Samira H. Daroub

Pyrosequencing enumerates and contrasts soil microbial diversity.

Estimates of the number of species of bacteria per gram of soil vary between 2000 and 8.3 million (Gans et al., 2005; Schloss and Handelsman, 2006). The highest estimate suggests that the number may be so large as to be impractical to test by amplification and sequencing of the highly conserved 16S rRNA gene from soil DNA (Gans et al., 2005). Here we present the use of high throughput DNA pyrosequencing and statistical inference to assess bacterial diversity in four soils across a large transect of the western hemisphere. The number of bacterial 16S rRNA sequences obtained from each site varied from 26 140 to 53 533. The most abundant bacterial groups in all four soils were the Bacteroidetes, Betaproteobacteria and Alphaproteobacteria. Using three estimators of diversity, the maximum number of unique sequences (operational taxonomic units roughly corresponding to the species level) never exceeded 52 000 in these soils at the lowest level of dissimilarity. Furthermore, the bacterial diversity of the forest soil was phylum rich compared to the agricultural soils, which are species rich but phylum poor. The forest site also showed far less diversity of the Archaea with only 0.009% of all sequences from that site being from this group as opposed to 4%–12% of the sequences from the three agricultural sites. This work is the most comprehensive examination to date of bacterial diversity in soil and suggests that agricultural management of soil may significantly influence the diversity of bacteria and archaea.

Characterization of Lead in Soils of a Rifle/Pistol Shooting Range in Central Florida, USA

The distribution of lead in soil samples collected from both surface (0 to 10 cm) and profile (O 0 to 10 cm, E 11 to 30 cm, Eb 31 to 50 cm, Bw 51 to 100 cm, and C 181 to 200 cm) at a 14-year-old rifle/pistol shooting range located in central Florida were determined using EPA Method 3051a (microwave, HNO3/HCl=3:1, v/v). In addition to total lead analysis, Toxicity Characteristic Leaching Procedure (TCLP) analysis was performed on corresponding samples to determine whether the soils would require special handling as hazardous waste if the soils were to be removed from the range. Total lead in surface soils varied from 330 to 17 850 mg Pb kg−1, with the greatest concentration in the middle of the backstop berm. The TCLP tests indicated that lead in all surface soils exceeded the 5 mg Pb L−1 critical level of federal regulation for solid wastes and hazardous wastes provided by the Resource Conservation and Recovery Act (RCRA) and would be characterized as hazardous waste. Sequential fractionation and X-ray diffraction (XRD) analyses revealed that lead carbonate existed predominantly (91.3%) in the berm soil. The weathering of lead bullets in the soil environments formed primarily as hydrocerussite (Pb3(CO3)2(OH)2), with small amounts of massicot (PbO) and cerussite (PbCO3). However, the elevated soil pH, caused by the oxidization and transformation process of elemental lead in lead bullets, could be a significant factor in limiting the migration of lead in the soil.

Long-term water quality trends after implementing best management practices in South Florida.

A mandatory best management practices (BMP) program was implemented in the Everglades Agricultural Area (EAA) farms basin-wide in 1995 as required by the Everglades Forever Act to reduce P loads in drainage water reaching the Everglades ecosystem. All farms in the EAA basin implement similar BMPs, and basin wide P load reductions have exceeded the 25% reduction required by law; however, differences remain in water quality between subbasins. Our objective was to determine long-term trends in P loads in discharge water in the EAA after implementing BMPs for 7 to10 yr and to explore reasons for differences in the performance of the subbasins. Two monitoring datasets were used, one from 10 research farms and the second from the EAA basin inflow and outflow locations. Mann-Kendall trend analysis was used to determine the degree of change in water quality trends. A decreasing trend in P loads was observed in general on sugarcane (Saccharum officinarum L.) farms, while mixed crop farms showed either decreasing or insignificant trends. The insignificant trends are probably related to management practices of mixed crop systems. Decreasing trends in P loads were observed in the outflow of the EAA basin, S5A, and S8 subbasins from 1992 to 2002. Inflow water from Lake Okeechobee had increasing P concentration from 1992 to 2006 with the highest trend in the east side of the lake. This analysis indicated there may be other factors impacting the success of BMPs in individual farms including cropping rotations and flooding of organic soils. Elevated P concentrations in Lake Okeechobee water used for irrigation may pose a future risk to degrade water quality on farms in the EAA, especially in the S5A subbasin.

