Plant microbiomes are an essential component of the plant host and are widely known to ameliorate environmental stress response. In the California Sierra Nevada, the endemic cutleaf monkeyflower (Mimulus laciniatus) is experiencing and adapting to various levels of stress associated with climate change, however, it is unclear to what extent the M. laciniatus microbiome is engaged in responding to this stress. This thesis aims to determine the monkeyflower microbiome along with their growing substrate (moss or soil) across a steep elevation gradient in the species range during the 2019 growing season. Specifically, we collected monkeyflower plants and divided those collections into compartment subsamples (roots, shoots, soil, and moss substrate) to assess the bacterial communities of each compartment by 16S rRNA gene sequencing to answer the following: (i) Do the M. laciniatus endophyte communities differ in composition and diversity across the M. laciniatus species range, and does this composition vary by elevation? (ii) Does the M. laciniatus endophyte community differ between plant compartments (e.g. root and shoot)? (iii) Do soil and moss, from which M. laciniatus grows, serve as a source of M. laciniatus endophytes, and do soil and moss communities vary in structure across the M. laciniatus species range? We found that the bacterial community structure of both plant compartments (roots and shoots) and the substrate (soil, and moss) differed across the range of M. laciniatus. Compartment strongly influenced bacterial communities, supporting the idea that plant compartments provide micro-habitats for bacterial communities. Elevation played a role in community variation when all samples were analyzed together. When compartment subsets were analyzed separately, population and site (nested in population) were significant drivers of variation in all four compartments. The differences between sites and population may be explained by the effect on bacterial communities by local climate, which could in turn reflect that the plant host selects beneficial microbes to support plant responses to the local environment, leading to habitat-adapted symbiosis. Alternatively, differences in bacterial community structure across the M. laciniatus species range could be due to differences in the metacommunity of locally available bacterial taxa to colonize compartments. This thesis highlights the importance of plant compartment and local climate or metacommunity in shaping bacterial communities along the elevation gradient of plant species.

The overall aim of my dissertation was to determine if subalpine conifers host a consistent core of endophytic bacteria. Endophytic bacteria, symbiotic bacteria found within healthy plant tissue, have been found in all plants studied to date. Endophytic bacteria can influence plant healthy through growth promotion, resistance to pests and pathogens, and confer plasticity to abiotic and biotic stress. Despite the role they can play in plant health, they have largely been examined in agricultural crops with little exploration of endophytic bacteria in long-lived forest trees. Forest trees may rely on bacterial endophytes to confer robustness to variable and environmental conditions they face over their lifetime. Subalpine conifers are long-lived trees that inhabit nutrient limited soils and harsh environments and therefore a good candidate to examine endophytic bacteria that may confer host robustness. In animals, core bacterial symbionts with important roles in host health are often consistently associated with individuals of a host species. To uncover potential beneficial endophytic bacteria, I used next-generation sequencing of the 16S rRNA bacterial gene to determine if subalpine conifers were host to a consistent core of endophytes. I found Pinus flexilis and Picea engelmannii at Niwot Ridge, CO were consistently dominated by a potential N2 fixing phylotype (AAB1). Next, to determine if the consistent AAB1 association was a result of shared environment (Niwot Ridge) or shared ancestry (Pinus and Picea), I characterized the needle endophytic community of P. flexilis and Pinus contorta at two distance sites: Niwot Ridge, CO and Horseshoe Meadows, CA. I found AAB1 to dominate the endophytic bacterial communities across both species and sites, suggesting the association was not dependent only on environment but may be a long-term association of AAB1 and Pinus and Picea. To further examine the distribution of AAB1 I examined the endophytic community of P. contorta at Tuolumne Meadows, CA. AAB1was present but not in high abundance, indicating the AAB1 association is facultative. I found that the remaining endophyte community (i.e. excluding AAB1) of P. flexilis, P. engelmanni and P. contorta was structured by geographic location. In contrast to the high elevation pines, a core and consistent endophytic bacteria taxa was not identified in Sequoia sempervirens and Sequoiadendron giganteum in the family Cupressaceae . In these species, the endophyte communities were highly variable within and across tree individuals and species. These results suggest that subalpine conifers in the Pinaceae associate with facultative but symbiotic N2 fixing endophytic bacteria, and that the symbiosis may have originated after the divergence of Pinaceae and the Cupressaceae.

Microbial organisms are important in all ecosystems, yet their diversity and distributions in different natural environments still need to be assessed. This doctoral thesis focuses on the study of macroecological distribution patterns and environmental constraints acting over different microbial groups inhabiting ephemeral ecosystems —vernal pools. Vernal pools are temporal wetlands, with contrasting desiccation periods and water availability periods; fundamentally they are well-delimited ecosystems, embedded in completely terrestrial habitats and populated by specialized organisms. 16s Hightroughput-sequencing was used for the analysis of Environmental DNA (eDNA) collected from different substrates in vernal pool ecosystems to investigate the microbial communities living in these endangered ecosystems.