was detected in crystals and in the secretion of salt glands. Our observations and experimental results provide evidence of Cd tolerance in A. schaueriana. As a new feature, despite the clear cellular physiological disorders, this plant is able to eliminate Cd through leaf salt glands and immobilise it in Ca crystals, representing fast mechanisms for Cd exclusion and complexation in leaves in heavy metal coastal polluted marine ecosystems.

Petroleum contamination and bioaugmentation in bacterial rhizosphere communities from Avicennia schaueriana

ABSTRACT Anthropogenic activity, such as accidental oil spills, are typical sources of urban mangrove pollution that may affect mangrove bacterial communities as well as their mobile genetic elements. To evaluate remediation strategies, we followed over the time the effects of a petroleum hydrocarbon degrading consortium inoculated on mangrove tree Avicennia schaueriana against artificial petroleum contamination in a phytoremediation greenhouse experiment. Interestingly, despite plant protection due to the inoculation, denaturing gradient gel electrophoresis of the bacterial 16S rRNA gene fragments amplified from the total community DNA indicated that the different treatments did not significantly affect the bacterial community composition. However, while the bacterial community was rather stable, pronounced shifts were observed in the abundance of bacteria carrying plasmids. A PCR-Southern blot hybridization analysis indicated an increase in the abundance of IncP-9 catabolic plasmids. Denaturing gradient gel electrophoresis of naphthalene dioxygenase (ndo) genes amplified from cDNA (RNA) indicated the dominance of a specific ndo gene in the inoculated petroleum amendment treatment. The petroleum hydrocarbon degrading consortium characterization indicated the prevalence of bacteria assigned to Pseudomonas spp., Comamonas spp. and Ochrobactrum spp. IncP-9 plasmids were detected for the first time in Comamonas sp. and Ochrobactrum spp., which is a novelty of this study.

Short-term cadmium exposure induces gas exchanges, morphological and ultrastructural disturbances in mangrove Avicennia schaueriana young plants.

Mangroves have been subject to more metal contamination, including cadmium (Cd). This study evaluated if a relatively short Cd exposure may induce metabolic, morphological and ultrastructural cell disturbance in Avicennia schaueriana. Cd induced evident constraints to seedlings since there was reduction in leaf gas exchanges and the plants did not survive for more than 10 days at a higher Cd exposure in controlled conditions. The highest Cd accumulation was observed in roots and gradually less in stem and leaves. Cadmium induced lignin deposition was observed in xylem cells of all vegetative organs. Intense sclerification in xylem cells, endoderm and change in the hypoderm organization were also detected. Cadmium clearly induced chloroplast deformities with ruptures of its membranes, thylakoids and core and provoked cytoplasm disorganization. These metal constraints under natural conditions for long term can lead to the accumulation of cellular and metabolic damages and jeopardize seedlings establishment and local biodiversity.

Petroleum contamination and bioaugmentation in bacterial rhizosphere communities from Avicennia schaueriana.

Anthropogenic activity, such as accidental oil spills, are typical sources of urban mangrove pollution that may affect mangrove bacterial communities as well as their mobile genetic elements. To evaluate remediation strategies, we followed over the time the effects of a petroleum hydrocarbon degrading consortium inoculated on mangrove tree Avicennia schaueriana against artificial petroleum contamination in a phytoremediation greenhouse experiment. Interestingly, despite plant protection due to the inoculation, denaturing gradient gel electrophoresis of the bacterial 16S rRNA gene fragments amplified from the total community DNA indicated that the different treatments did not significantly affect the bacterial community composition. However, while the bacterial community was rather stable, pronounced shifts were observed in the abundance of bacteria carrying plasmids. A PCR-Southern blot hybridization analysis indicated an increase in the abundance of IncP-9 catabolic plasmids. Denaturing gradient gel electrophoresis of naphthalene dioxygenase (ndo) genes amplified from cDNA (RNA) indicated the dominance of a specific ndo gene in the inoculated petroleum amendment treatment. The petroleum hydrocarbon degrading consortium characterization indicated the prevalence of bacteria assigned to Pseudomonas spp., Comamonas spp. and Ochrobactrum spp. IncP-9 plasmids were detected for the first time in Comamonas sp. and Ochrobactrum spp., which is a novelty of this study.

