Understanding how different tree species affect soil microbial communities is crucial for sustainable forestry and ecosystem management practices. Despite Black walnut (Juglans nigra L.) forestry having a rich history, the overall comprehension of how this hardwood species influences soil remains incomplete. In earlier studies, we examined the effects of hardwood plantations on soil chemical properties and their interaction with microbial biomass, however, we highlight the importance of studying the soil microbial communities and their relationship with soil properties in greater depth. Building on this foundation, our research focused on evaluating microbiome compositions beneath J. nigra and another hardwood, Northern red oak (Quercus rubra L.) after a decade of establishment. We uncovered intriguing patterns within the soil bacterial/archaeal and fungal structures by conducting meticulous analyses utilizing amplicon sequencing alongside soil chemical properties. Our findings underscore that tree species play a pivotal role in shaping soil microbial structures, a role that surpasses even seasonal and depth influences. Most notably, J. nigra stands out for its ability to enhance microbial diversity, as evidenced by increased alpha-diversity indices compared to baseline values. Conversely, Q. rubra tends to decrease these indices. Significant disparities in microbial composition between the two tree species were evident, with J. nigra exhibiting enrichment in certain taxa such as Nitrospira, Geobacter, and Bacillus while Q. rubra showed enrichment in others like Acidobacteriota and ectomycorrhizal fungi. Furthermore, we also observed differences in co-occurrence networks by delving deeper into the interconnections within the soil microbiota. In both fungal and bacterial/archaeal communities, J. nigra and Q. rubra notably decreased the number of connections within their networks, while Q. rubra increased some, suggesting a more interconnected network. These differences were further highlighted by network metrics with Q. rubra displaying a higher mean degree and clustering coefficient. Additionally, our analysis revealed that tree species influence soil chemical properties, either directly or indirectly, thereby affecting soil bacterial and fungal communities. In conclusion, our study elucidates the intricate interplay between tree species and soil microbiota, emphasizing the need to consider these relationships in forestry and ecosystem management practices.

Interactions between above- and below-ground monoculture forest plantation components are critical to tree growth and development. Within the Central Hardwoods Region (CHR), synergistic relationships between tree species and soil microbial community structure and function have received limited research attention. Soil microbes are integral to forest ecosystems as their activities intrinsically promote soil organic matter decomposition, nutrient cycling, and ecosystem functioning. Here, we examined soils from two perfectly aligned stands of black walnut (BW, Juglans nigra L.) and Northern red oak (RO, Quercus rubra L.) trees. Measurements of selected soil chemical properties, microbial community structure using ester-linked fatty acid methyl ester (EL-FAME), and soil enzyme activities (EAs) were used. Analysis of modifications within microbial communities showed a significant positive response to BW based upon soil EAs and microbial indicators, compared to RO. Seasonal comparisons predictably revealed higher microbial activities during summer. Fungi dominated the soil microbial community structure with a fungal/bacterial ratio of 2:1. Gram-positive rather than Gram-negative bacteria or actinomycetes dominated the bacterial community. The activity of the soil enzymes ß-glucosidase and arylsulfatase increased, but ß-glucosaminidase and acid phosphatase decreased. Additionally, acid phosphatase and arbuscular mycorrhizal fungi revealed strong correlations. The differences observed in biological properties, specifically microbial communities and EAs, highlight the varied responses to BW and RO soil biology and subsequent soil ecosystem functions. These results indicate that variations in microbial abundance and soil functions occur throughout the course of an entire year.

Biochar offers several benefits as a soil amendment, including increased soil fertility, carbon sequestration, and water-holding capacity in nutrient-poor soils.Here, we performed a series of enzyme assays on pine biochar-amended soils, comparing multiple enzyme activities (EAs) simultaneously determined in the same soil sample vs. the sum of individual EAs involved in the C, N, S, and P cycles to provide information of the impacts of biochar on biogeochemical cycling.The combination of these four EAs has been considered an indicator of soil health due to their role in the reactions that release bioavailable nutrients in the cycling of C (β-glucosidase), N and C (β-glucosaminidase), P (acid phosphomonoesterase), and S (arylsulfatase) in soils.Comparisons of the theoretical EAs and the CNPS activity assay approaches in the biochar-modified soil revealed similar activity trends with the different concentrations of added biochar.Two years after adding biochar, study results showed the amended soils did not retain more pNP substrate than the un-amended control soils in three different pH buffers (5.5, 5.8, and 6.5) commonly used in EA reactions.Finally, we performed a third experiment to determine if the biochar previously added to the EAs interfered with the reactions' enzyme or substrate.The results indicated that greater activity was measured using the combined assay, which suggests the CNPS activity method was less affected by biochar than the individual EAs.Our findings indicate that the potential soil biochemical-health index, CNPS activity (combination of four enzymes) assay is more robust than the individual EAs and can be used as an alternative tool to monitor soil functioning.

