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The physical and chemical changes that accompany permafrost thaw directly influence the microbial communities that mediate the decomposition of formerly frozen organic matter, leading to uncertainty in permafrost–climate feedbacks. Although changes to microbial metabolism and community structure are documented following thaw, the generality of post‐thaw assembly patterns across permafrost soils of the world remains uncertain, limiting our ability to predict biogeochemistry and microbial community responses to climate change. Based on our review of the Arctic microbiome, permafrost microbiology, and community ecology, we propose that Assembly Theory provides a framework to better understand thaw‐mediated microbiome changes and the implications for community function and climate feedbacks. This framework posits that the prevalence of deterministic or stochastic processes indicates whether the community is well‐suited to thrive in changing environmental conditions. We predict that on a short timescale and following high‐disturbance thaw (e.g., thermokarst), stochasticity dominates post‐thaw microbiome assembly, suggesting that functional predictions will be aided by detailed information about the microbiome. At a longer timescale and lower‐intensity disturbance (e.g., active layer deepening), deterministic processes likely dominate, making environmental parameters sufficient for predicting function. We propose that the contribution of stochastic and deterministic processes to post‐thaw microbiome assembly depends on the characteristics of the thaw disturbance, as well as characteristics of the microbial community, such as the ecological and phylogenetic breadth of functional guilds, their functional redundancy, and biotic interactions. These propagate across space and time, potentially providing a means for predicting the microbial forcing of greenhouse gas feedbacks to global climate change. [ABSTRACT FROM AUTHOR]

While soil organic carbon (C) is the foundation of productive and healthy ecosystems, the impact of the ecology of microorganisms on C-cycling remains unknown. We manipulated the diversity, applied here as species richness, of the microbial community present in similar soils on two contrasting land-covers—an adjacent pasture and forest—and observed the transformations of plant detritus and soil organic matter (SOM) using stable isotope (13C) tracing coupled with a novel nuclear magnetic resonance (NMR) experiment. The amount of detritus-C degraded was not affected by the microbial diversity (p > 0.05), however the fate of detritus- and SOM-C across the diversity gradient was complex and land cover-dependent. For example, in the pasture soil, higher diversity led to lower CO2 production (p = 0.001), a trend driven solely by SOM-C mineralization. There was no relationship between diversity and detritus-C mineralization or production of new mineral-associations after one year (p > 0.05). In contrast, in the forest soil higher diversity resulted in increased detritus-C (p = 0.01) and SOM-C (p = 0.0008) mineralization and decreased mineral-associated organic matter formation (p = 0.02). In both land cover types, retention efficiency—a measure that integrates both microbial physiology and the ability of the ecosystem to retain C—explained C loss and transformation trends. Overall, this demonstrates that the trajectory of C gained and lost is altered by land management-induced changes to microbial communities, soil structure, and chemical characteristics underlying SOM persistence. [ABSTRACT FROM AUTHOR]

Greenhouse gas (GHG) emissions from thawed permafrost are difficult to predict because they result from complex interactions between abiotic drivers and multiple, often competing, microbial metabolic processes. Our objective was to characterize mechanisms controlling methane (CH) and carbon dioxide (CO) production from permafrost. We simulated permafrost thaw for the length of one growing season (90 days) in oxic and anoxic treatments at 1 and 15 °C to stimulate aerobic and anaerobic respiration. We measured headspace CH and CO concentrations, as well as soil chemical and biological parameters (e.g. dissolved organic carbon (DOC) chemistry, microbial enzyme activity, NO production, bacterial community structure), and applied an information theoretic approach and the Akaike information criterion to find the best explanation for mechanisms controlling GHG flux. In addition to temperature and redox status, CH production was explained by the relative abundance of methanogens, activity of non-methanogenic anaerobes, and substrate chemistry. Carbon dioxide production was explained by microbial community structure and chemistry of the DOC pool. We suggest that models of permafrost CO production are refined by a holistic view of the system, where the prokaryote community structure and detailed chemistry are considered. In contrast, although CH production is the result of many syntrophic interactions, these actions can be aggregated into a linear approach, where there is a single path of organic matter degradation and multiple conditions must be satisfied in order for methanogenesis to occur. This concept advances our mechanistic understanding of the processes governing anaerobic GHG flux, which is critical to understanding the impact the release of permafrost C will have on the global C cycle. [ABSTRACT FROM AUTHOR]