Differential bioaccumulation and translocation patterns in three mangrove plants experimentally exposed to iron. Consequences for environmental sensing.

Avicennia schaueriana, Laguncularia racemosa and Rhizophora mangle were experimentally exposed to increasing levels of iron (0, 10, 20 and 100 mg L(-1) added Fe(II) in Hoagland's nutritive medium). The uptake and translocation of iron from roots to stems and leaves, Fe-secretion through salt glands (Avicennia schaueriana and Laguncularia racemosa) as well as anatomical and histochemical changes in plant tissues were evaluated. The main goal of this work was to assess the diverse capacity of these plants to detect mangroves at risk in an area affected by iron pollution (Vitoria, Espírito Santo, Brazil). Results show that plants have differential patterns with respect to bioaccumulation, translocation and secretion of iron through salt glands. L. racemosa showed the best environmental sensing capacity since the bioaccumulation of iron in both Fe-plaque and roots was higher and increased as the amount of added-iron rose. Fewer changes in translocation factors throughout increasing added-iron were observed in this species. Furthermore, the amount of iron secreted through salt glands of L. racemosa was strongly inhibited when exposed to added-iron. Among three studied species, A. schaueriana showed the highest levels of iron in stems and leaves. On the other hand, Rhizophora mangle presented low values of iron in these compartments. Even so, there was a significant drop in the translocation factor between aerial parts with respect to roots, since the bioaccumulation in plaque and roots of R. mangle increased as iron concentration rose. Moreover, rhizophores of R. mangle did not show changes in bioaccumulation throughout the studied concentrations. So far, we propose L. racemosa as the best species for monitoring iron pollution in affected mangroves areas. To our knowledge, this is the first detailed report on the response of these plants to increasing iron concentration under controlled conditions, complementing existing data on the behavior of the same plants under field exposure.

Antifungal potential of Avicennia schaueriana Stapf & Leech. (Acanthaceae) against Cladosporium and Colletotrichum species.

UNLABELLED: There is significant interest in research to develop plant extracts with fungicidal activities that are less harmful to the environment and human health than synthetic fungicides. This study aimed to evaluate the antifungal activity of the extracts of Avicennia schaueriana against Colletotrichum and Cladosporium species and to identify the compounds responsible for the activity. Leaves and stems of A. schaueriana were extracted with ethanol and partitioned with petroleum ether, chloroform and ethyl acetate. The antifungal activity of such extracts was tested by bioautography against Cladosporium sphaerospermum, Cladosporium cladosporioides and Colletotrichum lagenarium. Ethanolic extracts, petroleum ether and chloroform fractions of stems had the highest antifungal activity with several active bands (Rf = 0·72 and Rf = 0·55). In the agar dilution assay, ethanolic extract, petroleum ether and chloroform fractions of stems were the most efficacious, presenting 85, 62 and 63% growth inhibition of Colletotrichum gloeopsporioides and minimum inhibitory concentration values between 1 and 1·5 mg ml(-1) , respectively. Analysis carried out using gas chromatography coupled to a mass spectrometry of petroleum ether and chloroform fractions allowed the identification of fatty acids methyl esters, lupeol and naphthoquinones such as lapachol, α-lapachone, naphtho[2,3-b]furan-4,9-dione, 2-isopropyl- and avicenol-C. We may infer that the antifungal activity of A. schaueriana is due to the abundance of these compounds. SIGNIFICANCE AND IMPACT OF THE STUDY: This study shows that Avicennia schaueriana extracts have a high potential for the growth inhibition of Colletotrichum and Cladosporium ssp. and will provide a starting point for discovering new natural products with antifungal activity. Their development is of particular interest to organic production systems where synthetic fungicides cannot be used.

Effect of pollution by particulate iron on the morphoanatomy, histochemistry, and bioaccumulation of three mangrove plant species in Brazil.

