Validation of Soil Enzyme Activity Assay for a Biogeochemical Cycling Index in Biochar Amended Soils

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 (aryl-sulfatase) 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.

Enzymatic Hydrolysis of an Organic Sulfur Compound

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.

Microbial Compositions and Enzymes of a Forest Ecosystem in Alabama: Initial Response to Thinning and Burning Management Selections

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.