Forest floor and mineral soil manipulations influence the soil biogeochemical properties important for loblolly pine (Pinus taeda L.) tree growth. The impacts of forest floor manipulations on soil aggregate stability and the presence of ectomycorrhizal fungi (EMF), was assessed to elucidate the relationship between EMF abundance and aggregate stability. The study site consists of a 14-year-old loblolly pine plantation managed by Weyerhaeuser Company in the Lower Coastal Plain, approximately 8 miles east of New Bern, North Carolina, USA. The soil samples were collected from the top 7.62 cm of each soil treatment which includes three levels of forest floor retention: removed, control, and doubled and two levels of forest floor mixing with the mineral soil: mixed and unmixed. Ectomycorrhizal fungi abundance was evaluated by ester-linked fatty acid methyl ester analysis and microbial community functionality was assessed by acid-phosphatase activity measurement. Aggregate stability was assessed using the aggregate mean weight diameter approach. Results indicate that the forest floor manipulations had no significant impact on aggregate stability and EMF abundance. However, a positive relationship between EMF abundance and aggregate stability was identified. Removing the forest floor resulted in a soil bulk density increase of 0.18 g cm-3 compared to doubling the forest floor. Our results demonstrate that some mineral soil properties recover relatively quickly from forest floor manipulations. The study informs forest managers interested in how soil responds to forest floor manipulation and the interaction between EMF and aggregate stability.

Saprolite, weathered bedrock, is being used to dispose of domestic sewage through septic system drainfields, but the thickness of saprolite needed to remove biological contaminants is unknown for most saprolites. This study developed and tested a simple method for estimating the thickness of saprolite needed below septic drainlines to filter E. coli from wastewater using estimates of the volume of pores that are smaller than the length of the coliform (≤10 μm). Particle size distribution (texture) and water retention data were obtained for 12 different saprolites from the Piedmont and Mountain regions of North Carolina (N.C.). Saprolite textures ranged from clay loam to coarse sand. The volume of pores with diameters ≤10 μm were determined by water retention measurements for each saprolite. The data were used in an equation to estimate the saprolite thickness needed to filter E. coli. The estimated saprolite thicknesses ranged from 36 cm in the clay loam to 113 cm for the coarse sand. The average thickness across all samples was 58 cm. Saprolite thickness estimates increased as silt percentage decreased and as sand percentage and in situ saturated hydraulic conductivity increased. Silt percentage may be most useful for estimating appropriate saprolite thicknesses in the field.

Biochar offers several benefits as a soil amendment, including increased soil fertility, carbon sequestration, and water-holding capacity in nutrient-poor soils. In this study, soil samples with and without biochar additives were collected for two consecutive years from an experimental field plot to examine its effect on the microbial community structure and functions in sandy soils under peach-trees (Prunus persica). The four treatments evaluated consisted of two different rates of biochar incorporated into the soil (5%, and 10%, v/v), one “dynamic” surface application of biochar, and a 0% biochar control. Fatty acid methyl ester (FAME) analysis was used to assess the microbial community structure, and enzyme activities involved in C, N, P, and S nutrient cycling were used as a means of assessing soil functionality. Total FAME and bacterial indicators increased by 18% and 12%, respectively, in the 10% incorporated and 5% surface applied treatments. Biochar applications increased β-glucosaminidase and arylsulfatase activities, 5–30% and 12–46%, respectively. β-glucosidase and acid phosphatase activities decreased by approximately 18–35% and 5–22% in the 0–15 cm soils. The overall results suggest that biochar’s addition to the sandy soils stimulated microbial activity, contributing to the increased mean weight diameter (MWD), C sequestration, and consequential soil health. The changes in microbial community structure and functions may be useful predictors of modifications in soil organic matter (SOM) dynamics due to the long-term application of pine biochar in these systems.

