Ammonia-oxidizing archaea and bacteria differentially contribute to ammonia oxidation in soil under precipitation gradients and land legacy.

Background: Global change has accelerated the nitrogen cycle. Soil nitrogen stock degradation by microbes leads to the release of various gases, including nitrous oxide (N2O), a potent greenhouse gas. Ammonia-oxidizing archaea (AOA) and ammonia-oxidizing bacteria (AOB) participate in the soil nitrogen cycle, producing N2O. There are outstanding questions regarding the impact of environmental processes such as precipitation and land use legacy on AOA and AOB structurally, compositionally, and functionally. To answer these questions, we analyzed field soil cores and soil monoliths under varying precipitation profiles and land legacies. Results: We resolved 28 AOA and AOB metagenome assembled genomes (MAGs) and found that they were significantly higher in drier environments and differentially abundant in different land use legacies. We further dissected AOA and AOB functional potentials to understand their contribution to nitrogen transformation capabilities. We identified the involvement of stress response genes, differential metabolic functional potentials, and subtle population dynamics under different environmental parameters for AOA and AOB. We observed that AOA MAGs lacked a canonical membrane-bound electron transport chain and F-type ATPase but possessed A/A-type ATPase, while AOB MAGs had a complete complex III module and F-type ATPase, suggesting differential survival strategies of AOA and AOB. Conclusions: The outcomes from this study will enable us to comprehend how drought-like environments and land use legacies could impact AOA- and AOB-driven nitrogen transformations in soil.

Home-field advantage affects the local adaptive interaction between Andropogon gerardii ecotypes and root-associated bacterial communities.

Due to climate change, drought frequencies and severities are predicted to increase across the United States. Plant responses and adaptation to stresses depend on plant genetic and environmental factors. Understanding the effect of those factors on plant performance is required to predict species' responses to environmental change. We used reciprocal gardens planted with distinct regional ecotypes of the perennial grass Andropogon gerardii adapted to dry, mesic, and wet environments to characterize their rhizosphere communities using 16S rRNA metabarcode sequencing. Even though the local microbial pool was the main driver of these rhizosphere communities, the significant plant ecotypic effect highlighted active microbial recruitment in the rhizosphere, driven by ecotype or plant genetic background. Our data also suggest that ecotypes planted at their homesites were more successful in recruiting rhizosphere community members that were unique to the location. The link between the plants' homesite and the specific local microbes supported the "home field advantage" hypothesis. The unique homesite microbes may represent microbial specialists that are linked to plant stress responses. Furthermore, our data support ecotypic variation in the recruitment of congeneric but distinct bacterial variants, highlighting the nuanced plant ecotype effects on rhizosphere microbiome recruitment. These results improve our understanding of the complex plant host-soil microbe interactions and should facilitate further studies focused on exploring the functional potential of recruited microbes. Our study has the potential to aid in predicting grassland ecosystem responses to climate change and impact restoration management practices to promote grassland sustainability. IMPORTANCE In this study, we used reciprocal gardens located across a steep precipitation gradient to characterize rhizosphere communities of distinct dry, mesic, and wet regional ecotypes of the perennial grass Andropogon gerardii. We used 16S rRNA amplicon sequencing and focused oligotyping analysis and showed that even though location was the main driver of the microbial communities, ecotypes could potentially recruit distinct bacterial populations. We showed that different A. gerardii ecotypes were more successful in overall community recruitment and recruitment of microbes unique to the "home" environment, when growing at their "home site." We found evidence for "home-field advantage" interactions between the host and host-root-associated bacterial communities, and the capability of ecotypes to recruit specialized microbes that were potentially linked to plant stress responses. Our study aids in a better understanding of the factors that affect plant adaptation, improve management strategies, and predict grassland function under the changing climate.

Novel complete methanogenic pathways in longitudinal genomic study of monogastric age-associated archaea.

