In vivo exposure of insect AMP resistant Staphylococcus aureus to an insect immune system

Antimicrobial peptides (AMPs) are important immune effectors in insects. Bacteria have a limited number of ways to resist AMPs, and AMP-resistance is often costly. Recently, it has become clear that AMP activities in vitro and in vivo differ. Although some studies have followed the in vivo survival of AMP resistant pathogens, studying a pathogen resistant to the AMPs of that particular host has never been reported. Here, we infected the mealworm beetle Tenebrio molitor with Staphylococcus aureus strains that were evolved in vitro in the presence of one or two antimicrobial peptides from T. molitor. We found that the Tenebrio immune system could clear mutant Tenecin resistant strains at least as efficiently as sensitive controls. The bacterial load of Tenecin resistant S. aureus segregated by mutation. Strains with mutations in both the pmt and rpo operons showed the highest in vivo survival and therefore showed the lowest fitness cost amongst the evolved resistance mutations. In contrast, Tenecin resistant strains with mutations in the nsa and rpo operons showed much lower survival within the hosts. Our study shows that Tenecin resistant strains are phagocytosed at a lower rate. The nsa/rpo mutants were phagocytosed at a higher rate than other Tenecin resistant S. aureus strains. The differences in resistance against AMPs and phagocytosis did not translate into changes in virulence. AMP resistance, while a prerequisite for an infection in vertebrates, does not provide a survival advantage to S. aureus in a host environment that is dominated by AMPs.