Previously-frozen stores of organic carbon (C) are now subject to decomposition due to a warming Arctic climate and associated permafrost thaw; however, estimates of the amount of greenhouse gases (GHG) that may be released are not well constrained. Knowing more about the functions of the extant permafrost microbial community will inform this knowledge gap. The exploration of microbial functional traits may be useful to elucidate the relationship between microbial diversity and ecosystem function. We characterized the community traits and functional diversity of the bacterial and Archaeal component of the microbial community from three depths of permafrost, as well as the organic and mineral horizons of the seasonally-thawed active layer, by assessing 'substrate-use richness,' 'substrate preference,' 'growth rate,' 'and substrate specific growth rate.' We measured the microbial community response to 31 substrates with an EcoPlate (Biolog, Inc.) assay at three incubation temperatures (1, 10, and 20 °C) using a kinetic approach, and modeled the microbial response to each substrate with a modified logistic growth function. We hypothesized that the permafrost communities would be selected for high functional potential and activity at cold temperatures. Rather, we found that the permafrost community did not have a higher functional diversity or activity at 1 °C than the organic active layer soils. In addition, permafrost communities increased their growth rates with increasing temperature, indicating that the highest incubation temperature (20 °C) was below their temperature optimum for growth. As predicted, the permafrost communities did exhibit temperature dependent substrate preferences. Thus, permafrost microbial communities did not appear to be selected for higher metabolism and the ability to use a broad suite of substrates at low temperatures, which suggests that they may have limited function immediately following thaw when temperatures are near 0 °C. However, changes in community composition or additional permafrost warming will increase the functional capabilities of permafrost microbes to decompose the C stored in those soils. [ABSTRACT FROM AUTHOR]

Although permafrost soils contain vast stores of organic C, relatively little is known about the chemical composition of their constituent soil organic matter (SOM). Mineral permafrost and organic (OAL) and mineral active layer (MAL) soils from Sagwon Hills, AK were analyzed for total C and N content and SOM chemical composition using Fourier transformed mid-infrared spectroscopy (MidIR). We also investigated techniques for proper collection of MidIR spectra on high C soils, such as permafrost. Principal Components Analysis (PCA) of the MidIR spectra revealed that the OAL was different from the MAL and permafrost based on absorbance of various organic functional groups, such as hydroxyls, alkyls, carbonyls, amines, amides, and esters. The top of the permafrost (0-15 cm below the maximum active layer thaw depth) was also differentiated from the deeper permafrost (16-40 cm below) by the same organic functional groups. Spectral data suggested that there is more chemically labile C (e.g., hydroxyl, amine groups, carbohydrates) in the OAL than the top of the permafrost, which in turn has more labile C than the MAL and deeper permafrost. The chemical similarity between the top of the permafrost and the OAL, and its differences with the MAL, suggest that organic matter (OM) is introduced into the permafrost through cryoturbation. All the soils showed evidence of microbial processing, such as organic acids and car-boxylates, however the relative abundance of these compounds varied by soil depth. This study advances our understanding of permafrost C chemistry and the reactivity of constituent compounds. [ABSTRACT FROM AUTHOR]

Global climate change is already having significant impacts on arctic and alpine ecosystems, and ongoing increases in temperature and altered precipitation patterns will affect the strong seasonal patterns that characterize these temperature-limited systems. The length of the potential growing season in these tundra environments is increasing due to warmer temperatures and earlier spring snow melt. Here, we compare current and projected climate and ecological data from 20 Northern Hemisphere sites to identify how seasonal changes in the physical environment due to climate change will alter the seasonality of arctic and alpine ecosystems. We find that although arctic and alpine ecosystems appear similar under historical climate conditions, climate change will lead to divergent responses, particularly in the spring and fall shoulder seasons. As seasonality changes in the Arctic, plants will advance the timing of spring phenological events, which could increase plant nutrient uptake, production, and ecosystem carbon (C) gain. In alpine regions, photoperiod will constrain spring plant phenology, limiting the extent to which the growing season can lengthen, especially if decreased water availability from earlier snow melt and warmer summer temperatures lead to earlier senescence. The result could be a shorter growing season with decreased production and increased nutrient loss. These contrasting alpine and arctic ecosystem responses will have cascading effects on ecosystems, affecting community structure, biotic interactions, and biogeochemistry. [ABSTRACT FROM AUTHOR]

