Browsing by Author "Wrighton, Kelly C., committee member"

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Open Access
Analyzing longitudinal patterns of river metabolism in five distinct rivers
(Colorado State University. Libraries, 2021) Schmer, Natalie K., author; Ross, Matthew R. V., advisor; Wrighton, Kelly C., committee member; Covino, Tim P., committee member
The River Continuum and Serial Discontinuity Concepts are two common frameworks or theories to describe river ecosystem structure and function. While these concepts help explain how rivers should behave, the reality is that aquatic ecosystems are being changed and degraded due to dams which regulate flow and changes to land use and land cover that can negatively impact aquatic ecosystems. Ecosystem metabolism is an increasingly common way to monitor and assess river health and functioning. This analysis is a data synthesis to examine longitudinal coherence in productivity patterns and identify the strongest controls that disrupt coherence. I find that between discharge, dissolved oxygen, and metabolism, metabolism is the least predictable variable going downstream regardless of land use and river characteristics, but at the same time did show patterns consistent with the expected patterns of the River Continuum Concept or other theories of river function variation. This lack of coherence and predictability in river metabolism across sites within rivers highlights effects monitoring methods, river-specific characteristics, and the overall lack of satisfactory theories for how larger rivers behave, despite an abundance of riverine theories.
Open Access
Crop domestication impacts on rhizosphere interactions and nitrogen acquisition
(Colorado State University. Libraries, 2024) Hwang, Siwook, author; Fonte, Steven J., advisor; Machmuller, Megan B., committee member; Crews, Timothy E., committee member; Wrighton, Kelly C., committee member; Boot, Claudia M., committee member
Synthetic nitrogen (N) fertilizer is an essential pillar of modern industrial agriculture. Production and application of synthetic N fertilizer, however, are two of the most expensive, energy intensive, and environmentally deleterious processes in agriculture. Therefore, alternative means of providing N in an agroecosystem are of great interest in sustainable agriculture. While many solutions – from cover cropping to intercropping – have been suggested over time it remains unclear if the modern high-yielding crops can thrive in these alternative N conditions. Decades of breeding under high synthetic N input as well as the inherently annual nature of these modern cereal crops may prevent them from fully taking advantage of these alternative N sources. In this dissertation, I explored the impact of domestication on crop rhizosphere interactions and N acquisition, in both retrospective and prospective terms. First, I investigated how modern maize (Zea mays subsp. mays), and its wild relative Teosinte (Zea mays subsp. parviglumis) differed in their ability to adapt to, and take up, cover crop residue N and synthetic N inputs. We designed a 13C (carbon)/15N dual isotope labeling experiment in which we compared the C allocation patterns of modern maize and teosinte in response to synthetic (urea) and organic (cover crop residue) forms of N. Teosinte responded to organic N by increasing its biomass root-to-shoot (R:S) ratio by 50% compared to synthetic N, while modern maize maintained the same biomass R:S ratios in both N treatments. Recent photosynthate R:S ratio (measured using 13C-CO2, 7 weeks after establishment) was greater in organic N than in synthetic N treatments for both modern maize and teosinte (91% and 37%; respectively). Label-derived dissolved organic C (DOC), representing recent rhizodeposits, was 2.5 times greater in the organic N treatments for both genotypes. Modern maize took up a similar amount of organic N as teosinte using different C allocation strategies. Our findings suggest that intensive breeding under high N input conditions has not affected this modern maize hybrid's access to organic N sources while improving its ability to take up synthetic N. Next, I shifted my focus to the novel perennial grains Kernza and perennial wheat. Kernza® is a domesticated intermediate wheatgrass (IWG, Thinopyrum intermedium). Perennial wheat is a hybrid between Kernza/IWG and modern annual durum wheat (Triticum turgidum subsp. durum). Kernza, in addition to being a perennial, may still possess beneficial belowground traits that may have been lost in modern cereals through millennia of aboveground-focused plant breeding. If so, such traits may be passed down to perennial wheat. To characterize root architecture, exudate profiles, and microbial communities of Kernza and perennial wheat in relation to annual wheat, I conducted a greenhouse experiment. We grew three genotypes/species (Kernza, perennial wheat, annual wheat) and collected their root exudates after 8 weeks of growth. The exudates were analyzed via LC-MS/MS for their chemical composition. We extracted DNA from rhizosphere soils and sequenced them for 16S and ITS profiles. Lastly, we scanned the roots to analyze root distribution across different diameter classes. We found that perennial wheat invested more heavily into very fine (< 250 µm) roots compared to annual wheat and Kernza. Perennial wheat also exuded at a greater rate of exudates per amount of root biomass. We