In Brazil, some mangrove areas are subjected to air pollution by particulate iron from mining activities. However, the effect of this pollutant on mangrove plants is not well known. This study aimed to comparatively analyze the morphoanatomy, histochemistry, and iron accumulation in leaves of Avicennia schaueriana, Laguncularia racemosa, and Rhizophora mangle. Samples were collected from five mangrove sites of Espírito Santo state, each of which is exposed to different levels of particulate iron pollution. The amount of particulate material settled on the leaf surface was greater in A. schaueriana and L. racemosa, which contain salt glands. High iron concentrations were found in leaves of this species, collected from mangrove areas with high particulate iron pollution, which suggests the foliar absorption of this element. None of the samples from any of the sites showed morphological or structural damage on the leaves. Scanning electron microscopy (SEM) coupled to X-ray diffraction rendered a good method for evaluating iron on leaves surfaces. A histochemical test using Prussian blue showed to be an appropriate method to detect iron in plant tissue, however, proved to be an unsuitable method for the assessment of the iron bioaccumulation in leaves of A. schaueriana and R. mangle. So far, this study demonstrates the need of evaluating the pathway used by plants exposed to contaminated particulate matter to uptake atmospheric pollutants.

Nutrient removal in a closed silvofishery system using three mangrove species (Avicennia germinans, Laguncularia racemosa, and Rhizophora mangle).

The removal of ammonium (NH4(+)), nitrite (NO2(-)), nitrate (NO3(-)), and phosphate (PO4(-3)) in a closed silvofishery system was examined using three mangrove species (i.e., Avicennia germinans, Laguncularia racemosa, and Rhizophora mangle). Specifically, six closed tanks were installed for this experiment with a population of 60 Dormitator latifrons fishes per tank. We planted 40 seedlings in each of three experimental tanks separated by species, while the remaining tanks were used as control. During 15 weeks, nutrient concentrations among the three mangrove systems presented no significant differences (P>0.05). However, nutrient removal variability was minimum during the last 2-5 weeks. Mangroves presented an average efficiency of 63% for the removal of NH4(+) and NO2(-). Contrary, the average removal potential of NO3(-) and PO4(-3) was 50%. Results from this study suggest that the three mangrove species could be used in a closed silvofishery systems for the biological removal of NH4(+), NO2(-), NO3(-), and PO4(-3).

Cell viability and leakage of electrolytes in Avicennia germinans exposed to heavy metals.

The effect of heavy metal stress on the cell viability and leakage of electrolytes of Avicennia germinans leaf discs was investigated by the tissue tolerance test. Foliar discs were incubated with different Cd2+ or CU2+ concentrations for 24 h; thereafter, the cell membrane stability of the tissue was assayed by the cell viability Evans blue and leakage electrolytes methods. The results indicated that electrolyte leakage of the leaf discs increased 24 h after exposure to heavy metal stress, as shown by a reduction of the cell viability by 30% in discs exposed to higher doses of Cd2+ (0.546 M) and Cu2+ (0.7 M), respectively. Additionally, the histological analysis of the leaf discs exposed to heavy metal stress revealed that at higher Cd2+ and/or Cu2+ concentrations an increase in the intercellular spaces and destruction of mesophyll cells was observed 24 h after exposure. In summary, the biochemical and structural changes observed in foliar tissues of A. germinans suggest that higher cadmium and copper concentrations may result in structural changes and altered physiological characters in leaves.

The glutathione antioxidant system as a biomarker suite for the assessment of heavy metal exposure and effect in the grey mangrove, Avicennia marina (Forsk.) Vierh.

Alterations in the glutathione antioxidant system and lipid peroxidation in Avicennia marina were studied under laboratory and field conditions. The activity of glutathione peroxidase (GPx) was found to respond to Zn exposure, and a significant positive relationship between leaf Zn concentration and GPx activity was observed after 96 h and 8 weeks. Lipid hydroperoxides increased proportionally with increasing leaf Zn concentration after 2 and 8 weeks, while no changes in total glutathione were observed. Induction of GPx at 96 h predicted effects at the individual level at a later time interval (reduced biomass at 8 weeks). Results from the field revealed that increasing leaf metal concentration (Zn, Cu or Pb) produced a proportional increase in GPx activity whereas lipid hydroperoxides and total glutathione were not affected. The utility of GPx as an early warning biomarker is suggested, since GPx activity increases in a dose-dependant fashion in response to accumulated leaf metals, and is predictive of later effects on growth.