This article discusses the importance of ethical and inclusive engagement between non-Indigenous researchers and Indigenous communities in Arctic research. It acknowledges the historical mistreatment of Indigenous peoples by researchers and emphasizes the need to build respectful relationships. The article provides ten rules for researchers to follow, including researching the culture and history of the research site, understanding different approaches to knowledge production, and establishing reciprocal community relationships. It highlights the value of Indigenous knowledge and the need for collaboration with Indigenous communities for high-quality research outcomes. The document offers guidelines and best practices for researchers seeking to collaborate with Arctic Indigenous communities, emphasizing the importance of recognizing shared humanity, respectful contact methods, aligning research questions with community objectives, compensating Indigenous participants, and planning for the continuation of research and scientific engagement. The paper underscores the need for ethical collaboration that centers Indigenous sovereignty, empowerment, and self-determination, while respecting Indigenous communities and their land and resources. [Extracted from the article]

The ability of microorganisms to resuscitate from dormancy has major implications for ecosystem function. By conceptualizing the space of possible definitions of dormancy as a multidimensional trait space, we construct a pluralistic concept of dormancy that accommodates and allows intercomparison between distinct definitions of dormancy from a variety of disciplines. Our framework for describing and comparing definitions of dormancy facilitates cross-communication between subfields of microbiology. Our framework can be applied to extreme cases, such as permafrost, where it allows for discriminating between microorganisms that are adaptively dormant, and those that are simply stressed. Life can be stressful. One way to deal with stress is to simply wait it out. Microbes do this by entering a state of reduced activity and increased resistance commonly called 'dormancy'. But what is dormancy? Different scientific disciplines emphasize distinct traits and phenotypic ranges in defining dormancy for their microbial species and system-specific questions of interest. Here, we propose a unified definition of microbial dormancy, using a broad framework to place earlier discipline-specific definitions in a new context. We then discuss how this new definition and framework may improve our ability to investigate dormancy using multi-omics tools. Finally, we leverage our framework to discuss the diversity of genomic mechanisms for dormancy in an extreme environment that challenges easy definitions – the permafrost. [ABSTRACT FROM AUTHOR]

Purpose: Perennial forage systems (e.g., pastures and hayfields) cover a significant land area worldwide and are important for soil carbon storage; however, the degree to which these plant communities can be managed to accumulate and store soil carbon is not well understood. Additionally, less is known about how forage defoliation mediates C dynamics or if the effects of defoliation depend on plant community composition. Methods: To address these questions, we quantified soil C pools and C-degrading enzyme activity in a three-year experiment where bicultures of alfalfa-orchardgrass and red clover-orchardgrass were managed under treatments of contrasting defoliation frequency (3 vs. 5 cuts per year) and severity (i.e., cutting height; 10 cm vs. 5 cm residual forage height). Results: We found that more frequent defoliation resulted in greater permanganate oxidizable C (POX-C), particulate organic carbon (POC), mineral-associated organic C (MAOC), and the activity of β-glucosidase (BG) and cellobiohydrolase (CBH). The effects of defoliation frequency on total organic C (TOC) varied depending on biculture composition. Alfalfa-orchardgrass bicultures, characterized by a lower legume-grass proportion, had greater POX-C and lower hot-water extractable organic carbon (HWEOC) compared to red clover-orchardgrass which had a higher legume-grass proportion. We also found that BG activity and POX-C were positively correlated with MAOC and TOC. In contrast, we observed no effects of defoliation severity on any measured soil parameters. Conclusion: Our study highlights that defoliation frequency and relative abundance of legumes to grass are both potential management levers for increasing SOC storage in legume-grass agroecosystems. [ABSTRACT FROM AUTHOR]