The aim of this study was to evaluate the treatment effects of Ca(OH)(2) solutions applied at different concentrations (2.5%, 5%, and 7.5%) on the structural and chemical composition of sugarcane bagasse (SCB). The feasibility of using the SCB as a substrate for endoglucanase and beta-glucosidase production by Aspergillus niger through solid-state fermentation was also assessed. Sodium hydroxide (NaOH) 5% was used as a positive control/benchmark. Changes in SCB cell wall compositions was determined using C-13 solid-state nuclear magnetic resonance. While cellulosic fiber content and the production of endoglucanase (345.01 +/- 139 U.gDM(-1)) increased using Ca(OH)(2) as pretreatment for sugarcane bagasse, hemicellulose, lignin, alkyl C and carboxyl C content decreased. Comparisons between pretreated and non-pretreated SCB revealed that the highest values of beta-glucosidase (58.18 +/- 9.86 U.gDM(-1)) occurred in non-pretreated samples. The SCB pretreated with NaOH resulted in the highest delignification values (-49.88%) and the greatest hemicellulose remotion (-53.47%). However, this treatment also increased crystallinity index (CI; + 23.61%) in cellulose and showed the lowest endoglucanase (22.79 +/- 11 U.gDM(-1)) and beta-glucosidase (4.71 +/- 1.44 U.gDM(-1)) activities. This study claims that cellulose CI, hemicellulose, and other nutrients (e.g., amino acids) are important variables that should be considered along with delignification, in the substrates selection for cellulase production using filamentous fungi.

A good bioindicator of soil health is one that perceives minor changes due to changes in soil management and leads to changes in the processes of the soil ecosystem. The aim was to evaluate the use of soil bioindicators in the determination of the utility of two different Cedrela odorata forestry management systems. The litter and soil samples were collected from the rhizosphere of a monoculture (with no fertilization nor irrigation), and a co-culture (with fertilization and irrigation) stand with Citrus latifolia. The soil and litter bioindicators measured were the activity of acetylesterase (FDA), acid phosphatase (AcPh), alkaline phosphatase (AlkPh), laccase, and the estimation of the potential net rate of carbon mineralization (PNRCM). The results indicated that soil enzyme activities (FDA and phosphatases) in the litter were sensitive to the different management system treatments. Enzymatic activity, per unit dry weight, was higher in litter of co-culture (FDA = 1.05 nkat g-1 AcPh = 1.33 nkat g-1; AlkPh = 03.0 nkat g-1) than in litter of monoculture (FDA = 0.617nkat g-1; AcPh = 0.40 nkat g-1; AlkPh = 0. 983 nkat g-1). Differences in soil total nitrogen content, organic matter (OM) content, and micronutrients were found. In general, the co-culture stand (with fertilization and irrigation) with C. latifolia showed the best results.

A good bioindicator of soil health is one that perceives minor changes due to changes in soil management and leads to changes in the processes of the soil ecosystem. The aim was to evaluate the use of soil bioindicators in the determination of the utility of two different Cedrela odorata forestry management systems. The litter and soil samples were collected from the rhizosphere of a monoculture (with no fertilization nor irrigation), and a co-culture (with fertilization and irrigation) stand with Citrus latifolia. The soil and litter bioindicators measured were the activity of acetylesterase (FDA), acid phosphatase (AcPh), alkaline phosphatase (AlkPh), laccase, and the estimation of the potential net rate of carbon mineralization (PNRCM). The results indicated that soil enzyme activities (FDA and phosphatases) in the litter were sensitive to the different management system treatments. Enzymatic activity, per unit dry weight, was higher in litter of co-culture (FDA = 1.05 nkat g-1 AcPh = 1.33 nkat g-1; AlkPh = 03.0 nkat g-1) than in litter of monoculture (FDA = 0.617nkat g-1; AcPh = 0.40 nkat g-1; AlkPh = 0. 983 nkat g-1). Differences in soil total nitrogen content, organic matter (OM) content, and micronutrients were found. In general, the co-culture stand (with fertilization and irrigation) with C. latifolia showed the best results.

Saprolite, weathered bedrock, is being used to dispose of domestic sewage through septic system drainfields, but its ability to remove coliforms is unknown. This study determined if Escherichia coli could be removed by a sandy loam saprolite material. Triplicate columns containing saprolite were prepared with lengths of 30, 45, and 60 cm. A 215-mL solution containing 1 × 105 CFU/100 mL of non-toxic E. coli was applied to the top of each column for 5 days/week for 13 weeks, and selected outflow samples were analyzed for E. coli. Control columns had only tap water applied to them at the same time. Significantly higher (p ≤ 0.10 compared to controls) E. coli concentrations were only detected in samples collected at the end of week 3 for the 30-cm columns and week 4 for the 45-cm columns. E. coli concentrations were small and ranged from approximately 2 to 3 MPN/100 mL. No E. coli were detected in any outflow from the 60-cm columns. From weeks 5 to 13, E. coli concentrations from all columns were either undetectable or not significantly different from the control. The results showed that 60 cm of sandy loam saprolite was sufficient for the removal of E. coli from simulated wastewater.