Dial organic carbon fluctuations in a mangrove tidal creek in Sepetiba bay, Southeast Brazil

The carbon (C) concentration and flux, as dissolved organic carbon (DOC), particulate organic carbon (POC) and macrodetritus (MD), were quantified through 4 tidal cycles in a mangrove tidal creek in Southeastern Brazil. DOC was the major fraction of the total C concentration, accounting for 68 and 61 percent of the total C concentration during ebb and flood periods respectively. Concentrations of DOC (Ebb = 3,41 ± 0,57 mgC.L-1 and Flood = 3,55 ± 0,76 mgC.L-1) and POC (Ebb = 1,73 ± 0,99 mgC.L-1 and Flood = 1,28 ± 0,45 mgC.L-1) were relatively similar during the four tidal cycles. Macrodetritus presented a wide variation with concentration peaks probably related to external forces, such as winds, which enrich the ebb flow with leaf litter. DOC and POC fluxes depended primarily on tidal and net water fluxes, whereas MD fluxes were not. The magnitude of the DOC and POC fluxes varied with the area flooded at high tide, but not the MD fluxes. DOC was the major form of carbon export to Sepetiba Bay. During the four tidal cycles, the forest exported a total of 1,2 kg of organic carbon per ha, mostly as DOC (60 percent), followed by POC (22 percent) and MD (18 percent).

As concentrações e fluxos de carbono orgânico sob forma de carbono orgânico dissolvido (COD), carbono orgânico particulado (COP) e macrodetritos (MD) foram quantificadas durante 4 ciclos de maré em canal de maré na Floresta Experimental de Itacuruçá, Baía de Sepetiba, RJ, litoral sudeste do Brasil. COD foi a fração mais importante para a concentração total de carbono orgânico, contribuindo com 68 e 61 por cento da concentração total de C nos períodos de maré vazante e enchente, respectivamente. As concentrações de COD (vazante = 3.41 ± 0.57 mgC.L-1 e enchente = 3.55 ± 0.76 mgC.L-1) e COP (vazante = 1.73 ± 0.99 mgC.L-1 e enchente = 1.28 ± 0.45 mgC.L-1) foram similares durante os 4 ciclos de maré. A fração macrodetritos apresentou uma ampla variabilidade com máximos de concentração relacionados a fatores externos como ventos, que enriqueceram as águas de vazante com macrodetritos. A magnitude dos fluxos de COD e COP, mas não os de macrodetritos, relacionaram-se com os fluxos de água e a conseqüente área inundada pela maré. A fração COD foi a mais importante forma de exportação de carbono orgânico pelo manguezal. Durante os 4 ciclos monitorados, a floresta exportou um total de 1.2 kg de carbono orgânico, 60 por cento sob forma de COD, seguido pelo COP (22 por cento) e pela fração macrodetritos (18 por cento).

Coordinated responses of phytochelatin synthase and metallothionein genes in black mangrove, Avicennia germinans, exposed to cadmium and copper.

To evaluate the role of phytochelatins and metallothioneins in heavy metal tolerance of black mangrove Avicennia germinans, 3-month-old seedlings were exposed to cadmium or copper for 30 h, under hydroponic conditions. Degenerate Mt2 and PCS primers were synthesized based on amino acid and nucleotide alignment sequences reported for Mt2 and PCS in other plant species found in GenBank. Total RNA was isolated from A. germinans leaves and two partial fragments of metallothionein and phytochelatin synthase genes were isolated. Gene expression was evaluated with reverse transcripatase-polymerase chain reaction (RT-PCR) amplification technique. Temporal analysis showed that low Cd2+ and Cu2+ concentrations caused a slight (but not significant) increase in AvMt2 expression after a 16 h exposure time, while AvPCS expression showed a significant increase under the same conditions but only after 4h. Results strongly suggest that the rapid increase in AvPCS expression may contribute to Cd2+ and Cu2+ detoxification. Moreover, we found that A. germinans has the capacity to over-express both genes (AvMt2 and AvPCS), which may constitute a coordinated detoxification response mechanism targeting non-essential metals. Nonetheless, our results confirm that AvPCS was the most active gene involved in the regulation of essential metals (e.g., Cu2+) in A. germinans leaves.

Dial organic carbon fluctuations in a mangrove tidal creek in Sepetiba bay, Southeast Brazil.