Permafrost thaw releases nutrients and metals from previously frozen soils and these nutrients may affect important biogeochemical processes including methane (CH4) production and oxidation. Here we assessed how concentrations of nutrients, solutes, and metals varied across four plant communities undergoing permafrost thaw and if these geochemical characteristics affected rates of CH4 production and oxidation. We tested nutrient limitation in CH4 production and oxidation by experimentally adding nitrogen (N), phosphorus (P) and a permafrost leachate to peat across these four plant communities. The upper 20 cm of permafrost contained 715 ± 298 mg m−2 of extractable inorganic N and 20 ± 6 mg m−2 of resin-extractable phosphorus (Presin), for a N:P ratio of 36:1. These low amounts of Presin coincide with high acid-digestible aluminum (Al), iron (Fe), and P concentrations in the permafrost soil and suggest that P may accumulate via sorption and constrain easily available forms of P for plants and microbes. Permafrost leachate additions decreased potential CH4 production rates up to 80% and decreased CH4 oxidation rates by 66%, likely due to inhibitory effects of N in the permafrost. In contrast, organic and inorganic P additions increased CH4 oxidation rates up to 36% in the tall graminoid fen, a community where phosphate availability was low and CH4 production was high. Our results suggest that (1) inorganic N is available immediately from permafrost thaw, while (2) P availability is controlled by sorption properties, and (3) plant community, nutrient stoichiometry, and metal availability modulate how permafrost thaw affects CH4 production and oxidation. [ABSTRACT FROM AUTHOR]

Predicting and mitigating changes in soil carbon (C) stocks under global change requires a coherent understanding of the factors regulating soil organic matter (SOM) formation and persistence, including knowledge of the direct sources of SOM (plants vs. microbes). In recent years, conceptual models of SOM formation have emphasized the primacy of microbial‐derived organic matter inputs, proposing that microbial physiological traits (e.g., growth efficiency) are dominant controls on SOM quantity. However, recent quantitative studies have challenged this view, suggesting that plants make larger direct contributions to SOM than is currently recognized by this paradigm. In this review, we attempt to reconcile these perspectives by highlighting that variation across estimates of plant‐ versus microbial‐derived SOM may arise in part from methodological limitations. We show that all major methods used to estimate plant versus microbial contributions to SOM have substantial shortcomings, highlighting the uncertainty in our current quantitative estimates. We demonstrate that there is significant overlap in the chemical signatures of compounds produced by microbes, plant roots, and through the extracellular decomposition of plant litter, which introduces uncertainty into the use of common biomarkers for parsing plant‐ and microbial‐derived SOM, especially in the mineral‐associated organic matter (MAOM) fraction. Although the studies that we review have contributed to a deeper understanding of microbial contributions to SOM, limitations with current methods constrain quantitative estimates. In light of recent advances, we suggest that now is a critical time to re‐evaluate long‐standing methods, clearly define their limitations, and develop a strategic plan for improving the quantification of plant‐ and microbial‐derived SOM. From our synthesis, we outline key questions and challenges for future research on the mechanisms of SOM formation and stabilization from plant and microbial pathways. [ABSTRACT FROM AUTHOR]

Rapid Arctic environmental change affects the entire Earth system as thawing permafrost ecosystems release greenhouse gases to the atmosphere. Understanding how much permafrost carbon will be released, over what time frame, and what the relative emissions of carbon dioxide and methane will be is key for understanding the impact on global climate. In addition, the response of vegetation in a warming climate has the potential to offset at least some of the accelerating feedback to the climate from permafrost carbon. Temperature, organic carbon, and ground ice are key regulators for determining the impact of permafrost ecosystems on the global carbon cycle. Together, these encompass services of permafrost relevant to global society as well as to the people living in the region and help to determine the landscape-level response of this region to a changing climate. [ABSTRACT FROM AUTHOR]

