Microbial community shifts reflect losses of native soil carbon with pyrogenic and fresh organic matter additions and are greatest in low-carbon soils.

Soil organic carbon (SOC) plays an important role in regulating global climate change, carbon and nutrient cycling in soils, and soil moisture. Organic matter (OM) additions to soils can affect the rate at which SOC is mineralized by microbes, with potentially important effects on SOC stocks. Understanding how pyrogenic organic matter (PyOM) affects the cycling of native SOC (nSOC) and the soil microbes responsible for these effects is important for fire-affected ecosystems as well as for biochar-amended systems. We used an incubation trial with five different soils from National Ecological Observatory Network sites across the US and 13C-labelled 350°C corn stover PyOM and fresh corn stover OM to trace nSOC-derived CO2 emissions with and without PyOM and OM amendments. We used high-throughput sequencing of rRNA genes to characterize bacterial, archaeal, and fungal communities and their response to PyOM and OM in soils that were previously stored at -80°C. We found that the effects of amendments on nSOC-derived CO2 reflected the unamended soil C status, where relative increases in C mineralization were greatest in low-C soils. OM additions produced much greater effects on nSOC-CO2 emissions than PyOM additions. Furthermore, the magnitude of microbial community composition change mirrored the magnitude of increases in nSOC-CO2, indicating a specific subset of microbes were likely responsible for the observed changes in nSOC mineralization. However, PyOM responders differed across soils and did not necessarily reflect a common "charosphere". Overall, this study suggests that soils that already have low SOC may be particularly vulnerable to short-term increases in SOC loss with OM or PyOM additions.Importance Soil organic matter (SOM) has an important role in global climate change, carbon and nutrient cycling in soils, and soil moisture dynamics. Understanding the processes that affect SOM stocks is important for managing these functions. Recently, understanding how fire-affected organic matter (or "pyrogenic" organic matter (PyOM)) affects existing SOM stocks has become increasingly important, both due to changing fire regimes, and to interest in "biochar" - pyrogenic organic matter that is produced intentionally for carbon management or as an agricultural soil amendment. We found that soils with less SOM were more prone to increased losses with PyOM (and fresh organic matter) additions, and that soil microbial communities changed more in soils that also had greater SOM losses with PyOM additions. This suggests that soils that already have low SOM content may be particularly vulnerable to short-term increases in SOM loss, and that a subset of the soil microbial community is likely responsible for these effects.

Root Carbon Interaction with Soil Minerals Is Dynamic, Leaving a Legacy of Microbially Derived Residues.

Minerals preserve the oldest, most persistent soil carbon, and mineral characteristics appear to play a critical role in the formation of soil organic matter (SOM) associations. To test the hypothesis that roots, and differences in carbon source and microbial communities, influence mineral SOM associations over short timescales, we incubated permeable mineral bags in soil microcosms with and without plants, inside a 13CO2 labeling chamber. Mineral bags contained quartz, ferrihydrite, kaolinite, or soil minerals isolated via density separation. Using 13C-nuclear magnetic resonance, Fourier transform ion cyclotron resonance mass spectrometry, and lipidomics, we traced carbon deposition onto minerals, characterizing total carbon, 13C enrichment, and SOM chemistry over three growth stages of Avena barbata. Carbon accumulation was rapid and mineral-dependent but slowed with time; the accumulated amount was not significantly affected by root presence. However, plant roots strongly shaped the chemistry of mineral-associated SOM. Minerals incubated in a plant rhizosphere were associated with a more diverse array of compounds (with different functional groups-carbonyl, aromatics, carbohydrates, and lipids) than minerals incubated in an unplanted bulk soil control. We also found that many of the lipids that sorbed to minerals were microbially derived, including many fungal lipids. Together, our data suggest that diverse rhizosphere-derived compounds may represent a transient fraction of mineral SOM, rapidly exchanging with mineral surfaces.

Microbial community assembly differs across minerals in a rhizosphere microcosm.