suspect that the greater proportion of very fine roots in perennial wheat led to greater surface area and greater specific exudation rate, and that this may be related to hybrid vigor. We did not find evidence of a genotype effect on root exudate or microbial community composition. However, root exudates (overall metabolite profiles) significantly correlated with root architecture (distribution of root volume over different diameter classes) and the microbial community composition. These interactions represent a potential pathway through which plants can exert influence over the rhizosphere microbial community. Overall, these results emphasize the importance of root architecture in mediating belowground interactions. Understanding rhizosphere dynamics and the response to domestication and hybridization can guide further development of robust perennial cereal crops. In a third experiment, I studied how Kernza, perennial wheat, and annual responded to cereal-legume intercropping (biculture) in the field. To do so, we planted each of the three genotypes in monoculture or in biculture with alfalfa (Medicago sativa). We sampled their rhizosphere over two growing seasons and extracted soil DNA to construct rhizosphere 16S and ITS profiles. We hypothesized that 1) rhizosphere microbial community composition of annual wheat and Kernza will be most dissimilar from each other with perennial wheat intermediate, and 2) microbial community composition will shift in biculture, with the greatest change in Kernza and the smallest in annual wheat. We found that the rhizosphere 16S profiles differed significantly from the other two genotypes but the 16S profile of perennial wheat did not differ from that of annual wheat. Perennial wheat seemingly inherited microbial recruitment traits of its annual parent more so than its perennial parent's. Interestingly, inclusion of legumes led to the convergence, rather than divergence, of 16S profiles among genotypes. We postulate that the competitive pressure of alfalfa may have led to this convergence of 16S profiles across genotypes. The fungal community did not show evidence of genotype effect. However, the fungal community composition changed over two years in monoculture but not in biculture. This result implies that fungal community may become distinct over time if it is influenced by only one genotype (i.e., monoculture) rather than two (i.e., biculture). In conclusion, we found evidence of genotype-driven microbial community assembly that changed with legume's competitive pressure. The inheritability of microbial assembly was present but skewed towards the annual parent. Our study demonstrates the importance of including rhizosphere interactions in our evaluation of novel cereal crops in and out of cereal-legume biculture. In a final study, I investigated how rhizosphere microbial ecology of these three genotypes (Kernza, annual wheat, and perennial wheat) could be linked to their ability to acquire N from neighboring alfalfa plants. We designed a greenhouse study in which we planted all three cereals in monoculture or in biculture with alfalfa and used 15N leaf feeding technique to track the movement of N from alfalfa to cereals. In addition, we also extracted DNA from the soil and sequenced it for 16S rRNA profiles. Arbuscular mycorrhizal fungi (AMF) infection rate was also measured on all cereals and legumes. We hypothesized that: 1) annual wheat would produce the greatest biomass but Kernza would have highest proportion of legume derived N in its biomass, 2) all microbial communities will shift in biculture, with the greatest change in Kernza and the smallest in annual wheat, and 3) Kernza would have the highest rate of infection from AMF, especially in biculture. Surprisingly, we found no evidence of genotype or cropping system (monoculture or biculture) effect on either proportion or absolute amount of N derived from legume. We did find, however, that DOC concentration was higher in cereal rhizosphere grown in biculture than in monoculture, suggesting greater belowground investment in exudates when the grasses are grown with a legume. Despite this trend, annual wheat had much lower microbial biomass carbon (MBC) level in its rhizosphere compared to the perennials, in biculture. We contended that this may be due to substrate suitability of annual wheat's rhizodeposit. We also found that AMF infection rate was in fact the lowest in Kernza. Lastly, we found that 16S profiles of all three cereals shifted towards that of alfalfa in biculture. This trend might suggest microbial spillover, wherein rhizosphere microbial community of one genotype colonizes that of a neighboring plant, from alfalfa rhizosphere. Overall, we demonstrated that quantifying the N transfer in the rhizosphere can provide important insight into how these genotypes may be inducing changes in soil biogeochemistry in response to neighboring legumes. In summation, this dissertation provides links between crop genotype, root exudate chemistry and rate, microbial community assembly, and their biogeochemical consequences, in alternative N environments. Deepened understanding of how complex rhizosphere interactions may affect internal N cycling could be leveraged to further optimize these unique systems such as perennial cereal-legume biculture. In doing so, we will be one step closer to a more sustainable future, that is less reliant on synthetic N fertilizers.
