Soil phosphorus cycling across a 100-year deforestation chronosequence in the Amazon rainforest.

Deforestation of tropical rainforests is a major land use change that alters terrestrial biogeochemical cycling at local to global scales. Deforestation and subsequent reforestation are likely to impact soil phosphorus (P) cycling, which in P-limited ecosystems such as the Amazon basin has implications for long-term productivity. We used a 100-year replicated observational chronosequence of primary forest conversion to pasture, as well as a 13-year-old secondary forest, to test land use change and duration effects on soil P dynamics in the Amazon basin. By combining sequential extraction and P K-edge X-ray absorption near edge structure (XANES) spectroscopy with soil phosphatase activity assays, we assessed pools and process rates of P cycling in surface soils (0-10 cm depth). Deforestation caused increases in total P (135-398 mg kg-1 ), total organic P (Po ) (19-168 mg kg-1 ), and total inorganic P (Pi ) (30-113 mg kg-1 ) fractions in surface soils with pasture age, with concomitant increases in Pi fractions corroborated by sequential fractionation and XANES spectroscopy. Soil non-labile Po (10-148 mg kg-1 ) increased disproportionately compared to labile Po (from 4-5 to 7-13 mg kg-1 ). Soil phosphomonoesterase and phosphodiesterase binding affinity (Km ) decreased while the specificity constant (Ka ) increased by 83%-159% in 39-100y pastures. Soil P pools and process rates reverted to magnitudes similar to primary forests within 13 years of pasture abandonment. However, the relatively short but representative pre-abandonment pasture duration of our secondary forest may not have entailed significant deforestation effects on soil P cycling, highlighting the need to consider both pasture duration and reforestation age in evaluations of Amazon land use legacies. Although the space-for-time substitution design can entail variation in the initial soil P pools due to atmospheric P deposition, soil properties, and/or primary forest growth, the trend of P pools and process rates with pasture age still provides valuable insights.

Metagenomes from Eastern Brazilian Amazonian Floodplains in the Wet and Dry Seasons.

Here, we report the metagenomes from two Amazonian floodplain sediments in eastern Brazil. Tropical wetlands are well known for their role in the global carbon cycle. Microbial information on this diversified and dynamic landscape will provide further insights into its significance in regional and global biogeochemical cycles.

Increased soil moisture intensifies the impacts of forest-to-pasture conversion on methane emissions and methane-cycling communities in the Eastern Amazon.

Climatic changes are altering precipitation patterns in the Amazon and may influence soil methane (CH4) fluxes due to the differential responses of methanogenic and methanotrophic microorganisms. However, it remains unclear if these climate feedbacks can amplify land-use-related impacts on the CH4 cycle. To better predict the responses of soil CH4-cycling microorganisms and emissions under altered moisture levels in the Eastern Brazilian Amazon, we performed a 30-day microcosm experiment manipulating the moisture content (original moisture; 60%, 80%, and 100% of field capacity - FC) of forest and pasture soils. Gas samples were collected periodically for gas chromatography analysis, and methanogenic archaeal and methanotrophic bacterial communities were assessed using quantitative PCR and metagenomics. Positive and negative daily CH4 fluxes were observed for forest and pasture, indicating that these soils can act as both CH4 sources and sinks. Cumulative emissions and the abundance of methanogenesis-related genes and taxonomic groups were affected by land use, moisture, and their interaction. Pasture soils at 100% FC had the highest abundance of methanogens and CH4 emissions, 22 times higher than forest soils under the same treatment. Higher ratios of methanogens to methanotrophs were found in pasture than in forest soils, even at field capacity conditions. Land use and moisture were significant factors influencing the composition of methanogenic and methanotrophic communities. The diversity and evenness of methanogens did not change throughout the experiment. In contrast, methanotrophs exhibited the highest diversity and evenness in pasture soils at 100% FC. Taken together, our results suggest that increased moisture exacerbates soil CH4 emissions and microbial responses driven by land-use change in the Amazon. This is the first report on the microbial CH4 cycle in Amazonian upland soils that combined one-month gas measurements with advanced molecular methods.