Permafrost thaw can alter the soil environment through changes in soil moisture, frequently resulting in soil saturation, a shift to anaerobic decomposition, and changes in the plant community. These changes, along with thawing of previously frozen organic material, can alter the form and magnitude of greenhouse gas production from permafrost ecosystems. We synthesized existing methane (CH4) and carbon dioxide (CO2) production measurements from anaerobic incubations of boreal and tundra soils from the geographic permafrost region to evaluate large-scale controls of anaerobic CO2 and CH4 production and compare the relative importance of landscape-level factors (e.g., vegetation type and landscape position), soil properties (e.g., pH, depth, and soil type), and soil environmental conditions (e.g., temperature and relative water table position). We found fivefold higher maximum CH4 production per gram soil carbon from organic soils than mineral soils. Maximum CH4 production from soils in the active layer (ground that thaws and refreezes annually) was nearly four times that of permafrost per gram soil carbon, and CH4 production per gram soil carbon was two times greater from sites without permafrost than sites with permafrost. Maximum CH4 and median anaerobic CO2 production decreased with depth, while CO2:CH4 production increased with depth. Maximum CH4 production was highest in soils with herbaceous vegetation and soils that were either consistently or periodically inundated. This synthesis identifies the need to consider biome, landscape position, and vascular/moss vegetation types when modeling CH4 production in permafrost ecosystems and suggests the need for longer-term anaerobic incubations to fully capture CH4 dynamics. Our results demonstrate that as climate warms in arctic and boreal regions, rates of anaerobic CO2 and CH4 production will increase, not only as a result of increased temperature, but also from shifts in vegetation and increased ground saturation that will accompany permafrost thaw. [ABSTRACT FROM AUTHOR]

Permafrost thaw could induce substantial carbon (C) emissions to the atmosphere, and thus trigger a positive feedback to climate warming. As the engine of biogeochemical cycling, soil microorganisms exert a critical role in mediating the direction and strength of permafrost C‐climate feedback. However, our understanding about the impacts of thermokarst (abrupt permafrost thaw) on microbial structure and function remains limited. Here we employed metagenomic sequencing to analyze changes in topsoil (0–15 cm) microbial communities and functional genes along a permafrost thaw sequence (1, 10, and 16 years since permafrost collapse) on the Tibetan Plateau. By combining laboratory incubation and a two‐pool model, we then explored changes in soil labile and stable C decomposition along the thaw sequence. Our results showed that topsoil microbial α‐diversity decreased, while the community structure and functional gene abundance did not exhibit any significant change at the early stage of collapse (1 year since collapse) relative to non‐collapsed control. However, as the time since the collapse increased, both the topsoil microbial community structure and functional genes differed from the control. Abundances of functional genes involved in labile C degradation decreased while those for stable C degradation increased at the late stage of collapse (16 years since collapse), largely driven by changes in substrate properties along the thaw sequence. Accordingly, faster stable C decomposition occurred at the late stage of collapse compared to the control, which was associated with the increase in relative abundance of functional genes for stable C degradation. These results suggest that upland thermokarst alters microbial structure and function, particularly enhances soil stable C decomposition by modulating microbial functional genes, which could reinforce a warmer climate over the decadal timescale. [ABSTRACT FROM AUTHOR]

The magnitude of carbon (C) loss to the atmosphere via microbial decomposition is a function of the amount of C stored in soils, the quality of the organic matter, and physical, chemical, and biological factors that comprise the environment for decomposition. The decomposability of C is commonly assessed by laboratory soil incubation studies that measure greenhouse gases mineralized from soils under controlled conditions. Here, we introduce the Soil Incubation Database (SIDb) version 1.0, a compilation of time series data from incubations, structured into a new, publicly available, open-access database of C flux (carbon dioxide, CO2 , or methane, CH4). In addition, the SIDb project also provides a platform for the development of tools for reading and analysis of incubation data as well as documentation for future use and development. In addition to introducing SIDb, we provide reporting guidance for database entry and the required variables that incubation studies need at minimum to be included in SIDb. A key application of this synthesis effort is to better characterize soil C processes in Earth system models, which will in turn reduce our uncertainty in predicting the response of soil C decomposition to a changing climate. We demonstrate a framework to fit curves to a number of incubation studies from diverse ecosystems, depths, and organic matter content using a built-in model development module that integrates SIDb with the existing SoilR package to estimate soil C pools from time series data. The database will help bridge the gap between point location measurements, which are commonly used in incubation studies, and global remote-sensed data or data products derived from models aimed at assessing global-scale rates of decomposition and C turnover. The SIDb version 1.0 is archived and publicly available at 10.5281/zenodo.3871263 (Sierra et al., 2020), and the database is managed under a version-controlled system and centrally stored in GitHub (https://github.com/SoilBGC-Datashare/sidb , last access: 26 June 2020). [ABSTRACT FROM AUTHOR]