BACKGROUND: Archaea perform critical roles in the microbiome system, including utilizing hydrogen to allow for enhanced microbiome member growth and influencing overall host health. With the majority of microbiome research focusing on bacteria, the functions of archaea are largely still under investigation. Understanding methanogenic functions during the host lifetime will add to the limited knowledge on archaeal influence on gut and host health. In our study, we determined lifelong archaea dynamics, including detection and methanogenic functions, while assessing global, temporal and host distribution of our novel archaeal metagenome-assembled genomes (MAGs). We followed 7 monogastric swine throughout their life, from birth to adult (1-156 days of age), and collected feces at 22 time points. The samples underwent gDNA extraction, Illumina sequencing, bioinformatic quality and assembly processes, MAG taxonomic assignment and functional annotation. MAGs were utilized in downstream phylogenetic analysis for global, temporal and host distribution in addition to methanogenic functional potential determination. RESULTS: We generated 1130 non-redundant MAGs, representing 588 unique taxa at the species level, with 8 classified as methanogenic archaea. The taxonomic classifications were as follows: orders Methanomassiliicoccales (5) and Methanobacteriales (3); genera UBA71 (3), Methanomethylophilus (1), MX-02 (1), and Methanobrevibacter (3). We recovered the first US swine Methanobrevibacter UBA71 sp006954425 and Methanobrevibacter gottschalkii MAGs. The Methanobacteriales MAGs were identified primarily during the young, preweaned host whereas Methanomassiliicoccales primarily in the adult host. Moreover, we identified our methanogens in metagenomic sequences from Chinese swine, US adult humans, Mexican adult humans, Swedish adult humans, and paleontological humans, indicating that methanogens span different hosts, geography and time. We determined complete metabolic pathways for all three methanogenic pathways: hydrogenotrophic, methylotrophic, and acetoclastic. This study provided the first evidence of acetoclastic methanogenesis in archaea of monogastric hosts which indicated a previously unknown capability for acetate utilization in methanogenesis for monogastric methanogens. Overall, we hypothesized that the age-associated detection patterns were due to differential substrate availability via the host diet and microbial metabolism, and that these methanogenic functions are likely crucial to methanogens across hosts. This study provided a comprehensive, genome-centric investigation of monogastric-associated methanogens which will further improve our understanding of microbiome development and functions.

Pseudomonas cultivated from Andropogon gerardii rhizosphere show functional potential for promoting plant host growth and drought resilience.

BACKGROUND: Climate change will result in more frequent droughts that can impact soil-inhabiting microbiomes (rhizobiomes) in the agriculturally vital North American perennial grasslands. Rhizobiomes have contributed to enhancing drought resilience and stress resistance properties in plant hosts. In the predicted events of more future droughts, how the changing rhizobiome under environmental stress can impact the plant host resilience needs to be deciphered. There is also an urgent need to identify and recover candidate microorganisms along with their functions, involved in enhancing plant resilience, enabling the successful development of synthetic communities. RESULTS: In this study, we used the combination of cultivation and high-resolution genomic sequencing of bacterial communities recovered from the rhizosphere of a tallgrass prairie foundation grass, Andropogon gerardii. We cultivated the plant host-associated microbes under artificial drought-induced conditions and identified the microbe(s) that might play a significant role in the rhizobiome of Andropogon gerardii under drought conditions. Phylogenetic analysis of the non-redundant metagenome-assembled genomes (MAGs) identified a bacterial genome of interest - MAG-Pseudomonas. Further metabolic pathway and pangenome analyses recovered genes and pathways related to stress responses including ACC deaminase; nitrogen transformation including assimilatory nitrate reductase in MAG-Pseudomonas, which might be associated with enhanced drought tolerance and growth for Andropogon gerardii. CONCLUSIONS: Our data indicated that the metagenome-assembled MAG-Pseudomonas has the functional potential to contribute to the plant host's growth during stressful conditions. Our study also suggested the nitrogen transformation potential of MAG-Pseudomonas that could impact Andropogon gerardii growth in a positive way. The cultivation of MAG-Pseudomonas sets the foundation to construct a successful synthetic community for Andropogon gerardii. To conclude, stress resilience mediated through genes ACC deaminase, nitrogen transformation potential through assimilatory nitrate reductase in MAG-Pseudomonas could place this microorganism as an important candidate of the rhizobiome aiding the plant host resilience under environmental stress. This study, therefore, provided insights into the MAG-Pseudomonas and its potential to optimize plant productivity under ever-changing climatic patterns, especially in frequent drought conditions.