Belowground changes to community structure alter methane-cycling dynamics in Amazonia.

Amazonian rainforest is undergoing increasing rates of deforestation, driven primarily by cattle pasture expansion. Forest-to-pasture conversion has been associated with increases in soil methane (CH4) emission. To better understand the drivers of this change, we measured soil CH4 flux, environmental conditions, and belowground microbial community structure across primary forests, cattle pastures, and secondary forests in two Amazonian regions. We show that pasture soils emit high levels of CH4 (mean: 3454.6 ± 9482.3 µg CH4 m-2 d-1), consistent with previous reports, while forest soils on average emit CH4 at modest rates (mean: 9.8 ± 120.5 µg CH4 m-2 d-1), but often act as CH4 sinks. We report that secondary forest soils tend to consume CH4 (mean: -10.2 ± 35.7 µg CH4 m-2 d-1), demonstrating that pasture CH4 emissions can be reversed. We apply a novel computational approach to identify microbial community attributes associated with flux independent of soil chemistry. While this revealed taxa known to produce or consume CH4 directly (i.e. methanogens and methanotrophs, respectively), the vast majority of identified taxa are not known to cycle CH4. Each land use type had a unique subset of taxa associated with CH4 flux, suggesting that land use change alters CH4 cycling through shifts in microbial community composition. Taken together, we show that microbial composition is crucial for understanding the observed CH4 dynamics and that microorganisms provide explanatory power that cannot be captured by environmental variables.

Diazotrophs Show Signs of Restoration in Amazon Rain Forest Soils with Ecosystem Rehabilitation.

Biological nitrogen fixation can be an important source of nitrogen in tropical forests that serve as a major CO2 sink. Extensive deforestation of the Amazon is known to influence microbial communities and the biogeochemical cycles they mediate. However, it is unknown how diazotrophs (nitrogen-fixing microorganisms) respond to deforestation and subsequent ecosystem conversion to agriculture, as well as whether they can recover in secondary forests that are established after agriculture is abandoned. To address these knowledge gaps, we combined a spatially explicit sampling approach with high-throughput sequencing of nifH genes. The main objectives were to assess the functional distance decay relationship of the diazotrophic bacterial community in a tropical forest ecosystem and to quantify the roles of various factors that drive the observed changes in the diazotrophic community structure. We observed an increase in local diazotrophic diversity (α-diversity) with a decrease in community turnover (ß-diversity), associated with a shift in diazotrophic community structure as a result of the forest-to-pasture conversion. Both diazotrophic community turnover and structure showed signs of recovery in secondary forests. Changes in the diazotrophic community were primarily driven by the change in land use rather than differences in geochemical characteristics or geographic distances. The diazotroph communities in secondary forests resembled those in primary forests, suggesting that at least partial recovery of diazotrophs is possible following agricultural abandonment.IMPORTANCE The Amazon region is a major tropical forest region that is being deforested at an alarming rate to create space for cattle ranching and agriculture. Diazotrophs (nitrogen-fixing microorganisms) play an important role in supplying soil N for plant growth in tropical forests. It is unknown how diazotrophs respond to deforestation and whether they can recover in secondary forests that establish after agriculture is abandoned. Using high-throughput sequencing of nifH genes, we characterized the response of diazotrophs' ß-diversity and identified major drivers of changes in diazotrophs from forest-to-pasture and pasture-to-secondary-forest conversions. Studying the impact of land use change on diazotrophs is important for a better understanding of the impact of deforestation on tropical forest ecosystem functioning, and our results on the potential recovery of diazotrophs in secondary forests imply the possible restoration of ecosystem functions in secondary forests.

Boosting landfill gas production from lignin-containing wastes via termite hindgut microorganism.