Metal contamination of food and water resources is a known public health issue in Arctic and sub-Arctic communities due to the proximity of many communities to mining and drilling sites. In addition, permafrost thaw may release heavy metals sequestered in previously frozen soils, potentially contaminating food and water resources by increasing the concentration of metals in freshwater, plants, and wildlife. Here we assess the enrichment of selected heavy metals in Alaskan soils by synthesizing publicly available data of soil metal concentrations. We analyzed data of soil concentrations of arsenic, chromium, mercury, nickel, and lead from over 1,000 samples available through the USGS Alaskan Geochemical Database to evaluate 1) the spatial distribution of sampling locations for soil metal analysis, 2) metal concentrations in soils from different land cover types and depths, and 3) the occurrence of soils in Alaska with elevated metal concentrations relative to other soils. We found substantial clustering of sample sites in the southwestern portion of Alaska in discontinuous and sporadic permafrost, while the continuous permafrost zone in Northern Alaska and the more populous Interior are severely understudied. Metal concentration varied by land cover type but lacked consistent patterns. Concentrations of chromium, mercury, and lead were higher in soils below 10 cm depth, however these deeper soils are under-sampled. Arsenic, chromium, mercury, nickel and lead concentrations exceeded average values for US soils by one standard deviation or more in 3.7% to 18.7% of the samples in this dataset. Our analysis highlights critical gaps that impede understanding of how heavy metals in thawing permafrost soils may become mobilized and increase exposure risk for Arctic communities. [ABSTRACT FROM AUTHOR]

The magnitude of carbon (C) loss to the atmosphere via microbial decomposition is a function of the amount of C stored in soils, the quality of the organic matter, and physical, chemical and biological factors that comprise the environment for decomposition. The decomposability of C is commonly assessed by laboratory soil incubation studies that measure greenhouse gases mineralized from soils under controlled conditions. Here, we introduce the Soil Incubation Database (SIDb) version 1.0, a compilation of time series data from incubations, structured into a new, publicly available database of C flux (carbon dioxide, CO2, or methane, CH4). In addition to open access, the SIDb project also provides a platform for the development of tools for reading and analysis of incubation data as well as documentation for future use and development. In addition to introducing SIDb, we provide reporting guidance for database entry and the required variables that incubation studies need at minimum to be included in SIDb. A key application of this synthesis effort is to better characterize soil C processes in Earth system models, which will in turn reduce our uncertainty in predicting the response of soil C decomposition to a changing climate. We demonstrate a framework to fit curves to a number of incubation studies from diverse ecosystems, depths, and organic matter content using a built-in model development module that integrates SIDb with the existing SoilR package to estimate soil C pools from time series data. The database will help bridge the gap between site-level measurements, which are commonly used in incubation studies, and global remote-sensed data or data products derived from models aimed at assessing global-scale rates of decomposition and C turnover. [ABSTRACT FROM AUTHOR]

Accelerating permafrost thaw across high-latitude ecosystems increases organic matter (OM) availability to soil microorganisms. The chemical composition of permafrost-derived OM, which influences microbial metabolic efficiency, may determine whether the Arctic remains a critical carbon sink in the future or shifts to net a source of greenhouse gases. During a 90-day, simulated thaw experiment, we measured shifts in the chemistry of soil and dissolved organic matter pools sourced from surface (0–10 cm) and subsurface (16–25 cm) permafrost layers. We manipulated temperature (1 versus 15 °C) and redox (drained versus saturated) to mimic field-relevant conditions and tested whether variability in OM chemistry could be explained by the activities of six hydrolytic enzymes that catalyze carbon, nitrogen, and phosphorus mineralization. We found that permafrost depth and enzyme activities were significantly correlated with shifts in soil OM (SOM) functional group chemistries, as measured by diffuse reflectance infrared Fourier transform (DRIFT) spectroscopy. Greater variability in dissolved OM (DOM) chemistries were driven by redox and pH. Our results suggest depth-resolved analyses of permafrost vulnerability to enzymatic hydrolysis may help explain distinct patterns in SOM versus DOM cycling that can be used to constrain modeled projections of Arctic carbon storage during global change. • Hydrolytic enzymes strongly influenced permafrost soil organic matter chemistry. • Redox and pH explained shifts in dissolved organic matter chemistry. • DRIFT-detectable functional bands are early indicators of permafrost decomposition. • Different pathways of SOM versus DOM mineralization may influence Arctic C storage. [ABSTRACT FROM AUTHOR]