Stability and volatility shape the gut bacteriome and Kazachstania slooffiae dynamics in preweaning, nursery and adult pigs.

The gut microbiome plays important roles in the maintenance of health and pathogenesis of diseases in the growing host. In order to fully comprehend the interplay of the gut microbiome and host, a foundational understanding of longitudinal microbiome, including bacteria and fungi, development is necessary. In this study, we evaluated enteric microbiome and host dynamics throughout the lifetime of commercial swine. We collected a total of 234 fecal samples from ten pigs across 31 time points in three developmental stages (5 preweaning, 15 nursery, and 11 growth adult). We then performed 16S rRNA gene amplicon sequencing for bacterial profiles and qPCR for the fungus Kazachstania slooffiae. We identified distinct bacteriome clustering according to the host developmental stage, with the preweaning stage exhibiting low bacterial diversity and high volatility amongst samples. We further identified clusters of bacteria that were considered core, increasing, decreasing or stage-associated throughout the host lifetime. Kazachstania slooffiae was absent in the preweaning stage but peaked during the nursery stage of the host. We determined that all host growth stages contained negative correlations between K. slooffiae and bacterial genera, with only the growth adult stage containing positive correlates. Our stage-associated bacteriome results suggested the neonate contained a volatile gut microbiome. Upon weaning, the microbiome became relatively established with comparatively fewer perturbations in microbiome composition. Differential analysis indicated bacteria might play distinct stage-associated roles in metabolism and pathogenesis. The lack of positive correlates and shared K. slooffiae-bacteria interactions between stages warranted future research into the interactions amongst these kingdoms for host health. This research is foundational for understanding how bacteria and fungi develop singularly, as well as within a complex ecosystem in the host's gut environment.

Bacterial but Not Fungal Rhizosphere Community Composition Differ among Perennial Grass Ecotypes under Abiotic Environmental Stress.

Environmental change, especially frequent droughts, is predicted to detrimentally impact the North American perennial grasslands. Consistent dry spells will affect plant communities as well as their associated rhizobiomes, possibly altering the plant host performance under environmental stress. Therefore, there is a need to understand the impact of drought on the rhizobiome, and how the rhizobiome may modulate host performance and ameliorate its response to drought stress. In this study, we analyzed bacterial and fungal communities in the rhizospheres of three ecotypes (dry, mesic, and wet) of dominant prairie grass, Andropogon gerardii. The ecotypes were established in 2010 in a common garden design and grown for a decade under persistent dry conditions at the arid margin of the species' range in Colby, Kansas. The experiment aimed to answer whether and to what extent do the different ecotypes maintain or recruit distinct rhizobiomes after 10 years in an arid climate. In order to answer this question, we screened the bacterial and fungal rhizobiome profiles of the ecotypes under the arid conditions of western Kansas as a surrogate for future climate environmental stress using 16S rRNA and ITS2 metabarcoding sequencing. Under these conditions, bacterial communities differed compositionally among the A. gerardii ecotypes, whereas the fungal communities did not. The ecotypes were instrumental in driving the differences among bacterial rhizobiomes, as the ecotypes maintained distinct bacterial rhizobiomes even after 10 years at the edge of the host species range. This study will aid us to optimize plant productivity through the use of different ecotypes under future abiotic environmental stress, especially drought. IMPORTANCE In this study, we used a 10-year long reciprocal garden system, and reports that different ecotypes (dry, mesic, and wet) of dominant prairie grass, Andropogon gerardii can maintain or recruit distinct bacterial but not fungal rhizobiomes after 10 years in an arid environment. We used both 16S rRNA and ITS2 amplicons to analyze the bacterial and fungal communities in the rhizospheres of the respective ecotypes. We showed that A. gerardii might regulate the bacterial community to adapt to the arid environment, in which some ecotypes were not adapted to. Our study also suggested a possible tradeoff between the generalist and the specialist bacterial communities in specific environments, which could benefit the plant host. Our study will provide insights into the plant host regulation of the rhizosphere bacterial and fungal communities, especially during frequent drought conditions anticipated in the future.

