Described mechanisms of antimicrobial resistance (AMR) involve resistance to the direct effects of the antibiotic, and can be monitored by in vitro susceptibility testing and genetic methods. However, obligate human pathogens such as Group A Streptococcus (GAS) only experience antibiotic exposure in the context of an infection. The most widely-used treatment for GAS skin and soft tissue infections is a synergistic combination of trimethoprim and sulfamethoxazole (co-trimoxazole). Co-trimoxazole targets sequential steps in the bacterial folate biosynthesis pathway, with all known resistance mechanisms requiring individual resistance to each antibiotic. We have discovered a novel mechanism of co-trimoxazole resistance that involves an AMR protein (ThfT) that modifies the substrate recognition of a GAS ABC transporter, such that ThfT-positive GAS strains can acquire the end products of the folate synthesis pathway directly from the host. As these metabolites are abundant in vivo but lacking in susceptibility testing media, ThfT-mediated resistance is "host dependent" and not detectable by existing phenotypic or genetic methods. Evidence of transfer of thfT between GAS and related species suggests that resistance could disseminate widely without detection. Our study highlights the importance of understanding antibiotic activity in the context of the infections that they are used to treat, and should serve as a paradigm for investigating additional mechanisms of host-dependent AMR in medically-important bacterial pathogens.