Lignocellulose comprises a significant portion of municipal solid waste (MSW) - 40-70% in developed countries, including paper, wood, and yard waste. Cellulose and hemicellulose are often shielded by lignin, posing a barrier to waste decomposition and landfill gas generation. Unfortunately, lignin is resistant to microbial degradation under low-oxygen conditions that normally occur in MSW landfills. The bacterium strain TAV5, microaerophilic and member of phylum Verrucomicrobia, isolated from the hindgut of the Reticulitermes flavipes termite, the most widely distributed subterranean termite in North America. Its genome contains genes associated with methylotrophic competency which code for enzymes that structurally modify lignin. The overall goal of this research was to use TAV5 to modify lignin and boost methane production from MSW. Batch-scale reactors (125 mL) were filled with paper, yard, or wood waste, and four ratios of mixed of waste. Reactors were seeded with different ratios of TAV5 to anaerobic digester (AD) microorganisms (representing landfill anaerobic microorganisms). Based on batch tests, optimal ratios of TAV5 to AD microorganisms were used to seed wastes (mixed, yard, and wood) in 6-gallon reactors. Addition of TAV5 increased methane production from mixed waste, yard waste, and wood, by 49%, 34%, and 297%, respectively. TAV5 decreased acid soluble lignin by 7-39%, depending on waste type. TAV5 grown under aerobic conditions and room temperature (not requiring a heated anaerobic chamber) was found to remain viable and increase methane production under low-level oxygen conditions (1-2%). This finding will potentially lessen costs for growing large volumes of it for seeding landfills.

Draft Genome Sequences of Strains TAV3 and TAV4 (Verrucomicrobia: Opitutaceae), Isolated from a Wood-Feeding Termite, and In Silico Analysis of Their Polysaccharide-Degrading Enzymes.

Here, we report the high-quality draft genome sequences of Opitutaceae sp. strains TAV3 and TAV4, which were isolated from the hindgut of the wood-feeding termite Reticulitermes flavipes Using a combination of Illumina and PacBio sequencing, we constructed nearly complete assemblies totaling 5.84 and 5.91 Mbp in length for strains TAV3 and TAV4, respectively. In addition, we report an in silico analysis of potential lignocellulose-digesting enzymes present in these strains.

Amazon forest-to-agriculture conversion alters rhizosphere microbiome composition while functions are kept.

The conversion of native forest to agriculture is the main cause of microbial biodiversity loss in Amazon soils. In order to better understand this effect, we used metagenomics to investigate microbial patterns and functions in bulk soil and rhizosphere of soybean, in a long-term forest-to-agriculture conversion. Long-term forest-to-agriculture led to microbial homogenization and loss of diversity in both bulk soil and rhizosphere, mainly driven by decreasing aluminum concentration and increased cations saturation in soil, due to liming and fertilization in long-term no-till cropping. Data revealed that long-term no-till cropping culminated in a decrease in Acidobacteria, Actinobacteria and Proteobacteria abundances. However, α- and ß-Proteobacteria abundances were higher in the rhizosphere than in bulk soil, regardless of the time after forest-to-agriculture conversion. Changes in functional potential occurred predominantly in bulk soil, with decreases in functions related to potassium metabolism and virulence, disease and defense, while functions related to nucleic acids metabolism increased. Functions in the soybean rhizosphere remained stable, except for those related to potassium metabolism, which decreased after 20-year no-till cropping. Together, our results show that the soybean root system selects microbial taxa via trade-offs, to maintain functional resilience in the rhizosphere microbiome over time.

Deforestation impacts network co-occurrence patterns of microbial communities in Amazon soils.