Perspective: Simple State Communities to Study Microbial Interactions: Examples and Future Directions.

Microbial interactions in natural environments are intricately complex. High numbers and rich diversity of microorganisms, along with compositional heterogeneities complicate the cause. It is essential to simplify these complex communities to understand the microbial interactions. We proposed a concept of "simple state community," which represents a subset of microbes and/or microbial functions of the original population that is necessary to build a stable community. By combining microbial culturing and high-throughput sequencing, we can better understand microbe-microbe and microbe-host interactions. To support our proposed model, we used carbon-based and nitrogen-based media to capture the simple state communities. We used 16S rRNA amplicon sequencing and assigned taxonomic identity to the bacterial populations before and after simple state communities. We showed that simple state communities were a subset of the original microbial communities at both phyla and genera level. We further used shotgun metagenomics to gain insights into the functional potential of the assembled simple state communities. Our proposed model supported the goal of simplifying the complex communities across diverse systems to provide opportunity to facilitate comprehension of both the structure and function of the subset communities. Further applications of the concept include the high-throughput screening of simple state communities using the BIOLOG® system and continuous culturing (Chemostat). This concept has the potential to test diverse experimental hypotheses in simplified microbial communities, and further extend that knowledge to answer the overarching questions at a more holistic level.

Conduit-assisted Allograft Neurorrhaphy for the Treatment of Intractable Lower Extremity Pain Due to Neuromas-in-continuity.

We present a novel technique for the management of intractable lower extremity pain, due to neuromas-in-continuity of two peripheral nerves, through combined neurectomies proximal to the zone of initial injury and subsequent bridging utilizing an allograft-coupled conduit construct. METHODOLOGY: A retrospective chart review of 36 patients (18 women and 18 men) with recalcitrant nerve pain secondary to neuromas-in-continuity of two peripheral nerves following lower extremity trauma was conducted. Subjects underwent superficial peroneal nerve (SPN) to deep peroneal nerve neurorrhaphy (19 patients) or SPN to sural nerve neurorrhaphy (17 patients) proximal to the zone of initial injury. Patient demographics, comorbidities, procedure details, complications, and preoperative and postoperative pain assessments using a visual analog scale were evaluated. RESULTS: Residual nerve pain from previous lower extremity trauma was included. Analysis of preprocedure and postprocedure visual analog scale scores demonstrated a mean decrease of 7.45 points (mean: pre 8.89, mean: post 1.44). All patients voiced satisfaction with postoperative ambulatory tolerance and pain relief at last follow-up (mean: 30.86 months). CONCLUSIONS: The sequelae of neuromas-in-continuity of the SPN, deep peroneal nerve, and sural nerves were noted to have significantly improved with proximal neurectomy and subsequent bridging utilizing a nerve allograft and conduit construct. We present this coaptation technique as a viable treatment option for reduction in neurogenic pain involving peripheral nerve injury of two dermatome distributions.

Surgical Treatment Protocol for Peripheral Nerve Dysfunction of the Lower Extremity: A Systematic Approach.

To date, more than 150 surgical techniques have been described for the treatment of intractable nerve pain. However, owing to their technical complexity, as well as the lack of comparative studies in the literature, there is currently no consensus on the appropriate management of this often debilitating condition. Therefore, we present our surgical algorithm, based on Seddon's classification to differentiate the degree of nerve injury, and subsequent treatment course for the management of lower extremity neurogenic pain.

Understanding the Arterial Anatomy and Dermal Perfusion of the Lower Extremity with Clinical Application.

Respecting vascular territories is crucial for flap design and viability. There are 6 angiosomes in the foot and ankle originating from the posterior tibial, anterior tibial, and peroneal arteries. Reliable perforators from the 3 major source vessels of the lower extremity are found in 5-cm intervals within intermuscular septa. Computed tomography angiography, Doppler, and infrared cameras can be used preoperatively to help identify the perforators of the lower extremity.