Zika virus (ZIKV) and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) are the etiologic agents responsible for the two major pandemics of the 21st century (so far).

By virtue of its capacity for mosquito-borne, sexual, and vertical transmission, as well as its association with congenital Zika syndrome (CZS), ZIKV remains a significant global public health risk. Mounting evidence identifies the male reproductive tract as a significant ZIKV reservoir; however, data regarding the duration of ZIKV persistence, potential for sexual transmission, and male genitourinary sequelae remain sparse. Furthermore, while microcephaly is a well-documented, extreme manifestation of CZS, there are cases of normocephalic newborns with confirmed prenatal ZIKV exposure but no observable congenital defects exhibiting neurologic deficits and behavioral abnormalities over time. These and other long-term developmental sequelae to prenatal ZIKV infection are not well described.

I addressed these gaps in knowledge using archived tissues from 1) 51 ZIKV-inoculated male macaques from past collaborative research projects; and 2) two juvenile rhesus macaque infants exposed prenatally to ZIKV. In the male macaque cohort, I identified persistent ZIKV RNA and infectious virus in testis, epididymis, seminal vesicle, and prostate gland of sexually mature male macaques, with detection as late as 60 days post-inoculation. ZIKV RNA localized primarily to testicular stem cells/sperm precursors and epithelial cells, including Sertoli cells, epididymal duct epithelium, and glandular epithelia of the seminal vesicle and prostate gland. ZIKV infection was associated with microscopic evidence of inflammation in the epididymis and prostate gland of sexually mature males, which could have significant effects on male fertility. In the two juvenile macaques, I identified CNS microcalcifications and macrostructural developmental abnormalities within the CNS visual pathway, specifically disorganization, blending of layers, laminar discontinuities, and regions of low cell density within the lateral geniculate nucleus.

COVID-19 disease, caused by SARS-CoV-2, varies from asymptomatic to severe respiratory disease, progressing to acute respiratory distress syndrome (ARDS), multiorgan dysfunction, and death in a subset of patients. Clinical evidence of coagulopathy and microscopic indicators of pulmonary vascular damage are increasingly reported; however, published autopsy data in human patients remain relatively scarce and SARS-CoV-2 induced vascular lesions have not been fully characterized. I used a previously described Syrian golden hamster model of COVID-19 disease to demonstrate that regions of active SARS-CoV-2 induced pulmonary inflammation exhibit ultrastructural evidence of endothelial injury with platelet marginalization and marked perivascular and subendothelial mononuclear inflammation composed primarily of macrophages. SARS-CoV-2 antigen/RNA was not associated with affected blood vessels. Taken together, these findings suggest that the prominent microscopic vascular lesions in SARS-CoV-2 inoculated hamsters (and by extrapolation, humans) are primarily due to indirect endothelial damage, likely secondary to immune dysfunction.

These studies emphasize the importance of animal models such as nonhuman primates and hamsters to study pathogenesis of emerging viral diseases.

Arthropod borne viral (Arboviral) disease accounts for 17% of the total infectious disease burden, afflicting over 100 million people annually. Global expansion of mosquito-borne arboviruses demands integrated approaches to vector control and public health surveillance. However, disparate outcomes in laboratory vector competence studies complicates risk assessment of mosquito species as vectors. While the contribution of mosquito and viral genetics has enjoyed much attention, the effects of mosquito microbiota on arboviral transmission potential are poorly understood. For Aedes aegypti, which is an effective vector for many arboviruses including Zika virus (ZIKV), the microbiota is primarily environmentally derived and dominantly resides in the gut. Chapter 1 reviews the current knowledge of Ae. aegypti vector competence for Zika virus as well as known effects that mosquito microbiota have on vector competence. Chapter 2 assesses the impact of microbes acquired from the larval habitat on Ae. aegypti development and ZIKV transmission. Adult female mosquitoes that emerged from microbially rich larval water derived from cemetery headstones were found to harbor more diverse microbiota and consistently lower ZIKV infection and transmission rates than their laboratory counterparts reared in laboratory tap water. However, microbial community compositions varied between experiments despite a consistent phenotype. Together, the results suggest that wild Ae. aegypti are likely less competent vectors than conventionally determined in the lab where larvae are typically reared in tap water, and that this effect is mediated by mosquito interactions with their microbiota. Chapter 3 investigates the reversibility of larval microbe-mediated refraction of ZIKV after developmental maturity. A higher dissemination rate was observed in Ae. aegypti depleted of gut microbes during pupation, and this was linked to reduced blood digestion efficiency. Results of this work suggest an immuno-metabolomic mechanism by which gut microbes confer resistance to ZIKV dissemination, by way of nonstructural midgut modifications. Overall, work presented in this dissertation emphasizes the importance of environmental microbes as a source of variation in infection susceptibility that demands consideration when conducting vector competence studies. It also highlights the complex interactions between mosquito, virus, and all the symbionts in between that play shape transmission out in nature.