Co-occurrence networks allow for the identification of potential associations among species, which may be important for understanding community assembly and ecosystem functions. We employed this strategy to examine prokaryotic co-occurrence patterns in the Amazon soils and the response of these patterns to land use change to pasture, with the hypothesis that altered microbial composition due to deforestation will mirror the co-occurrence patterns across prokaryotic taxa. In this study, we calculated Spearman correlations between operational taxonomic units (OTUs) as determined by 16S rRNA gene sequencing, and only robust correlations were considered for network construction (-0.80 ≥ P ≥ 0.80, adjusted P < 0.01). The constructed network represents distinct forest and pasture components, with altered compositional and topological features. A comparative analysis between two representative modules of these contrasting ecosystems revealed novel information regarding changes to metabolic pathways related to nitrogen cycling. Our results showed that soil physicochemical properties such as temperature, C/N and H++Al3+ had a significant impact on prokaryotic communities, with alterations to network topologies. Taken together, changes in co-occurrence patterns and physicochemical properties may contribute to ecosystem processes including nitrification and denitrification, two important biogeochemical processes occurring in tropical forest systems.

Determining landscape-level drivers of variability for over fifty soil chemical elements.

Syntheses of large datasets have allowed increased clarity of distribution patterns and variation in soil major and trace elements. However, the drivers of variation in topsoil elements across biogeographical scales are not well understood. Our aim was to (1) identify how landscape-scale climate, geographical features, and edaphic factors influence soil elements, and (2) determine key environmental thresholds for shifts in soil element concentration. We analyzed patterns of variation in topsoil elements using 9830 samples collected across 39,000km2 in subtropical land in southeast China. Canonical correlations and multiple linear regressions were used to model variations of each element across mean annual temperature (MAT), mean annual precipitation (MAP), land use, spatial topography, and soil pH. Element concentrations show significant latitudinal and longitudinal trends, and are significantly influenced by climate, land use, spatial topography, and soil pH. Longitude, pH, MAT, and MAP were the environmental factors most tightly correlated with element concentrations. Climate and soil pH drove positive or negative alterations in soil elements, with threshold indicators of MAP=1000mm/1500mm, MAT=17.8°C/18.0°C, and pH=5.8/5.0, respectively. Our results indicate topsoil elements have structural and functional thresholds of climate and soil pH in relatively wet and acidic environments. Our findings can facilitate holistic soil element concentration predictions and help elucidate the specific influences of climate and soil pH, enabling development of more complete biogeochemical models.

Draft Genome Sequence of "Candidatus Spirobacillus cienkowskii," a Pathogen of Freshwater Daphnia Species, Reconstructed from Hemolymph Metagenomic Reads.

We report here the near-complete genome sequence of "Candidatus Spirobacillus cienkowskii," a spiral-shaped, red-pigmented uncultivated bacterial pathogen of Daphnia spp. The genome is 2.74 Mbp in size, has a GC content of 32.1%, and contains genes associated with bacterial motility and the production of carotenoids, which could explain the distinctive red color of hosts infected with this pathogen.

New Biological Insights Into How Deforestation in Amazonia Affects Soil Microbial Communities Using Metagenomics and Metagenome-Assembled Genomes.

Deforestation in the Brazilian Amazon occurs at an alarming rate, which has broad effects on global greenhouse gas emissions, carbon storage, and biogeochemical cycles. In this study, soil metagenomes and metagenome-assembled genomes (MAGs) were analyzed for alterations to microbial community composition, functional groups, and putative physiology as it related to land-use change and tropical soil. A total of 28 MAGs were assembled encompassing 10 phyla, including both dominant and rare biosphere lineages. Amazon Acidobacteria subdivision 3, Melainabacteria, Microgenomates, and Parcubacteria were found exclusively in pasture soil samples, while Candidatus Rokubacteria was predominant in the adjacent rainforest soil. These shifts in relative abundance between land-use types were supported by the different putative physiologies and life strategies employed by the taxa. This research provides unique biological insights into candidate phyla in tropical soil and how deforestation may impact the carbon cycle and affect climate change.

Burkholderia pseudomallei-absent soil bacterial community results in secondary metabolites that kill this pathogen.

