Natural variation in viral susceptibility of Caenorhabditis elegans

The nematode Caenorhabditis elegans belongs to the world’s most powerful genetic model organisms. Studying the genome of this nematode is facilitated by the androdiecious (male-hermaphrodite) mode of reproduction. A single hermaphrodite can start a population that will contain of hundreds of genetically identical individuals after only a couple of days. Males are not necessary for reproduction but are used to recombine genomes, for instance to introduce a mutation into a population. Although the nematode has provided a wealth of genetic knowledge, a large part of the C. elegans genome does not have a known function yet. Many of the genes without an assigned role will likely have a function in natural populations, for example by providing protection against the natural pathogens that are ubiquitous in nature. Viruses strongly shape the genome of their host and in this thesis, the interaction between the positive-strand RNA virus, the Orsay virus (OrV), and C. elegans was studied.

Chapter 2 reviews quantitative genetic studies investigating antiviral defense and discusses the practical tools to perform these studies in model organisms. Natural genetic variation in the genome of the host leads to different viral susceptibilities for individuals of the same species. Understanding the consequences of host genetic variation is expected to lead to better treatments and personalized medicine for human patients. But studying viral infections in humans comes with ethical and practical challenges. Therefore, quantitative genetic studies in model organisms can help to better understand the mechanisms by which host genetic variation defines viral susceptibility. To study host-virus interactions in three genetic model organisms, mouse (Mus musculus), fruit fly (Drosophila melanogaster) and nematode (C. elegans), different tools have been developed over the years. These include two- or multiparent Recombinant Inbred Lines (RILs) and Introgression Lines (ILs) that are genetic mosaics of their parental strains. Additionally, Genome Wide Association Studies (GWAS) link genetic variation in populations of genetically distinct individuals to viral susceptibility. The use of these tools has led to identification of genetic variants that contribute to viral susceptibility in evolutionary conserved pathways and has improved the understanding of human-infecting viruses such as West Nile, Influenza and Ebola.

Chapter 3 investigates natural genetic variation in antiviral defense in C. elegans populations. Because viruses represent such a strong selective pressure, their natural presence also leads to genetic changes in the genome of their host. Traces of pathogenic selection were identified in C. elegans by analyzing the genetic composition of antiviral genes of the Intracellular Pathogen Response (IPR) in strains that were collected worldwide. This led to the discovery that natural strains carry a limited set of pals-gene variants, a class of genes fundamental within the IPR, that are maintained in C. elegans populations by balancing selection. Only three haplotypes were found worldwide for the IPR regulators pals-22 and pals-25 and two strains with distinct pals-22/pals-25 haplotypes had a different IPR activity. The pals-22/pals-25 haplotype of the standard reference strain, Bristol N2, was most common in currently samples strains. The N2 strain had low basal IPR expression that strongly increased upon OrV infection. Contrary, the Hawaiian strain CB4856, had a constantly active IPR that did not change after OrV exposure. The strain CB4856 had lower viral susceptibility than the N2 strain after short term exposure, which might relate to IPR activity.

Chapter 4 studies the genetic loci that determine viral susceptibility in N2 and CB4856. To uncover genetic variants that underlie the different viral susceptibility of N2 and CB4856, a quantitative trait locus (QTL) mapping approach was used. Thereto, a panel of N2xCB4856 RILs was infected and statistical associations between viral susceptibility and genetic background linked to chromosome IV. Using ILs from both genetic backgrounds, a small region containing 34 polymorphic genes was found to affect the viral susceptibility of N2 and CB4856. One of the genes located in this region is the IPR gene cul-6. This gene contains a single nucleotide polymorphism between N2 and CB4856 at a conserved site near the binding domain with another cul-6 complex member. After infection of CRISPR-Cas9 allele swap strains it became clear that cul-6 contributes to the viral susceptibility. Nevertheless, having a CB4856 cul-6 allele only did not confer resistance. Together with the finding that multiple genetic loci contribute to differences between N2 and CB4856 this shows that viral susceptibility is a complex trait with a polygenic basis.

Chapter 5 investigated viral infection in mixed-sex populations of C. elegans. Sex is another factor that is genetically determined, and sexual differences can underlie different viral susceptibilities. Both sexes (males and hermaphrodites) of three genetically distinct strains were exposed to the OrV. Males of the reference strain N2 were more resistant to the OrV than hermaphrodites. This could result from higher IPR activity in males under standard conditions, possibly protecting them from infection. Viral presence can also change population and mating dynamics. Indeed, male frequencies increased in the isogenic populations of the genotypes CB4856 and JU1580. Moreover, males rather mated with healthy than infected hermaphrodites. Together, this shows that viral infection can result in flexible outcrossing in C. elegans populations.

The research presented in this thesis displays how viral presence can have shaped the genome of C. elegans and how genetic variation determines viral susceptibility to date. Although these observations have been made in the laboratory, they were placed in the context of field observations to provide a better ecological context for this model organism. The findings made here suggest an evolutionary advantage for individuals with an active IPR haplotype under pathogenic conditions. Moreover, these results indicate an ecological advantage of having males, even in the frequently isogenic populations that are found in nature. Together, this thesis invites for investigation of host-virus interactions in a more natural set-up to fully incorporate lab and field studies.