The carbon (C) concentration and flux, as dissolved organic carbon (DOC), particulate organic carbon (POC) and macrodetritus (MD), were quantified through 4 tidal cycles in a mangrove tidal creek in Southeastern Brazil. DOC was the major fraction of the total C concentration, accounting for 68 and 61% of the total C concentration during ebb and flood periods respectively. Concentrations of DOC (Ebb = 3.41 +/- 0.57 mgC.L-1 and Flood = 3.55 +/- 0.76 mgC.L-1) and POC (Ebb = 1.73 +/- 0.99 mgC.L-1 and Flood = 1.28 +/- 0.45 mgC.L-1) were relatively similar during the four tidal cycles. Macrodetritus presented a wide variation with concentration peaks probably related to external forces, such as winds, which enrich the ebb flow with leaf litter. DOC and POC fluxes depended primarily on tidal and net water fluxes, whereas MD fluxes were not. The magnitude of the DOC and POC fluxes varied with the area flooded at high tide, but not the MD fluxes. DOC was the major form of carbon export to Sepetiba Bay. During the four tidal cycles, the forest exported a total of 1.2 kg of organic carbon per ha, mostly as DOC (60%), followed by POC (22%) and MD (18%).

The influence of radical architecture on cadmium bioaccumulation in the black mangrove, Avicennia germinans L.

Two groups of Avicennia germinans plants with differences in the radical architecture were exposed under hydroponic conditions to 95ppm of cadmium (Cd) for a period of 24h. Later, Cd concentration in roots, stems and leaves was determined by graphite furnace atomic absorption spectrophotometry. Our results showed that, for both groups of plants, the roots accumulated higher concentration of Cd as compared to stems and leaves, though, the plants of group B displayed enhanced radical architecture, better growth performance, and lower Cd concentration as compared to plants of group A. In contrast, low values of leaves/roots Cd transportation index, and bioaccumulation factor were found in plants of group B. These results suggest that the higher radical architecture developed in plants of group B might better adjust the uptake of Cd as a result of an integrated network of multiple response processes for instances, production of organic acids, antioxidative replay, cell-wall lignification and/or suberization. Further studies will be focused in understanding the role of the radical system in mangrove plants with the rhizosphere activation and root adsorption to soil Cd under natural conditions.

Nitrogen limitation of growth and nutrient dynamics in a disturbed mangrove forest, Indian River Lagoon, Florida.

The objectives of this study were to determine effects of nutrient enrichment on plant growth, nutrient dynamics, and photosynthesis in a disturbed mangrove forest in an abandoned mosquito impoundment in Florida. Impounding altered the hydrology and soil chemistry of the site. In 1997, we established a factorial experiment along a tree-height gradient with three zones, i.e., fringe, transition, dwarf, and three fertilizer treatment levels, i.e., nitrogen (N), phosphorus (P), control, in Mosquito Impoundment 23 on the eastern side of Indian River. Transects traversed the forest perpendicular to the shoreline, from a Rhizophora mangle-dominated fringe through an Avicennia germinans stand of intermediate height, and into a scrub or dwarf stand of A. germinans in the hinterland. Growth rates increased significantly in response to N fertilization. Our growth data indicated that this site is N-limited along the tree-height gradient. After 2 years of N addition, dwarf trees resembled vigorously growing saplings. Addition of N also affected internal dynamics of N and P and caused increases in rates of photosynthesis. These findings contrast with results for a R. mangle-dominated forest in Belize where the fringe is N-limited, but the dwarf zone is P-limited and the transition zone is co-limited by N and P. This study demonstrated that patterns of nutrient limitation in mangrove ecosystems are complex, that not all processes respond similarly to the same nutrient, and that similar habitats are not limited by the same nutrient when different mangrove forests are compared.

Trace metal retention in mangrove ecosystems in Guanabara Bay, SE Brazil.

Along contrasting environmental conditions (e.g., degree of trace metal contamination and mangrove forest structural development), sediments of Laguncularia racemosa-dominated mangrove stands in Guanabara Bay (SE Brazil) presented a trend of trace metal accumulation in forms with low potential of remobilization and biotic uptake. Concurrently, a relatively low transfer of sediment-bound metals to L. racemosa leaves was observed, which may moderate the metal export from the forests via leaf litter transport and the metal availability to enter in food chains based on leaf consumption.