Burkholderia pseudomallei is a Gram-negative bacterium found in soil and the causative agent of a severe disease in humans and animals known as melioidosis. It is intrinsically resistant to many antibiotics and has been reported resistant to the drugs of choice; ceftazidime. Microbial communities in soil in the presence and absence of B. pseudomallei were investigated using metagenomics approach. The variation in bacterial species diversity was significantly higher in soil samples without B. pseudomallei. Abundances of phyla Actinobacteria and Firmicutes were found significantly higher in B. pseudomallei-negative soils. Bacillus amyloliquefaciens KKU1 in phylum Firmicutes was discovered from negative soil and its secondary metabolites could inhibit clinical, environmental and drug resistant isolates of B. pseudomallei, together with some pathogenic Gram-negative but not Gram-positive bacteria. The antimicrobial activity from KKU 1 against B. pseudomallei was abolished when treated with proteinase K, stable in a wide range of pH and remained active after heating at 100 °C for 15 min. Precipitated proteins from KKU1 were demonstrated to cause lysis and corrugated surfaces of B. pseudomallei. The minimum inhibitory concentrations and minimum bactericidal concentrations of the precipitated proteins from KKU1 against B. pseudomallei were 0.97 µg/ml and 3.9 µg/ml. Interestingly, Native SDS-PAGE showed small active compounds of less than 6 kDa, along with other information collectively suggesting the properties of antimicrobial peptides. For the first time, culture-independent information in melioidosis endemic area could lead to a suspected source of metabolites that may help defense against B. pseudomallei and other pathogenic Gram-negative bacteria.

Long-term effects of grazing and topography on extra-radical hyphae of arbuscular mycorrhizal fungi in semi-arid grasslands.

Grazing and topography have drastic effects on plant communities and soil properties. These effects are thought to influence arbuscular mycorrhizal (AM) fungi. However, the simultaneous impacts of grazing pressure (sheep ha-1) and topography on plant and soil factors and their relationship to the production of extra-radical AM hyphae are not well understood. Our 10-year study assessed relationships between grazing, plant species richness, aboveground plant productivity, soil nutrients, edaphic properties, and AM hyphal length density (HLD) in different topographic areas (flat or sloped). We found HLD linearly declined with increasing grazing pressure (1.5-9.0 sheep ha-1) in sloped areas, but HLD was greatest at moderate grazing pressure (4.5 sheep ha-1) in flat areas. Structural equation modeling indicates grazing reduces HLD by altering soil nutrient dynamics in sloped areas, but non-linearly influences HLD through plant community and edaphic changes in flat areas. Our findings highlight how topography influences key plant and soil factors, thus regulating the effects of grazing pressure on extra-radical hyphal production of AM fungi in grasslands. Understanding how grazing and topography influence AM fungi in semi-arid grasslands is vital, as globally, severe human population pressure and increasing demand for food aggravate the grazing intensity in grasslands.

Soil microbial community dynamics and assembly under long-term land use change.

We evaluated the bacterial and archaeal community dynamics and assembly in soils under forest, grassland and no-till cropping, using a high-throughput shotgun metagenomics approach. No significant alterations in alpha diversity were observed among different land uses, but beta diversity in grassland was lower than that observed in forest and no-till soils. Grassland communities showed assembly that predominantly followed the neutral model, i.e. high homogenizing selection with moderate dispersion, leading to biotic homogenization. Both no-till and forest soil communities were found to have assembly that predominantly followed a niche model, i.e. low rates of dispersal and weak homogenizing selection, resulting in maintenance of higher beta diversity relative to grasslands, indicating niche specialization or variable selection. Taken together, our results indicate that the patterns of assembly and their governing processes are dependent on the land use employed after deforestation, with consequences for taxa turnover and microbial functional potential.

Distinct Biogeographic Patterns for Archaea, Bacteria, and Fungi along the Vegetation Gradient at the Continental Scale in Eastern China.