Mineral-associated microbes drive many critical soil processes, including mineral weathering, soil aggregation and cycling of mineral-sorbed organic matter. To investigate the interactions between soil minerals and microbes in the rhizosphere, we incubated three types of minerals (ferrihydrite, kaolinite and quartz) and a native soil mineral fraction near roots of a common Californian annual grass, Avena barbata, growing in its resident soil. We followed microbial colonization of these minerals for up to 2.5 months - the plant's lifespan. Bacteria and fungi that colonized mineral surfaces during this experiment differed across mineral types and differed from those in the background soil, implying that microbial colonization was the result of processes in addition to passive movement with water to mineral surfaces. Null model analysis revealed that dispersal limitation was a dominant factor structuring mineral-associated microbial communities for all mineral types. Once bacteria arrived at a mineral surface, capacity for rapid growth appeared important, as ribosomal copy number was significantly correlated with relative enrichment on minerals. Glomeromycota (a phylum associated with arbuscular mycorrhizal fungi) appeared to preferentially associate with ferrihydrite surfaces. The mechanisms enabling the colonization of soil minerals may be foundational in shaping the overall soil microbiome composition and development of persistent organic matter in soils.

Spatiotemporal dynamics of the bacterial microbiota on lacustrine Cladophora glomerata (Chlorophyta).

The branched periphytic green alga Cladophora glomerata, often abundant in nearshore waters of lakes and rivers worldwide, plays important ecosystem roles, some mediated by epibiotic microbiota that benefit from host-provided surface, organic C, and O2 . Previous microscopy and high-throughput sequencing studies have indicated surprising epibiont taxonomic and functional diversity, but have not included adequate consideration of sample replication or the potential for spatial and temporal variation. Here, we report the results of 16S rRNA amplicon-based phylum-to-genus taxonomic analysis of Cladophora-associated bacterial epibiota sampled in replicate from three microsites and at six times during the open-water season of 2014, from the same lake locale (Picnic Point, Lake Mendota, Dane Co., WI, USA) explored by high-throughput sequencing studies in two previous years. Statistical methods were used to test null hypotheses that the bacterial community: (i) is homogeneous across microsites tested, and (ii) does not change over the course of a growth season or among successive years. Results indicated a dynamic microbial community that is more strongly influenced by sampling day during the growth season than by microsite variation. A surprising diversity of bacterial genera known to be associated with the key function of methane-oxidation (methanotrophy), including relatively high-abundance of Crenothrix, Methylomonas, Methylovulum, and Methylocaldum-showed intraseasonal and interannual variability possibly related to temperature differences, and microsite preferences possibly related to variation in methane abundance. By contrast, a core assemblage of bacterial genera seems to persist over a growth season and from year to year, possibly transmitted by a persistent attached host resting stage.

Carbon mineralizability determines interactive effects on mineralization of pyrogenic organic matter and soil organic carbon.

Soil organic carbon (SOC) is a critical and active pool in the global C cycle, and the addition of pyrogenic organic matter (PyOM) has been shown to change SOC cycling, increasing or decreasing mineralization rates (often referred to as priming). We adjusted the amount of easily mineralizable C in the soil, through 1-day and 6-month preincubations, and in PyOM made from maple wood at 350 °C, through extraction. We investigated the impact of these adjustments on C mineralization interactions, excluding pH and nutrient effects and minimizing physical effects. We found short-term increases (+20-30%) in SOC mineralization with PyOM additions in the soil preincubated for 6 months. Over the longer term, both the 6-month and 1-day preincubated soils experienced net ∼10% decreases in SOC mineralization with PyOM additions. Additionally, the duration of preincubation affected interactions, indicating that there may be no optimal preincubation time for SOC mineralization studies. We show conclusively that mineralizability of SOC in relation to PyOM-C is an important determinant of the effect of PyOM additions on SOC mineralization.

Corrigendum to "An open-source, automated, gas sampling peripheral for laboratory incubation experiments using cavity ring-down spectroscopy" [HardwareX 10 (2021) e00208].

[This corrects the article DOI: 10.1016/j.ohx.2021.e00208.].

Experimentally determined traits shape bacterial community composition one and five years following wildfire.

