The dynamics of mucosal cytomegalovirus replication and barriers to dissemination during acquisition of infection

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Cytomegalovirus (CMV) infects more than 90% of humans, and is primarily acquired at mucosal surfaces. Upon exposure, CMV initially replicates in mucosal epithelium and then spreads to establish life-long infection in distant anatomic sites. However, we have found that CMV acquisition is highly inefficient in humans, and infection is frequently transient without spread beyond the mucosa. We have determined that these natural CMV transmission dynamics are best explained by weak cell-to-cell spread from only 1 or very few infected mucosal cells at the start of infection. Thus, although life-long CMV infection eventually becomes established in most people, this typically occurs only after large numbers of exposures, most of which many are rapidly extinguished.This project aims to better define the basis of this severe transmission bottleneck, to understand why CMV infection does not occur. We will elucidate the earliest events of CMV acquisition, using in vitro, murine, and mathematical models to define the viral and host conditions that determine whether mucosal CMV replication persists and spreads, or terminates following exposure to a range of viral inoculum sizes. We hypothesize that CMV dynamics in mucosal epithelium are influenced by stochastic as well as host immune factors, and that infection terminates when viral replication does not reach a threshold required for dissemination. We also predict that innate immune barriers can be overcome by larger viral inocula. We will address these questions following specific aims: Aim 1: Define the dynamics of CMV mucosal replication and spread in cultured cells and mice. We will use wild-type and recombinant CMV strains to measure viral expansion in cultured epithelial cells and in the mucosa of mice by live bioimaging, PCR, and histopathology. We will also determine the frequency of self-limited mucosal infections vs. disseminated infections over a range of inoculum sizes.Aim 2: Identify the role of host innate immunity in determining successful CMV infection. We will identify the immune responses that constrain CMV spread in mucosal cells, by inducing or blocking antiviral factors as well as transcriptomics using in vitro and murine systems, including knock-out mice.Aim 3: Construct mathematical models to elucidate determinants of successful CMV infection. Detailed in vitro and murine data will be used to supplement our extensive human data, and allow refinement and expansion of our mathematical models of CMV acquisition. Model predictions will represent hypotheses to be tested in further rounds of experiments.This work will be conducted by pre- and postdoctoral trainees, in a highly collaborative interdisciplinary learning environment at UBC. By integrating state-of-the-art virology, immunology, and mathematical modeling data, we will gain fundamental insights into the determinants of whether a natural CMV infection succeeds or fails.