Effect of cropping and low-chemical input systems on soil phosphorus fractions

The adoption of alternative management practices has been shown to increase soil organic matter. However, the effect of adopting these practices on soil phosphorus (P), especially organic P, is not clear. We evaluated the effect of such practices-mainly no-tillage, zero and low-chemical input, organic-based, row crop agricultural systems-on soil P and compared them with conventional agriculture and perennial farming systems. We also compared soil P under conventional agriculture to an adjacent forest site and a never-tilled native successional community site in southwest Michigan. Sequential fractionation analysis of soil inorganic and organic P fractions showed that long-term conventional row crop agriculture resulted in a 79% reduction of NaOH-extractable organic P compared with adjacent forested sites. The calcium phosphate pool and the residual P fraction, however, increased under conventional agriculture compared with the forest site, probably because of fertilizer inputs. Adoption of no-tillage and low-chemical input systems with a winter leguminous cover crop in the rotation for 7 years did not increase organic P significantly in any of the fractions extracted from the annual cropping systems. However, organic P extracted by NaOH increased to 22.1% after adoption of continuous alfalfa for the same period compared with 11.4% extracted under conventionally tilled annual cropping systems. We conclude that continuous alfalfa can help restore soils to their native P fertility levels by taking up P from the stable residual fraction and transforming it into moderately labile organic P through root death. We found no evidence that low chemical input organic based systems are sustainable with regard to P as there is no evidence that organic P is accumulating as a result of the use of cover crops. Further investigation is warranted after these soils become P limiting and more years have passed under the same treatments.

Ca. Nitrososphaera and Bradyrhizobium are inversely correlated and related to agricultural practices in long-term field experiments

Agricultural land management, such as fertilization, liming, and tillage affects soil properties, including pH, organic matter content, nitrification rates, and the microbial community. Three different study sites were used to identify microorganisms that correlate with agricultural land use and to determine which factors regulate the relative abundance of the microbial signatures of the agricultural land-use. The three sites included in this study are the Broadbalk Experiment at Rothamsted Research, UK, the Everglades Agricultural Area, Florida, USA, and the Kellogg Biological Station, Michigan, USA. The effects of agricultural management on the abundance and diversity of bacteria and archaea were determined using high throughput, barcoded 16S rRNA sequencing. In addition, the relative abundance of these organisms was correlated with soil features. Two groups of microorganisms involved in nitrogen cycle were highly correlated with land use at all three sites. The ammonia oxidizing-archaea, dominated by Ca. Nitrososphaera, were positively correlated with agriculture while a ubiquitous group of soil bacteria closely related to the diazotrophic symbiont, Bradyrhizobium, was negatively correlated with agricultural management. Analysis of successional plots showed that the abundance of ammonia oxidizing-archaea declined and the abundance of bradyrhizobia increased with time away from agriculture. This observation suggests that the effect of agriculture on the relative abundance of these genera is reversible. Soil pH and NH3 concentrations were positively correlated with archaeal abundance but negatively correlated with the abundance of Bradyrhizobium. The high correlations of Ca. Nitrososphaera and Bradyrhizobium abundances with agricultural management at three long-term experiments with different edaphoclimatic conditions allowed us to suggest these two genera as signature microorganisms for agricultural land use.

