Browsing by Subject "microbial ecology"

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The biogeography, ecophysiology, and functional potential of phototrophic Chloroflexi in alkaline hot springs: from marker genes to metagenomes
(2022-07) Bennett, Annastacia
Alkaline hot springs in Yellowstone National Park (YNP) provide a framework to study the relationship between photoautotrophs and temperature. Previous work has focused on understanding how Cyanobacteria (oxygenic phototrophs) vary with temperature, sulfide, and pH – but many questions remain regarding the ecophysiology of anoxygenic photosynthesis due to the taxonomic and metabolic diversity of these taxa. Phototrophs within the Cyanobacteria and Chloroflexi groups are frequently observed in alkaline hot springs. Decades of research has determined that temperature constrains Cyanobacteria in alkaline hot springs, but factors that constrain the distribution of phototrophic Chloroflexi remain unresolved. In Chapter 1, I review the key findings from Chloroflexi isolate and in situ studies in the two hot springs that have been the focus of decades of work in YNP. I highlight the metabolic and ecological diversity of characterized phototrophic Chloroflexi as it relates to nitrogen fixation, carbon cycling, and sulfur cycling. Additionally, I introduce how foundational studies have informed next generation sequencing efforts in this dissertation and in other works. In Chapter 2, I examined the distribution of genes involved in phototrophy, carbon fixation, and nitrogen fixation in eight alkaline (pH 7.3-9.4) hot spring sites near the upper temperature limit of photosynthesis (~71ºC) in YNP using metagenome sequencing. Based on genes encoding key reaction center proteins, geographic isolation plays a larger role than temperature in selecting for distinct phototrophic Chloroflexi while genes typically associated with autotrophy in anoxygenic phototrophs did not have distinct distributions with temperature. However, I recovered Calvin Cycle gene variants associated with Chloroflexi, an alternative carbon fixation pathway in anoxygenic photoautotrophs. Lastly, I recovered several abundant nifH (nitrogen fixation gene) sequences associated with Roseiflexus providing further evidence that genes involved in nitrogen fixation in Chloroflexi are more common than previously assumed. Together, these results add to the body of work focused on the distribution and functional potential of phototrophic bacteria in Yellowstone National Park hot springs and support the hypothesis that a combination of abiotic and biotic factors impact the distribution of phototrophic bacteria in hot springs. In Chapter 3, I employed a combination of 16S rRNA gene sequencing and inorganic carbon photoassimilation microcosms, to test the hypothesis that temperature would constrain the activity and composition of phototrophic Cyanobacteria and Chloroflexi. I expected diversity and rates of photoassimilation to decrease with increasing temperature. I report 16S rRNA amplicon sequencing along with carbon isotope signatures and photoassimilation from 45-72ºC in two alkaline hot springs. I found that Roseiflexus, Chloroflexus (Chloroflexi) and Leptococcus (Cyanobacteria) operational taxonomic units (OTUs) have distinct distributions with temperature. This distribution suggests that, like phototrophic Cyanobacteria, temperature selects for specific phototrophic Chloroflexi taxa. The richness of phototrophic Cyanobacteria decreased with increasing temperature along with a decrease in oxygenic photosynthesis, whereas Chloroflexi richness and rates of anoxygenic photosynthesis did not decrease with increasing temperature, even as temperatures approaches the upper limit of photosynthesis (~72 - 73ºC). Our carbon isotopic data suggest an increasing prevalence of 3-hydroxypropionate bicycle (3-HPB) with decreasing temperature coincident with photoautotrophic Chloroflexi. Together these results indicate temperature plays a role in defining the niche space of phototrophic Chloroflexi (as has been observed for Cyanobacteria), but other factors such as morphology, geochemistry, or metabolic diversity of Chloroflexi, in addition to temperature, could determine the niche space of this highly versatile group. Finally, in Chapter 4, I build on the work in Rabbit Creek from Chapter 3 by conducting a phylogenomic and pangenome analysis of 17 Chloroflexales metagenome assembled genomes (MAGs). I hypothesized that Chloroflexus and Roseiflexus would harbor unique core genomes and that temperature would select for certain accessory genes. I built a phylogenomic tree with 17 Rabbit Creek MAGs and 15 NCBI isolate genomes and found that the Rabbit Creek MAGs represent genera with characterized isolates as well as novel taxa. I examined the functional potential of Roseiflexus and Chloroflexus MAGs and found that Roseiflexus core genomes were depleted in both 3-HPB and sulfide oxidation functions while Chloroflexus core genomes contained sulfide:quinone oxidoreductases (SQRs) and several steps in the 3-HPB. Furthermore, I found both Roseiflexus and Chloroflexus core genomes contained genes for carbon assimilation and storage, suggesting this is an integral function for Chloroflexi in the Rabbit Creek ecosystem. Lastly, I recovered several MAGs that did not align with reference genomes but contained genes for type-II reaction centers and thiosulfate oxidation. Additionally, the unclassified MAGs were more like other Rabbit Creek MAGs. This work builds on the body of work in phototrophic Chloroflexi in alkaline hot springs and further constrains the role of Chloroflexi in carbon and sulfur cycling.