Wildfires represent major ecological disturbances, burning 2-3% of Earth's terrestrial area each year with sometimes drastic effects above- and belowground. Soil bacteria offer an ideal, yet understudied system within which to explore fundamental principles of fire ecology. To understand how wildfires restructure soil bacterial communities and alter their functioning, we sought to translate aboveground fire ecology to belowground systems by determining which microbial traits are important post-fire and whether changes in bacterial communities affect carbon cycling. We employed an uncommon approach to assigning bacterial traits, by first running three laboratory experiments to directly determine which microbes survive fires, grow quickly post-fire and/or thrive in the post-fire environment, while tracking CO2 emissions. We then quantified the abundance of taxa assigned to each trait in a large field dataset of soils one and five years after wildfires in the boreal forest of northern Canada. We found that fast-growing bacteria rapidly dominate post-fire soils but return to pre-burn relative abundances by five years post-fire. Although both fire survival and affinity for the post-fire environment were statistically significant predictors of post-fire community composition, neither are particularly influential. Our results from the incubation trials indicate that soil carbon fluxes post-wildfire are not likely limited by microbial communities, suggesting strong functional resilience. From these findings, we offer a traits-based framework of bacterial responses to wildfire.

Disturbance by soil mixing decreases microbial richness and supports homogenizing community assembly processes.

The spatial heterogeneity of soil's microhabitats warrants the study of ecological patterns and community assembly processes in the context of physical disturbance that disrupts the inherent spatial isolation of soil microhabitats and microbial communities. By mixing soil at various frequencies in a 16-week lab incubation, we explored the effects of physical disturbance on soil bacterial richness, community composition, and community assembly processes. We hypothesized that well-mixed soil would harbor a less rich microbial community, with community assembly marked by homogenizing dispersal and homogeneous selection. Using 16S rRNA gene sequencing, we inferred community assembly processes, estimated richness and differential abundance, and calculated compositional dissimilarity. Findings supported our hypotheses, with > 20% decrease in soil bacterial richness in well-mixed soil. Soil mixing caused communities to diverge from unmixed controls (Bray-Curtis dissimilarity; 0.75 vs. 0.25), while reducing within-group heterogeneity. Our results imply that the vast diversity observed in soil may be supported by spatial heterogeneity and isolation of microbial communities, and also provide insight into the effects of physical disturbance and community coalescence events. By isolating and better understanding the effects of spatial heterogeneity and disconnectivity on soil microbial communities, we can better extrapolate how anthropogenic disturbances may affect broad soil functions.

Effects of physical, chemical, and biological ageing on the mineralization of pine wood biochar by a Streptomyces isolate.

If biochar is to be used for carbon (C) management, we must understand how weathering or ageing affects biochar C mineralization. Here, we incubated aged and unaged eastern white pine wood biochar produced at 350 and 550°C with a Streptomyces isolate, a putative biochar-decomposing microbe. Ageing was accelerated via three different processes, namely, (a) physical ageing-subjecting biochar to alternating freeze-thaw and wet-dry cycles, (b) chemical ageing-treating biochar with concentrated hydrogen peroxide and (c) biological ageing-incubating biochar in the presence of nutrients and microorganisms. Elemental composition and surface chemistry (Fourier Transform Infrared spectroscopy) of biochar samples were compared before and after ageing. Biochar C mineralization between ageing treatments was significantly different in the case of 350°C biochar (p value = 0.03). Among the 350°C biochars, physical ageing resulted in the greatest increase (by 103%) in biochar C mineralization (p value = 0.05). However, in the case of 550°C biochar, ageing did not result in a significant change in biochar C mineralization (p value = 0.40). Biochar C mineralization was positively correlated with an increase in O/C ratio post-ageing (rs = 0.86, p value = 0.01). In the case of 350°C biochar, surface oxidation during ageing enhanced biochar degradation by the isolate. For 550°C biochar, however, ageing did not significantly increase biochar C mineralization, likely due to high condensed aromatic C content and lower surface oxidation during ageing. The results from our study suggest that low temperature aged biochar is more susceptible to biological degradation by soil microbes. These findings have implications for the use of biochar for long term C storage in soils.

Effects of short-interval reburns in the boreal forest on soil bacterial communities compared to long-interval reburns.

Increasing fire frequency in some biomes is leading to fires burning in close succession, triggering rapid vegetation change and altering soil properties. We studied the effects of short-interval (SI) reburns on soil bacterial communities of the boreal forest of northwestern Canada using paired sites (n = 44). Both sites in each pair had burned in a recent fire; one site had burned within the previous 20 years before the recent fire (SI reburn) and the other had not. Paired sites were closely matched in prefire ecosite characteristics, prefire tree species composition, and stand structure. We hypothesized that there would be a significant effect of short vs. long fire-free intervals on community composition and that richness would not be consistently different between paired sites. We found that Blastococcus sp. was consistently enriched in SI reburns, indicating its role as a strongly 'pyrophilous' bacterium. Caballeronia sordidicola was consistently depleted in SI reburns. The depletion of this endophytic diazotroph raises questions about whether this is contributing to-or merely reflects-poor conifer seedling recolonization post-fire at SI reburns. While SI reburns had no significant effect on richness, dissimilarity between short- and long-interval pairs was significantly correlated with difference in soil pH, and there were small significant changes in overall community composition.