Tree-based modeling of complex interactions of phosphorus loadings and environmental factors

Phosphorus (P) enrichment has been observed in the historic oligotrophic Greater Everglades in Florida mainly due to P influx from upstream, agriculturally dominated, low relief drainage basins of the Everglades Agricultural Area (EAA). Our specific objectives were to: (1) investigate relationships between various environmental factors and P loads in 10 farm basins within the EAA, (2) identify those environmental factors that impart major effects on P loads using three different tree-based modeling approaches, and (3) evaluate predictive models to assess P loads. We assembled thirteen environmental variable sets for all 10 sub-basins characterizing water level management, cropping practices, soils, hydrology, and farm-specific properties. Drainage flow and P concentrations were measured at each sub-basin outlet from 1992-2002 and aggregated to derive monthly P loads. We used three different tree-based models including single regression trees (ST), committee trees in Bagging (CTb) and ARCing (CTa) modes and ten-fold cross-validation to test prediction performances. The monthly P loads (MPL) during the monitoring period showed a maximum of 2528 kg (mean: 103 kg) and maximum monthly unit area P loads (UAL) of 4.88 kg P ha(-1) (mean: 0.16 kg P ha(-1)). Our results suggest that hydrologic/water management properties are the major controlling variables to predict MPL and UAL in the EAA. Tree-based modeling was successful in identifying relationships between P loads and environmental predictor variables on 10 farms in the EAA indicated by high R(2) (>0.80) and low prediction errors. Committee trees in ARCing mode generated the best performing models to predict P loads and P loads per unit area. Tree-based models had the ability to analyze complex, non-linear relationships between P loads and multiple variables describing hydrologic/water management, cropping practices, soil and farm-specific properties within the EAA.

Genome Sequence of Candidatus Nitrososphaera evergladensis from Group I.1b Enriched from Everglades Soil Reveals Novel Genomic Features of the Ammonia-Oxidizing Archaea

The activity of ammonia-oxidizing archaea (AOA) leads to the loss of nitrogen from soil, pollution of water sources and elevated emissions of greenhouse gas. To date, eight AOA genomes are available in the public databases, seven are from the group I.1a of the Thaumarchaeota and only one is from the group I.1b, isolated from hot springs. Many soils are dominated by AOA from the group I.1b, but the genomes of soil representatives of this group have not been sequenced and functionally characterized. The lack of knowledge of metabolic pathways of soil AOA presents a critical gap in understanding their role in biogeochemical cycles. Here, we describe the first complete genome of soil archaeon Candidatus Nitrososphaera evergladensis, which has been reconstructed from metagenomic sequencing of a highly enriched culture obtained from an agricultural soil. The AOA enrichment was sequenced with the high throughput next generation sequencing platforms from Pacific Biosciences and Ion Torrent. The de novo assembly of sequences resulted in one 2.95 Mb contig. Annotation of the reconstructed genome revealed many similarities of the basic metabolism with the rest of sequenced AOA. Ca. N. evergladensis belongs to the group I.1b and shares only 40% of whole-genome homology with the closest sequenced relative Ca. N. gargensis. Detailed analysis of the genome revealed coding sequences that were completely absent from the group I.1a. These unique sequences code for proteins involved in control of DNA integrity, transporters, two-component systems and versatile CRISPR defense system. Notably, genomes from the group I.1b have more gene duplications compared to the genomes from the group I.1a. We suggest that the presence of these unique genes and gene duplications may be associated with the environmental versatility of this group.

Best Management Practices and Long-Term Water Quality Trends in the Everglades Agricultural Area

The Everglades Agricultural Area (EAA) in South Florida, part of the historical Everglades, was initially drained in the early 20th century for agriculture and flood protection. The organic soils have been subject to subsidence caused by organic matter oxidation. Soils are deeper east of Lake Okeechobee compared to soils south of the lake. The area is mostly planted to sugarcane and other crops such as rice, vegetables, and sod. Concerns about quality of water leaving the EAA led to a regulatory program for mandatory best management practices (BMP) since 1995 to reduce phosphorus (P) loads out of the EAA by 25% compared to historical levels. The program is highly successful, with 100% grower participation and exceeding P load reduction required by law. Trend analysis conducted on selected EAA farms, subbasins, and whole basin show, in general, decreasing trends in P concentrations, drainage flow, and P loads. Differences are noted between farms and subbasins due to factors that include rainfall distribution, water management practices, irrigation water quality, soil type/depth, and cropping systems. Water management practices were the dominant factors affecting P loads out of the EAA. Water management research that targets farms with deeper soils is recommended to achieve additional P load reductions. Other practices to improve BMP performance include minimizing generation and transport of sediments from farm canals. The quality of irrigation water from Lake Okeechobee is of concern of its impact on BMP performance.

