Manipulating multi-level selection in a fungal entomopathogen reveals social conflicts and a method for improving biocontrol traits.

Changes in parasite virulence are commonly expected to lead to trade-offs in other life history traits that can affect fitness. Understanding these trade-offs is particularly important if we want to manipulate the virulence of microbial biological control agents. Theoretically, selection across different spatial scales, i.e. between- and within-hosts, shapes these trade-offs. However, trade-offs are also dependent on parasite biology. Despite their applied importance the evolution of virulence in fungal parasites is poorly understood: virulence can be unstable in culture and commonly fails to increase in simple passage experiments. We hypothesized that manipulating selection intensity at different scales would reveal virulence trade-offs in a fungal pathogen of aphids, Akanthomyces muscarius. Starting with a genetically diverse stock we selected for speed of kill, parasite yield or infectivity by manipulating competition within and between hosts and between-populations of hosts over 7 rounds of infection. We characterized ancestral and evolved lineages by whole genome sequencing and by measuring virulence, growth rate, sporulation and fitness. While several lineages showed increases in virulence, we saw none of the trade-offs commonly found in obligately-killing parasites. Phenotypically similar lineages within treatments often shared multiple single-nucleotide variants, indicating strong convergent evolution. The most dramatic phenotypic changes were in timing of sporulation and spore production in vitro. We found that early sporulation led to reduced competitive fitness but could increase yield of spores on media, a trade-off characteristic of social conflict. Notably, the selection regime with strongest between-population competition and lowest genetic diversity produced the most consistent shift to early sporulation, as predicted by social evolution theory. Multi-level selection therefore revealed social interactions novel to fungi and showed that these biocontrol agents have the genomic flexibility to improve multiple traits-virulence and spore production-that are often in conflict in other parasites.