Climate change impact of biochar cook stoves in western Kenyan farm households: system dynamics model analysis.

Cook stoves that produce biochar as well as heat for cooking could help mitigate indoor air pollution from cooking fires and could enhance local soils, while their potential reductions in carbon (C) emissions and increases in soil C sequestration could offer access to C market financing. We use system dynamics modeling to (i) investigate the climate change impact of prototype and refined biochar-producing pyrolytic cook stoves and improved combustion cook stoves in comparison to conventional cook stoves; (ii) assess the relative sensitivity of the stoves' climate change impacts to key parameters; and (iii) quantify the effects of different climate change impact accounting decisions. Simulated reductions in mean greenhouse gas (GHG) impact from a traditional, 3-stone cook stove baseline are 3.50 tCO(2)e/household/year for the improved combustion stove and 3.69-4.33 tCO(2)e/household/year for the pyrolytic stoves, of which biochar directly accounts for 26-42%. The magnitude of these reductions is about 2-5 times more sensitive to baseline wood fuel use and the fraction of nonrenewable biomass (fNRB) of off-farm wood that is used as fuel than to soil fertility improvement or stability of biochar. Improved cookstoves with higher wood demand are less sensitive to changes in baseline fuel use and rely on biochar for a greater proportion of their reductions.

An open-source, automated, gas sampling peripheral for laboratory incubation experiments using cavity ring-down spectroscopy.

Spectroscopic instruments are becoming increasingly popular for measuring the isotopic composition and fluxes of a wide variety of gases in both field and laboratory experiments. The popularity of these instruments has created a need for automated multiplexers compatible with the equipment. While there are several such peripherals commercially available, they are currently limited to only a small number of samples (≤16), which is insufficient for some studies. To support researchers in constructing custom, larger-scale systems, we present our design for a scalable gas sampling peripheral that can be programmed to autonomously sample up to 56 vessels - the "multiplexer". While originally designed to be used with a Picarro cavity ring-down spectroscopy (CRDS) system, the multiplexer design and data processing approach implemented can be easily adapted to serve as a gas sampling/delivery platform for a wide variety of instruments including other cavity ring-down systems and infra-red gas analyzers. We demonstrate the basic capabilities of the multiplexer by using it to autonomously sample head-space CO2 from 14 laboratory-incubated soils amended with 13C-enriched pyrogenic organic matter for analysis in a Picarro G2201-i cavity ring-down spectroscopy system.

Affordable Diffusion Microchamber Array Designs for Isolating Microbes in Classrooms and Laboratories.

Diffusion microchamber array (DMA) devices have provided contemporary microbiologists with a successful alternative to the century-old plating method using petri dishes to isolate and cultivate environmental bacteria. The "ichip" DMA device, developed by the research group of Slava Epstein, is exemplary of this technology, using membranes to culture rare or otherwise nonmodel bacterial taxa, whereby the target cells are nourished by environmental medium across the separating membrane. However, DMA devices have not sufficiently infiltrated the university-level microbiology curriculum, and the size and shape of the ichip make the device challenging for use by students. The following work provides guidance in the creation and use of a suite of DMA devices called "iplates," which are ichip-like tools for use in microbiology education. An iplate mimics the shape and size of a 96-well plate, making it a simpler, more affordable, and customizable design for the isolation and incubation of bacteria and other microorganisms using fresh environmental media. Iplates and their many customizations are intended to enable teachers, students, and researchers to isolate, grow, and analyze the widest possible diversity of microorganisms, complementing standard plating methods. We offer additional tips and tools to help instructors and students get started creating DMA devices to meet their unique needs and research interests.

Pyrolyzed Substrates Induce Aromatic Compound Metabolism in the Post-fire Fungus, Pyronema domesticum.