Conservation tillage practices can benefit soil health relative to conventional tillage; however, conservation tillage systems are often coupled with the use of pesticide seed treatments (PST; fungicides and insecticides, including neonicotinoids), which may undermine these benefits by disrupting microbial communities and processes. We examined the effects of tillage system (conventional moldboard plowing versus two conservation tillage practices: no tillage and strip tillage) and PST on microbial diversity and microbial function in a field experiment with a soybean–corn rotation. Enzyme activities and the microbial functional capacity to degrade a variety of substrates was intermediate in strip tillage compared with conventional and no tillage. Microbial community composition under strip tillage was similar to those under no tillage but distinct from conventional tillage, and community differences were associated with soil C/N and pH. However, tillage system had no effect on alpha diversity. Communities under strip tillage and no tillage had different substrate preferences to those under conventional tillage, which could be explained by the major C sources under each management practice. Pesticide seed treatment affected microbial substrate preference and altered the relative abundance of various fungal taxa, indicating a direct (i.e., toxic) effect. Despite this, PST had no effect on enzyme activities or alpha diversity of the microbial community. There were limited interactions between tillage and PST on microbial properties. Our results support that microbial communities are valuable indicators of soil health and show that strip tillage maintains soil microbial health better than conventional tillage but not as well as no tillage, and that PST may negatively affect soil microbial health. [Display omitted] • Microbial processes, such as elemental cycling, are fundamental to soil health. • Soil health under strip tillage was intermediate to no tillage and conventional tillage. • Tillage affected microbial structure and activity, but not diversity. • Pesticide seed treatments had a direct (toxic) effect on soil fungal communities. • Pesticide seed treatments may undermine some of the benefits of conservation tillage. [ABSTRACT FROM AUTHOR]

• POXC is an emerging soil health metric in need of methodological standardization. • Decreasing sieve size and increasing soil mass both increased POXC values. • Intra-lab variability and lower soil mass were the greatest sources of variability. • Decreased sieve size had a negligible effect on analytical variability. • Soils with lower SOC contents generally have higher variability. Soil organic matter is central to the soil health framework. Therefore, reliable indicators of changes in soil organic matter are essential to inform land management decisions. Permanganate oxidizable carbon (POXC), an emerging soil health indicator, has shown promise for being sensitive to soil management. However, strict standardization is required for widespread implementation in research and commercial contexts. Here, we used 36 soils—three from each of the 12 USDA soil orders—to determine the effects of sieve size and soil mass of analysis on POXC results. Using replicated measurements across 12 labs in the US and the EU (n = 7951 samples), we quantified the relative importance of 1) variation between labs, 2) variation within labs, 3) effect soil mass, and 4) effect of soil sieve size on the repeatability of POXC. We found a wide range of overall variability in POXC values across labs (0.03 to 171.8%; mean = 13.4%), and much of this variability was attributable to within-lab variation (median = 6.5%) independently of soil mass or sieve size. Greater soil mass (2.5 g) decreased absolute POXC values by a mean of 177 mg kg−1 soil and decreased analytical variability by 6.5%. For soils with organic carbon (SOC) >10%, greater soil mass (2.5 g) resulted in more frequent POXC values above the limit of detection whereas the lower soil mass (0.75 g) resulted in POXC values below the limit of detection for SOC contents <5%. A finer sieve size increased absolute values of POXC by 124 mg kg−1 while decreasing the analytical variability by 1.8%. In general, soils with greater SOC contents had lower analytical variability. These results point to potential standardizations of the POXC protocol that can decrease the variability of the metric. We recommend that the POXC protocol be standardized to use 2.5 g for soils <10% SOC. Sieve size was a relatively small contributor to analytical variability and therefore we recommend that this decision be tailored to the study purpose. Tradeoffs associated with these standardizations can be mitigated, ultimately providing guidance on how to standardize POXC for routine analysis. [ABSTRACT FROM AUTHOR]