IMPACT OF FORAGING BEHAVIOR, NUTRITION, AND GUT MICROBIOME ON CORONAVIRUS PREVALENCE IN BATS: LESSONS IN MINIMIZING THE IMPACTS OF HABITAT LOSS ON THE VIRAL ECOLOGY OF BATS

Abstract

Deforestation is a leading root cause of viral spillover from bats. This is hypothesized to be due to two primary reasons: (1) increased overlap between wildlife and humans/livestock creates new opportunities for contact and cross-species transmission to occur, and (2) loss of biodiversity results in dietary changes for wildlife that have negative impacts on their immune systems, causing greater infection intensity or viral shedding into the environment. The second of these hypotheses has received comparatively little attention. To counteract the potential impact of biodiversity loss on the bat immune system, the impact of resource scarcity on bat foraging behavior requires further study. Furthermore, the gut microbiome, which performs immunomodulation services and in bats is largely environmentally driven, is understudied in bats from an infectious disease perspective. Seasons, particularly in resource-scarce regions, can provide insights into wildlife behavior under both optimal and non-optimal conditions. These insights could promote improved understanding of how wildlife behave following land-use changes. The first research chapter of this dissertation compares the foraging habits of Angolan rousette bats (Myonycteris angolensis) in the dry and wet seasons of the Mount Elgon region of eastern Uganda. Critical seed dispersers, M. angolensis are frugivorous bats that specialize on fruits growing in montane forests and are often found preferentially on forest edges, frequently exposing them to ongoing deforestation or land-conversion efforts. To interrogate seasonal patterns in foraging behavior, GPS tracking and diet metabarcoding were leveraged to understand 1) the spatial and movement pattern differences and 2) the specific dietary changes across the two seasons. The gut microbiome (as interpreted through fecal samples) was examined to understand how these changes in diet and environment impact the enteric system of these bats, which carries implications for metabolism and immune functioning. This study showed that the abundant fruit in the wet season was not associated with higher dietary diversity, as originally hypothesized. Rather, fruit scarcity in the dry season was associated with more experimental foraging behavior and diverse diet selection, whereas the abundance of fruit in the wet season was associated with movement and dietary consistency. Furthermore, high dietary diversity was not linked to high microbial diversity of the gut. Conversely, high gut microbiome diversity was associated with a more stable diet characteristic of the wet season, though this is not necessarily a causal relationship and rather could be due to non-dietary seasonal differences. While this study focused on seasonality, which is predictable, it could have insights for how frugivorous bats would respond to unpredictable events, such as deforestation or climate disasters, that result in resource scarcity. Where the fruit bats from Chapter 2 are critical seed dispersers, insectivorous bats are important for agricultural pest control while also posing potential concerns for spillover of coronaviruses. As land conversion to agriculture creates dietary opportunity to feed on agricultural pests, there is likely to be change to their nutritional intake and their gut microbiome. To study dietary composition of insectivorous bats utilizing both forested and agricultural habitats, fecal samples were collected from insectivorous bats from four cave systems from the Mount Elgon region of Eastern Uganda across both dry and wet seasons from 2022 to 2023. Additionally, rectal and oral swabs were collected and tested for coronaviruses. In the second research chapter of this dissertation, these fecal samples were used to determine the diet of these bats using metabarcoding techniques. This study showed the immense agricultural services provided by the insectivorous bats in this region through the enriched composition that agricultural pests comprised in the bats’ diet. It also showed no correlation between viral RNA detection and diet or the gut microbiome. However, similar to the study in frugivorous bats (Chapter 2), an inverse correlation was seen between the diversity of the diet and the gut microbiome, despite having a positive correlation between their overall composition. It is possible that these field studies found no significant correlation between diet or microbiome with viral RNA detections because of confounding variables inherent in field-based studies, including environmental and the inability to determine time point in viral infections. Thus, the third research chapter (Chapter 4) examined potential links between diet, gut microbiome, and viral exposure in a controlled laboratory setting. For frugivorous bats, loss of native forests results in loss of native fruits, which are often more rich in protein and other nutrients than cultivated or invasive fruits. Reliance on cultivated fruits, often low in protein, can lead to nutritional deficiencies in fruit bats that could change immune regulation, potentially via the gut microbiome. Previous studies on fruit bats have found specific bacterial groups in the gut to be predictors of the strength of the immune response. Thus, understanding the relationship between diet, gut microbiome, and outcome of viral exposures is critical. To characterize these interactions in a controlled setting, we evaluated the impact of dietary protein restriction in Jamaican fruit bats (Artibeus jamaicensis) intranasally inoculated with Middle East respiratory syndrome coronavirus (MERS-CoV; Coronaviridae: Betacoronavirus). Gut microbiome diversity increased in MERS-CoV inoculated bats with nutrient sufficient diets, but did not increase in MERS-CoV inoculated bats with nutrient deficient diets. Bats not inoculated with MERS-CoV maintained stable gut microbiome diversity with nutrient sufficient diets and experienced a slight decrease with nutrient deficient diets. Data obtained through controlled experimental studies are critical in understanding how diet impacts the gut microbiome and mechanisms of immunomodulation. Our study examines this relationship through the lens of dietary health, where organisms are directly impacted by habitat loss. The results of this study have implications for public health surveillance, translational immunology, and bat conservation strategies.