Comparison Of Three Soil Test Methods For Estimating Plant-available Silicon

Silicon (Si) confers increased disease resistance and nutritional benefits for both sugarcane and rice. Sugarcane is the primary crop grown in the Everglades Agricultural Area (EAA) in south Florida, USA, and production inputs routinely include Si fertilization. Soil testing for Si is based on a 0.5 N acetic acid extraction procedure that was developed for rice grown on the organic and mineral soils found in the EAA. The objective of this study was to compare a Florida based acetic acid extraction protocol with the sodium acetate buffer method used in Japan and Korea, and the 0.01 M calcium chloride method used in Australia. The three procedures were used to extract Si from soil samples collected from 31 countries, collectively representing 137 mineral soils. The collectors were asked to sample Oxisols, Ultisols, and coarse textured soils. A subset of the soil collection was classified as deficient (requiring Si fertilization for rice and/or sugarcane), based on published critical soil-test Si values specific to each extraction procedure. The sodium acetate buffer extracted the greatest amounts of Si (0 to 509 mg kg−1), followed by acetic acid (1 to 239 mg L−1) and calcium chloride (3 to 109 mg kg−1). Acetic acid and sodium acetate buffer soil-test Si values were fairly well correlated (r=0.77) and both methods performed well across a wide range of soils. Results with calcium chloride were less well correlated with acetic acid (r=0.73) and were poorly related to sodium acetate buffer (r=0.57). When considering only the subset of soils testing at or below the critical value, the correlation between acetic acid and sodium acetate buffer extractions was improved (r=0.84). #This research was supported by the Florida Agricultural Experiment Station and approved for publication as Journal series No. R-08790.

Using flow calorimetry to determine the molar heats of cation and anion exchange and the point of zero net charge on amorphous aluminum hydroxides

A technique for determining the point of zero net charge (PZNC) using flow adsorption calorimetry was investigated. The rationale behind this method is that the heat of exchange, determined calorimetrically, is directly proportional to surface charge and that the PZNC can be equated to the pH at which the heats of cation exchange (CE) and anion exchange (AE) are equal. The technique was tested using an amorphous aluminum hydroxide (AHO) synthesized in our lab. The flow calorimetry was conducted by measuring the heat associated with CE and AE at pH 5.8, 8.0, and 10.5. Heats of AE were measured using 50 mM NaCl and NaNO3; heats of CE were measured using 50 mM NaNO3 and KNO3. CE and AE capacities were measured on the AHO samples after removal from the calorimeter. The heat associated with NO3− replacing Cl was −4.5 kJ/mol; that for K replacing Na exchange was −1.2 kJ/mol. To determine the PZNC, the calorimetric heat (mJ/mg of sample) was plotted against pH. The heat for CE was multiplied by the factor, 4.5/1.2, to account for the difference in the two molar heats of exchange. The PZNC of the AHO determined calorimetrically was about 9.5, consistent with PZNC values reported in the literature for aluminum hydroxide. In other experiments, it was shown that surface charging of the AHO was reversible with pH and that exposing the AHO to arsenate while in the flow calorimeter lowered the PZNC by 1 pH unit. The use of flow calorimetry to measure surface charge properties of soil colloids has several advantages over existing methods: (i) it requires very small sample sizes; (ii) it is quicker and easier to carry out; and (iii) it gives better control of pH and ionic strength. With flow calorimetry, it is also easy to measure the effect of specifically adsorbed species on surface charge, as was demonstrated in the case of arsenate.