20-EEID US-UK Modelling reassortment at the cellular, clinical, and phylogenetic level in emerging Bunyaviruses

Genome segmentation has important implications for viral gene expression control and RNA assembly into nascent virions. It also creates the potential for reassortment: the exchange of intact gene segments between viruses that coinfect the same cell. Reassortment is quantitatively different from intramolecular recombination in that it allows many distinct genotypes to emerge from a single coinfected cell. Not only does segmentation enhance genetic diversification but it also plays a unique role in the evolutionary history of many segmented viruses due to the rare occasions when a reassortant is successful at the population scale. A striking example of the emergence of a novel virus through reassortment from the Bunyavirales is that of Ngari virus. For influenza (IAV), the best characterised segmented virus, reassortment has facilitated the formation of pandemic strains in 1957, 1968 and 2009. Out of seven epidemic-prone diseases prioritised by the WHO 2018 R&D Blueprint as public health emergency with an urgent need for accelerated research, three are Bunyaviruses. Thus, the overarching hypothesis of this project is that reassortment of segmented viruses plays a major role not only to drive their diversification and evolution, but to dramatically alter their ecology and transmission dynamics. Specifically, we aim to 1) develop mathematical models of the intracellular life cycle for a family of segmented viruses and together with in vitro experiments quantify for the first time their viral replication dynamics and reassortment frequencies, and 2) develop standardised protocols for sequencing, as well as novel phylogenetic methods to quantify the evolutionary and epidemiological implications of reassortment for Crimean-Congo hemorrhagic fever virus (CCHFV). A biobank with clinical and field samples from key CCHFV endemic regions in Turkey and Tajikistan will be set up in this project. Clinical and field data will be leveraged so that our methods and results have the potential to inform control strategies and predict outbreak risk. The quantitative methods developed in this project will contribute to the One Health approach advocated by Sorvillo et al. for CCHFV. Our goal is to develop novel mathematical and phylodynamic methods to quantify, at the cellular and the epidemic levels, reassortment frequency and fitness for segmented Bunyaviruses. This will enhance our understanding of CCHFV evolution and epidemiology in Turkey and Tajikistan. To achieve this goal, this US-UK collaborative project brings together expertise in phylogenetics, mathematical modelling, BSL-3 and BSL-4 capabilities, as well as field and clinical expertise in Turkey and Tajikistan with direct access to large collections of human, animal host and vector samples. We will quantify reassortment patterns, bringing together mathematical models with in vitro experiments of selected Bunyaviruses in arthropod and vertebrate cell lines. Statistical inference will allow us to estimate the basic reproduction number for each reassortant and cellular host. At the population level, we will design DNA sequencing and phylodynamic methods to dissect the evolutionary and transmission history of a segmented virus. Thus, our phylogenetic analysis of the field data to be collected in Turkey and Tajikistan will critically evaluate the convolution of point mutations, recombination and reassortment of CCHFV. Phylodynamic methods, adapted and developed in this project, together with field data, will allow us to relate these evolutionary processes to the host switching, temporal and geographical patterns of CCHFV outbreaks. With clinical information and data from Turkey and Tajikistan we will parameterise a mathematical model of CCHFV nosocomial transmission to inform public health decision making. While we focus on the Bunyaviridae family, especially CCHFV, our quantitative methods will be made adjustable to other segmented viruses of great public health relevance.

This project aims to study the transmission, evolutionary and epidemiological dynamics of Bunyaviruses in a range of vectors and hosts. The proposal will bridge experimental, clinical and field data to examine and quantify how reassortment and other genetic diversification mechanisms work at the cellular and population levels, where both selection (e.g., increased infectivity) and epidemiological processes (e.g., adaptation to a new host) affect the relative distribution of reassorted viruses. The overarching hypothesis of this project is that reassortment of segmented viruses plays a major role not only to drive their evolution, but to dramatically alter their ecology and transmission dynamics. Specifically, this proposal involves i) laboratory experiments (BSL-3 and BSL-4) to quantify the dynamics of Bunyavirus in vitro infection and reassortment frequencies in different host cells and ii) CCHFV clinical and field data collection in Turkey and Tajikistan in humans, small vertebrates, livestock and ticks. We will develop novel mathematical models and phylogenetic methods for segmented viruses to analyse rich field data from multiple hosts and vectors, clinical data from hospitals, and in vitro data from BSL-3 and BSL-4 experiments. Together, this project will provide the most comprehensive data and modelling of a real segmented virus epidemic to date, significantly improving our understanding of Bunyavirus transmission, ecology and evolution. We have divided the project into two specific aims:

Aim 1: Develop and parameterise, with in vitro experimental data, mathematical models of the intracellular Bunyavirus life cycle and quantify in vitro reassortment frequencies in arthropod and vertebrate cells.

Aim 2: Deconvolute the joint processes of reassortment, recombination and point mutation in the phylogeny of CCHFV with data from Turkey and Tajikistan and quantify the transmission and evolutionary implications of reassortment.