The influence of environmental variation on the microbiome during early-life stages in reptiles

Abstract

My dissertation research seeks to understand how environmental variation, including maternal effects, might influence the microbiome of reptiles and how those differences translate to phenotypic variation. My research framework integrates both observational and experimental science through field and lab-based methods. Documenting environmentally mediated changes in the microbiome and their effects on hosts will provide a robust foundation for understanding the role of microbiome plasticity in shaping host phenotypes, including growth, physiology, and behavior. For my first chapter, I sought to understand how gut homeostasis is influenced by environmental variation (in the form of aquatic pollutants like estrogen). I experimentally assigned 23 hatchling American alligators (Alligator mississippiensis) to three ecologically relevant treatments (control, low, and high estrogen concentrations) for ten weeks. Gut microbial samples were collected following diet treatments and microbial diversity was determined using 16S rRNA gene-sequencing. Individuals in estrogen-treatment groups had decreased microbial diversity, but a greater relative abundance of operational taxonomic units than those in the control group. This effect was dose-dependent; as individuals were exposed to more estrogen, their microbiota became less diverse, less rich, and less even. Findings from this study suggest that environmental contamination can influence wildlife populations at the internal, microbial level, which may lead to future deleterious health effects. For my second chapter, I sought to effectively sample and manipulate the microbiome of eggshells. Although most vertebrates are oviparous, little is known about microorganisms on the surface of eggshells and their functions, particularly on eggs of non-avian reptiles. I developed a novel method to effectively sample (i.e., whole-egg sonication) and manipulate the eggshell microbiome of non-avian reptiles while minimizing contamination from external sources. Overall, my results provide useful guidelines for future manipulative studies that examine the source and function of the eggshell microbiome. For my third chapter, I experimentally manipulated the maternal gut microbiome using antibiotics and evaluate consequences on offspring phenotype in the brown anole lizard (Anolis sagrei). DNA was extracted from maternal gut tissue and cloacal samples and sequenced at the 16S rRNA gene. Eggs were incubated and embryo/hatchling phenotypes were recorded (e.g., survival, hatchling morphology). I found that treatment mothers had reduced gut microflora diversity and produced larger eggs/hatchlings than control mothers. Findings from this study provide new insight into the role of maternal gut microbiota and its potential functional significance on offspring. For my concluding chapter, I conducted a systematic review on vertical transmission of microbiota in non-human animals. I found that many studies examining vertical transmission of microbiomes fail to collect whole microbiome samples from both maternal and offspring sources, particularly for oviparous vertebrates. An ideal microbiome study incorporates host factors, microbe-microbe interactions, and environmental factors. Together, results from my dissertation suggest that the gut microbiome is highly influenced by environmental variation, including maternal effects, in ways that may affect offspring fitness. As evolutionary biologists continue to merge microbiome science and ecology, examining microbiomes in oviparous taxa may provide insight into how microbiota shape host phenotypes.