Open Access
Metagenomic insights into microbial colonization & persistence in subsurface fractured shales
(Colorado State University. Libraries, 2023) Amundson, Kaela K., author; Wilkins, Michael J., advisor; Wrighton, Kelly C., committee member; Borch, Thomas, committee member; Ross, Matthew, committee member
Microorganisms are pervasive yet important components of hydraulically fractured shale systems. Subsurface shales harbor oil & gas and require unconventional techniques, such as hydraulic fracturing, to access these trapped hydrocarbons. Shale microbiomes are of crucial importance as they can directly impact the recovery of oil & gas and associated infrastructure. The overarching theme of this dissertation was to characterize the metabolisms and key traits that underpin the colonization and persistence of fractured shale microbiomes using a multi-omic approach to better understand the microbial impact on this important ecosystem. In Chapter 1, I first discuss the importance of subsurface shales as an important energy reserve, summarize what is known about microorganisms in these ecosystems, and highlight the strength of using a metagenomic approach to studying shale microbiomes. Subsurface shales are heterogeneous – varying in their mineral content, temperature, and other physiochemical conditions. The microbial communities that persist can have substantial impact on the fractured shale ecosystem and contribute to common challenges in hydrocarbon recovery such as corrosion, souring, and bioclogging. The literature review presented here highlights the need to study the functional potential of shale microbiomes as most studies have mostly focused solely on taxonomic composition of persisting microbial communities, and a vast majority of these studies have focused on samples from wells in the Appalachian Basin. However, functional potential of shale microbiomes across a variety of physiochemical conditions must be considered in order to gain an understanding of the role of microorganisms and what possible influences they may have on hydraulically fractured shales systems. Here, I highlight the need (1) to study the whole community at a functional scale and (2) apply a metagenomic approach to a variety of less characterized shale basins to gain a holistic understanding of shale microbiomes and the effects they may have on the broader ecosystem. In Chapter 2, I apply this metagenomic approach to study the persisting shale microbiomes of three fractured shale wells in the Anadarko Basin – a western shale basin characterized by elevated temperature and salinity. No studies using metagenomics had been applied to shale basins in the western United States prior to this research. We sampled five wells in the Anadarko basin over a timeseries >500 days and preformed NMR metabolomics and metagenomic sequencing to uncover the dominant metabolisms, community composition, and other functional traits of the Anadarko shale microbiome. This system was dominated by a fermentative microbial community and a less-abundant sub-community of inferred sulfate reducing microorganisms. Using paired NMR metabolomics and metagenomics, I demonstrated how many fermentative microorganisms have the potential to degrade common complex polymers, such as guar gum, and have potential to produce organic acids that may serve as electron donors for sulfate and thiosulfate reducing microorganisms. Thus, in this study I provided a framework for how carbon may move through the closed fractured shale ecosystem to sustain the microbial community. Finally, I investigated viral presence and diversity across all thirty-six metagenomes and found that inputs were large sources of viral diversity, but that only an extremely small proportion of viruses recovered from produced fluids were genetically similar to viruses previously reported from fractured shales. I observed that a majority of the dominant and persisting genomes encoded a CRISPR-Cas viral defense system, likely in response to the viral community. This highlights viral defense as another key trait for persisting microorganisms, as viruses are the only predators to bacteria and archaea in fractured shale ecosystems. Overall, this study expanded our knowledge of sulfate and thiosulfate reducing microorganisms in fractured shales, demonstrated the potential for common chemical inputs such as guar gum to be utilized by shale microbiomes, and highlighted how other key traits, such as CRISPR-Cas viral defense systems, may be a crucial trait for persisting shale microbiomes. Building on results from viral analyses in Chapter 2, in Chapter 3 I next sought to investigate the temporal dynamics between hosts and viruses to better understand the role of microbial defense against viruses in fractured shale ecosystems. To do this, I sampled six shale wells in the Denver-Julesburg Basin for >800 days, performed metagenomic sequencing, and identified host (bacterial & archaeal) and viral genomes from this data. I observed evidence of ongoing host defense to viral predation at both the community and genome-level through quantifying spacers from CRISPR arrays in metagenomic reads and MAGs. Through these analyses leveraging timeseries sampling and age differences between the shale wells, I provided evidence that suggested migration toward CRISPR arrays that may be more efficient at protecting the microbial host against a wider suite of viruses. Finally, I observed a temporal increase in host-viral co-existence in the closed, fractured shale ecosystem – suggesting the CRISPR defense does not entirely protect against viral predation. Chapter 4 ultimately leverages the approaches, insights, and data gained from Chapters 2 and 3 to study shale microbiomes at a cross-basin, geographic scale. Here, I collected samples from many collaborators who have previously worked in shale systems, performed metagenomic sequencing, and processed all samples in a standardized pipeline to build a comprehensive genomic shale database. In total, this database contains 978 unique MAGs and >7 million unique genes recovered from 209 metagenomic samples obtained from 36 fractured shale wells spanning eleven shale basins from North America, China, and the United Kingdom. In this chapter I analyze the functional potential of shale microbiomes at a genome-resolved level to better understand the geographic distribution of microbial metabolisms and other key traits that likely contribute to colonization and persistence of microorganisms. Here I also leveraged bioinformatic tools to build a custom annotation summary toolkit to process and analyze the large amount of sequencing data for traits of interest. The complete absence of a taxonomic core microbiome across shale basins illustrated in this chapter underscores the necessity of a genome-resolved and functional approach to studying shale microbiomes. Results from analyzing shale microbiomes at this scale could ultimately help to inform microbial management of fractured shale systems. The final chapter of this dissertation (Chapter 5) summarizes the key findings of my research into fractured shale microbiomes, and the mechanisms that may promote microbial colonization, persistence, and survival in these relatively harsh and economically relevant ecosystems. In this chapter I conclude this work by discussing future directions and lingering knowledge gaps for studying fractured shale microbiomes, as well as implications of these findings for other subsurface engineered ecosystems. Ultimately, this body of work contributes a substantial amount of new and informative insights into the functional potentials of persisting shale microbiomes across broad geographic scales.