Manganese-oxidizing fungi support bioremediation through the conversion of manganese ions into manganese oxide deposits that in turn adsorb manganese and other heavy metal ions from the environment. Manganese-oxidizing fungi were immobilized onto nanofiber surfaces to assist remediation of heavy metal–contaminated water. Two fungal isolates, Coniothyrium sp. and Coprinellus sp., from a Superfund site (Lot 86, Farm Unit #1) water treatment system were incubated in the presence of nanofibers. Fungal hyphae had strong association with nanofiber surfaces. Upon fungal attachment to manganese chloride–seeded nanofibers, Coniothyrium sp. catalyzed the conformal deposition of manganese oxide along hyphae and nanofibers, but Coprinellus sp. catalyzed manganese oxide only along its hyphae. Fungi–nanofiber hybrids removed various heavy metals from the water. Heavy metal ions were adsorbed into manganese oxide crystalline structure, possibly by ion exchange with manganese within the manganese oxide. Hybrid materials of fungal hyphae and manganese oxides confined to nanofiber-adsorbed heavy metal ions from water.

AbstractBiochar may enhance soil health in sandy soils under peach [Prunus persica (L.) Batsch] tree production. A study was initiated in the Sandhills of North Carolina by applying pine‐biochar at two different rates (5 and 10%, v/v) and types of application (incorporation to 30 cm [‐Inc] and surface [‐Sur]) in 2017. Soil organic matter (SOM) dynamics was assessed via soil organic carbon (SOC), total nitrogen (TN), permanganate‐oxidizable carbon (POXC), aggregate fractions (macro‐, micro‐aggregates, and clay + silt particles), and enzyme activities (CNPS activity) at 0–15 and 15–30 cm in 2018 and 2019. After the first year of pine‐biochar application, POXC decreased in all biochar treatments, SOC was higher in the 10%‐Inc treatment, whereas CNPS activities and macroaggregates were highest in the 5%‐Sur compared with the non‐treated control. These results suggest early changes in SOM dynamics and biogeochemical cycling and have the potential to guide producers with the long‐term application of pine‐biochar to sandy soils.

Sulfatases which cleave sulfate esters in biological systems are key enzymes that deserve special attention due to their significant roles in organic sulfur (OS) mineralization and inorganic sulfur () release. In this study, in-vitro experiments were conducted to evaluate S bonded substrate hydrolysis by a commercially available arylsulfatase (EC 3.1.6.1) from Aerobacter aerogenes. The enzyme-substrate interactions were assessed to determine: 1) rate of hydrolysis, 2) catalytic efficiency, 3) thermal stability, and 4) optimal pH of this enzyme. Arylsulfatase exhibited substrate hydrolysis with a high affinity for p-nitrophenyl sulfate (potassium 4-nitrophenyl sulfate (pNPS)). The optimum activity for the enzyme was observed to occur at a pH of 7.1. The optimal temperature was 37°C but ranged from 35°C - 45°C. The apparent Km and Kcat of the enzyme for pNPS hydrolysis at the optimal pH, and temperature were determined to be 1.03 mM and 75.73 μM/min, respectively. This work defines the catalytic and kinetic properties of arylsulfatase (EC 3.1.6.1) and confirms the optimal conditions for sulfatase activity testing. The resulting information is useful in elucidating the contributions that individual enzymes have for specific reactions rather than relying on traditional total enzyme activity measurements.

The internally transcribed spacer (ITS) region between the small subunit ribosomal RNA gene and large subunit ribosomal RNA gene is a widely used phylogenetic marker for fungi and other taxa. The eukaryotic ITS contains the conserved 5.8S rRNA and is divided into the ITS1 and ITS2 hypervariable regions. These regions are variable in length and are amplified using primers complementary to the conserved regions of their flanking genes. Previous work has shown that removing the conserved regions results in more accurate taxonomic classification. An existing software program, ITSx, is capable of trimming FASTA sequences by matching hidden Markov model profiles to the ends of the conserved genes using the software suite HMMER. ITSxpress was developed to extend this technique from marker gene studies using Operational Taxonomic Units (OTU’s) to studies using exact sequence variants; a method used by the software packages Dada2, Deblur, QIIME 2, and Unoise. The sequence variant approach uses the quality scores of each read to identify sequences that are statistically likely to represent real sequences. ITSxpress enables this by processing FASTQ rather than FASTA files. The software also speeds up the trimming of reads by a factor of 14-23 times on a 4-core computer by temporarily clustering highly similar sequences that are common in amplicon data and utilizing optimized parameters for Hmmsearch. ITSxpress is available as a QIIME 2 plugin and a stand-alone application installable from the Python package index, Bioconda, and Github.