Wildfires represent a fundamental and profound disturbance in many ecosystems, and their frequency and severity are increasing in many regions of the world. Fire affects soil by removing carbon in the form of CO2 and transforming remaining surface carbon into pyrolyzed organic matter (PyOM). Fires also generate substantial necromass at depths where the heat kills soil organisms but does not catalyze the formation of PyOM. Pyronema species strongly dominate soil fungal communities within weeks to months after fire. However, the carbon pool (i.e., necromass or PyOM) that fuels their rise in abundance is unknown. We used a Pyronema domesticum isolate from the catastrophic 2013 Rim Fire (CA, United States) to ask whether P. domesticum is capable of metabolizing PyOM. Pyronema domesticum grew readily on agar media where the sole carbon source was PyOM (specifically, pine wood PyOM produced at 750°C). Using RNAseq, we investigated the response of P. domesticum to PyOM and observed a comprehensive induction of genes involved in the metabolism and mineralization of aromatic compounds, typical of those found in PyOM. Lastly, we used 13C-labeled 750°C PyOM to demonstrate that P. domesticum is capable of mineralizing PyOM to CO2. Collectively, our results indicate a robust potential for P. domesticum to liberate carbon from PyOM in post-fire ecosystems and return it to the bioavailable carbon pool.

Dynamics of microbial community composition and soil organic carbon mineralization in soil following addition of pyrogenic and fresh organic matter.

Pyrogenic organic matter (PyOM) additions to soils can have large impacts on soil organic carbon (SOC) cycling. As the soil microbial community drives SOC fluxes, understanding how PyOM additions affect soil microbes is essential to understanding how PyOM affects SOC. We studied SOC dynamics and surveyed soil bacterial communities after OM additions in a field experiment. We produced and mixed in either 350 °C corn stover PyOM or an equivalent initial amount of dried corn stover to a Typic Fragiudept soil. Stover increased SOC-derived and total CO2 fluxes (up to 6x), and caused rapid and persistent changes in bacterial community composition over 82 days. In contrast, PyOM only temporarily increased total soil CO2 fluxes (up to 2x) and caused fewer changes in bacterial community composition. Of the operational taxonomic units (OTUs) that increased in response to PyOM additions, 70% also responded to stover additions. These OTUs likely thrive on easily mineralizable carbon (C) that is found both in stover and, to a lesser extent, in PyOM. In contrast, we also identified unique PyOM responders, which may respond to substrates such as polyaromatic C. In particular, members of Gemmatimonadetes tended to increase in relative abundance in response to PyOM but not to fresh organic matter. We identify taxa to target for future investigations of the mechanistic underpinnings of ecological phenomena associated with PyOM additions to soil.

A dual-isotope approach to allow conclusive partitioning between three sources.

Stable isotopes have proved to be a transformative tool; their application to distinguish between two sources in a mixture has been a cornerstone of biogeochemical research. However, quantitatively partitioning systems using two stable isotopes (for example, (13)C and (12)C) has been largely limited to only two sources, and systems of interest often have more than two components, with interactive effects. Here we introduce a dual-isotope approach to allow conclusive partitioning between three sources, using only two stable isotopes. We demonstrate this approach by partitioning soil CO2 emissions derived from microbial mineralization of soil organic carbon (SOC), added pyrogenic organic matter (PyOM) and root respiration. We find that SOC mineralization in the presence of roots is 23% higher (P<0.05) when PyOM is also present. Being able to discern three sources with two isotopes will be of great value not only in biogeochemical research, but may also expand hitherto untapped methodologies in diverse fields.

Characterization of biochars to evaluate recalcitrance and agronomic performance.

Biochars (n=94) were found to have ash contents from 0.4% to 88.2%, volatile matter from 13.2% to 70.0%, and fixed carbon from 0% to 77.4% (w/w). Greater pyrolysis temperature for low-ash biochars increased fixed carbon, but decreased it for biochars with more than 20% ash. Nitrogen recovery varied depending on feedstock used to a greater extent (12-68%) than organic (25-45%) or total C (41-76%) at a pyrolysis temperature of 600 °C. Fixed carbon production ranged from no enrichment in poultry biochar to a 10-fold increase in corn biochar (at 600 °C). Prediction of biochar stability was improved by a combination of volatile matter and H:C ratios corrected for inorganic C. In contrast to stability, agronomic utility of biochars is not an absolute value, as it needs to meet local soil constraints. Woody feedstock demonstrated the greatest versatility with pH values ranging from 4 to 9.