Discovering Ecological and Evolutionary Principles Governing Microbial Community Responses to Bacteriophage Infection of a Cross-Feeding Synthetic Coculture and Implications for Phage-based Applications
(2020-08) Fazzino, Lisa
Bacterial viruses, called bacteriophage (phage), infect bacteria and alter microbial community structure. Phages are an untapped resource to manipulate agriculture and medically applicable microbial communities. Yet, we cannot predict how phage impact a microbial community. My research aims to uncover ecological and evolutionary principles governing responses of microbial communities that contain cross-feeding interactions, where one species provides nutrients to (‘feeds’) another, phage. I combine wet-lab experiments on an engineered microbial co-culture with mathematical modeling to explore aspects of phage infection that are difficult to manipulate experimentally. I use a cross-feeding bacterial co-culture with Escherichia coli (E. coli) and Salmonella enterica (S. enterica) bacterial strains. In this cross-feeding system, E. coli cannot produce methionine, but does produce acetate and galactose. E. coli is paired with S. enterica that over-produces methionine and consumes acetate and galactose that E. coli secretes. To this co-culture, I add phage that infect either species. I have asked how simple cross-feeding co-cultures respond to phage infection. In Chapter 2, I used mathematical modeling and wet-lab experiments to show that single phage infections can break the cross-feeding relationship by liberating nutrients previously sequestered in the infected bacterial cells, ultimately changing community composition, and that partial, not full, resistance was necessary for this effect. In Chapter 3, ‘cocktails’ made of two different phage suppressed community growth the longest in a novel formulation that targeted both the pathogenic bacterial species and the slowest growing cross-feeder. Mathematical modeling showed that this was a generalizable concept to all cross-feeding systems. In Chapter 4, despite impacting community structure, I found that long term co-evolution between phage and E. coli cross-feeding with S. enterica only had weak effects on rates of adaptation. Phage treatments tended to increase rates of adaptations, as predicted by the Red Queen hypothesis, and cross-feeding tended to decrease rates of adaptation, as predicted by the Red King hypothesis. Overall, this thesis helps set baseline expectations of how phage influence cross-feeding microbial communities.
Ecology of interspecies signaling among Streptomyces and its relationship to pathogen suppression
(2013-05) Vaz Jauri, Patricia
Interspecies signaling may be defined as the induced change in phenotype of one species by another that is not due to the metabolism of the signal. Although suggested to be a relatively widespread phenomenon, the role of signaling in natural soil communities has not been thoroughly studied. Within Streptomyces communities in soil, understanding the impacts of interspecies signaling on species interactions, and especially on nutrient competition and antagonism, may be key to effective Streptomyces-based suppression of plant pathogens. I evaluated the frequency of signaling interactions and their effect on inhibitory phenotypes of Streptomyces isolated from natural prairies. Signaling among Streptomyces was frequent, and observed in 35% of all interactions. Isolates from the same location in soil were more likely to signal one another than isolates from different locations, suggesting local selection for signaling interactions. Signaling was similarly more frequent between isolates that had similar nutrient use profiles. Finally, closely-related isolates were more likely to increase inhibition towards one another via signaling than distantly-related isolates. In chapter 2, subinhibitory concentrations of antibiotics were studied as signals, specifically in relation to their capacities to shift nutrient use among Streptomyces. We found that some antibiotics altered nutrient use by Streptomyces in ways that could reduce nutrient competition among isolates. Finally, pathogen suppression and signaling were evaluated in soils with different cropping histories. Pathogen suppression by Streptomyces varied significantly among soils, and suppressive activity was positively correlated with bacterial density. Among Streptomyces from these plots, shifts in inhibitory phenotypes in response to signaling by another isolate were very frequent (~ 50% of all interactions). Overall, signaling in Streptomyces is frequent and varies with spatial origin, nutrient overlap, antagonistic phenotype, and genetic relatedness among isolates, as well as soil cropping history. Moreover, some antibiotics have the potential to act as signals that can significantly alter nutrient competition among Streptomyces. Variation in signaling has significant potential to mediate pathogen suppression in soil communities.