Prescribed burning and tree thinning are commonly used restoration practices for US forests management to increase forest productivity and enhance plant and animal diversity. The impact of these practices in Alabama’s Bankhead National Forest (BNF) to soil microbial components and overall forest soil health are unknown. We hypothesized that microbial assemblages and enzyme activities are continuously changing in forest ecosystems especially due to management selections. Therefore, the objective of this study was to assess changes in microbial community compositions (fungal vs bacterial populations) via fatty acid methyl ester (FAME) profiling and several enzyme activities (β-glucosaminidase, acid phosphatase, arylsulfatase, β-glucosidase, xylanase, laccase, and manganese peroxidase) critical to soil organic matter (SOM) dynamics and biogeochemical cycling. In this forest, heavily-thinned plots without burning or less frequent burning treatments seemed to provide more favorable conditions (higher pH and lower C:N ratios) for C and N mineralization. This may explain a slight increase (by 12%) detected in fungi:bacteria (F:B) ratio in the heavily-thinned plots relative to the control. Thinned (lightly and heavily) plots showed greater ligninolytic (laccase and MnP) activities and lower β-glucosidase and β-glucosaminidase activities compared to the no-thinned plots probably due to increase depositions of woody recalcitrant C materials. We observed significant but negative correlations between the ligninolytic laccase and manganese peroxidase (Lac and MnP) enzymes respectively, with MBC (?0.45* and ?0.68** respectively) and MBN (?0.43* and ?0.65** respectively). Prescribed burning treatment reduced microbial biomass C and N of the 9-yr burned plot/lightly thinned plotsprobably due to depletion of labile C sources with the high temperatures, leaving mostly recalcitrant C sources as available soil substrates. Gram-positive bacteria (i15:0, a15:0, i17:0, and a17:0), actinomycetes (10-Me17:0, 10-Me18:0), AMF (16:1ω5c), and saprophytic fungi (18:1ω9c), largely contributed to the microbial compositions. This study bridges knowledge gaps in our understanding of microbial community compositions and enzyme-mediated processes in repeatedly burned and thinned forest ecosystems.

Manganese oxides, which may be biogenically produced in both pristine and contaminated environments, have a large affinity for many trace metals. In this study, water and Mn oxide-bearing biofilm samples were collected from the components of a pump and treat remediation system at a superfund site. To better understand the factors leading to their formation and their effects on potentially toxic metal fate, we conducted a chemical, microscopic, and spectroscopic characterization of these biofilm samples. Scanning electron microscopy revealed the presence of Mn oxides in close association with biological structures with morphologies consistent with fungi. X-ray absorption spectroscopy (XAS) and X-ray diffraction (XRD) revealed the oxides to be a mixture of layer and tunnel structure Mn(iv) oxides. In addition, XAS suggested that Ba, Co, and Zn all primarily bind to oxides in the biofilm in a manner that is analogous to synthetic or laboratory grown bacteriogenic Mn oxides. The results indicate that Mn oxides produced by organisms in the system may effectively scavenge metals, thus highlighting the potential utility of these organisms in designed remediation systems.

Elemental cycling is critical to the function of ecosystems and delivery of key ecosystem services because many of these elements are essential nutrients or detrimental toxicants that directly affect the health of organisms and ecosystems. Soil biogeochemical processes control the transformation, fate, and transport of elemental species in soils. In this chapter, we describe the biogeochemistry of carbon, nitrogen, and phosphorus, with an emphasis on the physical, chemical, and biological processes that drive their cycling at the microscopic to global scale. Additionally, how soils control the fate and transport of metals and synthetic organic chemicals in the environment is characterized. In an additional effort to provide transparent connections to the real world, case studies are presented to illustrate how proper management of soils can foster key biogeochemical processes to maximize ecosystem services and address current environmental issues. This chapter provides the background needed for an informed public to gain understanding of the multitude of soil biogeochemical functions and how soil provides and regulates services to society.