Theory predicts that selection for pathogen virulence and horizontal transmission is highest at the onset of an epidemic but decreases thereafter, as the epidemic depletes the pool of susceptible hosts. We tested this prediction by tracking the competition between the latent bacteriophage λ and its virulent mutant λcI857 throughout experimental epidemics taking place in continuous cultures of Escherichia coli. As expected, the virulent λcI857 is strongly favored in the early stage of the epidemic, but loses competition with the latent virus as prevalence increases. We show that the observed transient selection for virulence and horizontal transmission can be fully explained within the framework of evolutionary epidemiology theory. This experimental validation of our predictions is a key step towards a predictive theory for the evolution of virulence in emerging infectious diseases. Why are some pathogens more virulent than others? Theory predicts that pathogens that ‘keep their host alive’ can sometimes outcompete virulent pathogens in times when transmission to new susceptible hosts is unlikely. Yet, this prospect of finding a new susceptible host changes itself throughout an epidemic. In the early stage of an epidemic susceptible hosts are abundant and virulent pathogens that invest more into horizontal transmission should win the competition. Later on, the spread of the infection reduces the pool of susceptible hosts and may reverse the selection on virulence. This may favor benign pathogens after the acute phase of the epidemic. We model this transient benefit for virulence and predict both the epidemiology and the evolution of pathogens during an epidemic. To put these predictions to the test we monitor the competition of the temperate bacterial virus λ and its virulent mutant λcI857 in experimental epidemics. Our experimental results agree remarkably well with all our theoretical predictions. This demonstrates the ability of evolutionary epidemiology to predict selection for virulence in an ongoing epidemic.

Mutational (genetic) robustness is phenotypic constancy in the face of mutational changes to the genome. Robustness is critical to the understanding of evolution because phenotypically expressed genetic variation is the fuel of natural selection. Nonetheless, the evidence for adaptive evolution of mutational robustness in biological populations is controversial. Robustness should be selectively favored when mutation rates are high, a common feature of RNA viruses. However, selection for robustness may be relaxed under virus co-infection because complementation between virus genotypes can buffer mutational effects. We therefore hypothesized that selection for genetic robustness in viruses will be weakened with increasing frequency of co-infection. To test this idea, we used populations of RNA phage ϕ6 that were experimentally evolved at low and high levels of co-infection and subjected lineages of these viruses to mutation accumulation through population bottlenecking. The data demonstrate that viruses evolved under high co-infection show relatively greater mean magnitude and variance in the fitness changes generated by addition of random mutations, confirming our hypothesis that they experience weakened selection for robustness. Our study further suggests that co-infection of host cells may be advantageous to RNA viruses only in the short term. In addition, we observed higher mutation frequencies in the more robust viruses, indicating that evolution of robustness might foster less-accurate genome replication in RNA viruses.

The adaptive hypothesis invoked to explain why parasites harm their hosts is known as the trade-off hypothesis, which states that increased parasite transmission comes at the cost of shorter infection duration. This correlation arises because both transmission and disease-induced mortality (i.e. virulence) are increasing functions of parasite within-host density. There is, however, a glaring lack of empirical data to support this hypothesis. Here, we review empirical investigations reporting to what extent within-host viral accumulation determines the transmission rate and the virulence of vector-borne plant viruses. Studies suggest that the correlation between within-plant viral accumulation and transmission rate of natural isolates is positive. Unfortunately, results on the correlation between viral accumulation and virulence are very scarce. We found only very few appropriate studies testing such a correlation, themselves limited by the fact that they use symptoms as a proxy for virulence and are based on very few viral genotypes. Overall, the available evidence does not allow us to confirm or refute the existence of a transmission–virulence trade-off for vector-borne plant viruses. We discuss the type of data that should be collected and how theoretical models can help us refine testable predictions of virulence evolution.

Viral recombination can dramatically impact evolution and epidemiology. In viruses, the recombination rate depends on the frequency of genetic exchange between different viral genomes within an infected host cell and on the frequency at which such co-infections occur. While the recombination rate has been recently evaluated in experimentally co-infected cell cultures for several viruses, direct quantification at the most biologically significant level, that of a host infection, is still lacking. This study fills this gap using the cauliflower mosaic virus as a model. We distributed four neutral markers along the viral genome, and co-inoculated host plants with marker-containing and wild-type viruses. The frequency of recombinant genomes was evaluated 21 d post-inoculation. On average, over 50% of viral genomes recovered after a single host infection were recombinants, clearly indicating that recombination is very frequent in this virus. Estimates of the recombination rate show that all regions of the genome are equally affected by this process. Assuming that ten viral replication cycles occurred during our experiment—based on data on the timing of coat protein detection—the per base and replication cycle recombination rate was on the order of 2 × 10⁻⁵ to 4 × 10⁻⁵. This first determination of a virus recombination rate during a single multi-cellular host infection indicates that recombination is very frequent in the everyday life of this virus.

Co-infection may be beneficial in large populations of viruses because it permits sexual exchange between viruses that is useful in combating the mutational load. This advantage of sex should be especially substantial when mutations interact through negative epistasis. In contrast, co-infection may be detrimental because it allows virus complementation, where inferior genotypes profit from superior virus products available within the cell. The RNA bacteriophage φ6 features a genome divided into three segments. Co-infection by multiple φ6 genotypes produces hybrids containing reassorted mixtures of the parental segments. We imposed a mutational load on φ6 populations by mixing the wild-type virus with three single mutants, each harboring a deleterious mutation on a different one of the three virus segments. We then contrasted the speed at which these epistatic mutations were removed from virus populations in the presence and absence of co-infection. If sex is a stronger force, we predicted that the load should be purged faster in the presence of co-infection. In contrast, if complementation is more important we hypothesized that mutations would be eliminated faster in the absence of co-infection. We found that the load was purged faster in the absence of co-infection, which suggests that the disadvantages of complementation can outweigh the benefits of sex, even in the presence of negative epistasis. We discuss our results in light of virus disease management and the evolutionary advantage of haploidy in biological populations.

The transmission–virulence trade-off hypothesis is one of the few adaptive explanations of virulence evolution, and assumes that there is an overall positive correlation between parasite transmission and virulence. The shape of the transmission–virulence relationship predicts whether virulence should evolve toward either a maximum or to an intermediate optimum. A positive correlation between each of these traits and within-host growth is often suggested to underlie the relationship between virulence and transmission. There are few experimental tests of this hypothesis; this study reports on the first empirical test on a plant pathogen. We infected Brassica rapa plants with nine natural isolates of Cauliflower mosaic virus and then estimated three traits: transmission, virulence, and within-host viral accumulation. As predicted by the trade-off hypothesis, we observed a positive correlation between transmission and virulence, suggestive of the existence of an intermediate optimum. We discovered the unexpected existence of two groups of within-host accumulation, differing by at least an order of magnitude. When accumulation groups were not accounted for, within-host accumulation was correlated neither to virulence nor transmission, although our results suggest that within each group these correlations exist.