The Effect of Abiotic and Biotic Factors, Symbiont Exchange Between Host Species, and Host Migration on Fungal Symbiont Community Composition and Diversity
(2021-10) Watson, Monica
Microbiomes, the ubiquitous communities of microbial symbionts residing within hosts, play important roles in host health, development, and fitness. While recent research has characterized the microbiomes of many different host species, our understanding of how environmental factors affect microbial community dynamics is still in nascent stages. In this dissertation, I investigate how biotic and abiotic environmental factors affect the diversity and composition of microbial symbiont communities within different host species. In chapter 1, I use a full factorial experimental plot design and culture-based methods to examine the effects of nutrient addition, large animal herbivore exclusion, and host tissue specificity on the fungal symbiont communities, known as endophytes, residing within the grass species Andropogon gerardii. While neither nutrient addition nor herbivore exclusion alone significantly affect the diversity or composition of culturable endophytes, in combination, nutrient addition and herbivore exclusion were associated with greater fungal symbiont diversity than found in other treatments. Further, while different host tissues harbored distinct fungal communities, diversity was greater in all plant host tissues sampled from plots with both nutrient addition and herbivore exclusion treatments. In chapter 2, using field collections and both culture- and sequence-based methods I compare fungal diversity and community composition in a migratory agricultural pest insect, Spodoptera frugiperda, its plant host, Sorghum bicolor, and soil collected beneath infested host plants. Finding fungal communities in insects were much more variable compared to fungal communities in plants and soil, I estimated contributions of these differing sources of fungal symbionts to the insect microbiome. Surprisingly, I find that insect fungal communities were more commonly attributed to other insect sources than to the plants on which they were feeding or to soil sources. In chapter 3, I examine the fungal symbiont communities in an overwintering population of S. frugiperda to the fungal symbiont of a migratory population to ask whether fungal symbiont communities differ over the course of migration. Specifically, I ask if the prevalence and abundance of entomopathogenic taxa is different in overwintering and migratory populations. In an analysis of environmental sources of insect fungal symbionts, I examined the fungal symbiont communities of plant hosts on which they feed at the overwintering and migratory sites. Fungal communities were surprisingly similar over the course of migration in insect hosts. While most fungal entomopathogens occurred in similar prevalence and abundance, there were two OTUs that were significantly different in abundance in the two states with one more abundant in overwintering populations and one more abundant in migratory populations. There was little evidence of fungal symbiont exchange between insects and plants. Together, these chapters characterize how both abiotic and biotic environmental factors affect fungal symbiont communities in plants and insects, the extent to which fungal symbionts may transmit among different hosts and ecological compartments, and how migration impacts microbial symbiont communities. This work has important implications for our understanding of the factors affecting microbial symbiont community dispersal and our ability to predict the effect of the environment on microbial symbiont communities in agriculturally significant species.
Elevated Carbon Dioxide Alters the Structure of Soil Microbial Communities
(2012) Deng, Ye; He, Zhili; Xu, Meiying; Qin, Yujia; Van Nostrand, Joy D; Wu, Liyou; Roe, Bruce A; Wiley, Graham; Hobbie, Sarah E; Reich, Peter B; Zhou, Jizhong
Pyrosequencing analysis of 16S rRNA genes was used to examine impacts of elevated CO2(eCO(2)) on soil microbial communities from 12 replicates each from ambient CO2(aCO(2)) and eCO(2) settings. The results suggest that the soil microbial community composition and structure significantly altered under conditions of eCO(2), which was closely associated with soil and plant properties.
High-performance tools for precise microbiome characterization
(2018-08) Al-Ghalith, Gabriel
The microbiome, defined as the vast number of microorganisms inhabiting both human and non-human environments, has been associated with human disease as well as other important ecological phenomena. However, its quantitative study is complicated in part by measurement error and computational limitations, pointing to a need for more sensitive and reproducible DNA sequence analysis techniques. To this end, I have developed a variety of improved methods including a flexible short-read quality control pipeline, curated databases of marker genes and whole genomes, streamlined OTU picking software, and a high-throughput optimal aligner with taxonomy interpolation. Together, these methods represent advancements over traditional sequence analysis pipelines and may improve the quality of downstream statistical analyses.
Linking inhibitory phenotypes of soil Streptomyces to resource inputs, resource-use tradeoffs, and soil microbiome composition and diversity
(2019-01) Gieske, Miriam
Soil bacteria produce a diverse array of antibiotics which mediate interactions among microbes. Resource competition and tradeoffs between inhibitory ability and growth are thought to be important in shaping the evolution of inhibitory phenotypes, but tests of these ideas in naturally-occurring microbial populations remain scarce. In my dissertation, I used soil-borne Streptomyces isolates from a long-term agricultural experiment to examine the relationships between resource inputs (nitrogen fertilizer and crop residues), resource use traits, and inhibitory phenotypes, as well as relationships between Streptomyces inhibitory phenotypes and the diversity and composition of the soil microbiome. I found that long-term nitrogen addition resulted in lower frequencies of antibiotic inhibitory phenotypes among indigenous Streptomyces in nitrogen fertilized plots than in non-fertilized plots, while crop residue incorporation had only limited effects on inhibitor frequencies. Streptomyces isolates with greater ability to inhibit other Streptomyces had lower niche width and mean growth across carbon sources they utilized. This tradeoff between inhibitory ability and resource use was consistent across plots with different histories of nitrogen and crop residue inputs. The frequency of inhibitory phenotypes among Streptomyces was correlated with abundances of many individual OTUs from diverse phyla. Inhibitor frequency was also correlated with soil pH, but not with OTU richness or diversity. Taken together, my findings suggest that resource inputs can have substantial effects on the frequencies of antibiotic-producing microbes in soil microbial communities. However, these effects appear to mediated in complex ways by microbial densities, soil physicochemical characteristics, and/or changes in the composition of the soil microbiome. Further research is needed to better understand the mechanisms by which resource inputs affect the evolution of inhibitory capacities among naturally-occurring soil microbial populations.

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