The natural forest ecosystem in Eastern China, from tropical forest to boreal forest, has declined due to cropland development during the last 300 years, yet little is known about the historical biogeographic patterns and driving processes for the major domains of microorganisms along this continental-scale natural vegetation gradient. We predicted the biogeographic patterns of soil archaeal, bacterial, and fungal communities across 110 natural forest sites along a transect across four vegetation zones in Eastern China. The distance decay relationships demonstrated the distinct biogeographic patterns of archaeal, bacterial, and fungal communities. While historical processes mainly influenced bacterial community variations, spatially autocorrelated environmental variables mainly influenced the fungal community. Archaea did not display a distance decay pattern along the vegetation gradient. Bacterial community diversity and structure were correlated with the ratio of acid oxalate-soluble Fe to free Fe oxides (Feo/Fed ratio). Fungal community diversity and structure were influenced by dissolved organic carbon (DOC) and free aluminum (Ald), respectively. The role of these environmental variables was confirmed by the correlations between dominant operational taxonomic units (OTUs) and edaphic variables. However, most of the dominant OTUs were not correlated with the major driving variables for the entire communities. These results demonstrate that soil archaea, bacteria, and fungi have different biogeographic patterns and driving processes along this continental-scale natural vegetation gradient, implying different community assembly mechanisms and ecological functions for archaea, bacteria, and fungi in soil ecosystems. IMPORTANCE Understanding biogeographic patterns is a precursor to improving our knowledge of the function of microbiomes and to predicting ecosystem responses to environmental change. Using natural forest soil samples from 110 locations, this study is one of the largest attempts to comprehensively understand the different patterns of soil archaeal, bacterial, and fungal biogeography at the continental scale in eastern China. These patterns in natural forest sites could ascertain reliable soil microbial biogeographic patterns by eliminating anthropogenic influences. This information provides guidelines for monitoring the belowground ecosystem's decline and restoration. Meanwhile, the deviations in the soil microbial communities from corresponding natural forest states indicate the extent of degradation of the soil ecosystem. Moreover, given the association between vegetation type and the microbial community, this information could be used to predict the long-term response of the underground ecosystem to the vegetation distribution caused by global climate change.

Conversion of Amazon rainforest to agriculture alters community traits of methane-cycling organisms.

Land use change is one of the greatest environmental impacts worldwide, especially to tropical forests. The Amazon rainforest has been subject to particularly high rates of land use change, primarily to cattle pasture. A commonly observed response to cattle pasture establishment in the Amazon is the conversion of soil from a methane sink in rainforest, to a methane source in pasture. However, it is not known how the microorganisms that mediate methane flux are altered by land use change. Here, we use the deepest metagenomic sequencing of Amazonian soil to date to investigate differences in methane-cycling microorganisms and their traits across rainforest and cattle pasture soils. We found that methane-cycling microorganisms responded to land use change, with the strongest responses exhibited by methane-consuming, rather than methane-producing, microorganisms. These responses included a reduction in the relative abundance of methanotrophs and a significant decrease in the abundance of genes encoding particulate methane monooxygenase. We also observed compositional changes to methanotroph and methanogen communities as well as changes to methanotroph life history strategies. Our observations suggest that methane-cycling microorganisms are vulnerable to land use change, and this vulnerability may underlie the response of methane flux to land use change in Amazon soils.

Hill number as a bacterial diversity measure framework with high-throughput sequence data.

Bacterial diversity is an important parameter for measuring bacterial contributions to the global ecosystem. However, even the task of describing bacterial diversity is challenging due to biological and technological difficulties. One of the challenges in bacterial diversity estimation is the appropriate measure of rare taxa, but the uncertainty of the size of rare biosphere is yet to be experimentally determined. One approach is using the generalized diversity, Hill number (Na), to control the variability associated with rare taxa by differentially weighing them. Here, we investigated Hill number as a framework for microbial diversity measure using a taxa-accmulation curve (TAC) with soil bacterial community data from two distinct studies by 454 pyrosequencing. The reliable biodiversity estimation was obtained when an increase in Hill number arose as the coverage became stable in TACs for a ≥ 1. In silico analysis also indicated that a certain level of sampling depth was desirable for reliable biodiversity estimation. Thus, in order to attain bacterial diversity from second generation sequencing, Hill number can be a good diversity framework with given sequencing depth, that is, until technology is further advanced and able to overcome the under- and random-sampling issues of the current sequencing approaches.