Antibiotics are primarily found in the environment due to human activity, which has been reported to influence the structure of biotic communities and the ecological functions of soil and water ecosystems. Nonetheless, their effects in other terrestrial ecosystems have not been well studied. As a result of oxidative stress in organisms exposed to high levels of antibiotics, genotoxicity can lead to DNA damage and, potentially, cell death. In addition, in symbiotic organisms, removal of the associated microbiome by antibiotic treatment has been observed to have a big impact on the host, e.g., corals. The lung lichen Lobaria pulmonaria has more than 800 associated bacterial species, a microbiome which has been hypothesized to increase the lichen's fitness. We artificially exposed samples of L. pulmonaria to antibiotics and a stepwise temperature increase to determine the relative effects of antibiotic treatments vs. temperature on the mycobiont and photobiont gene expression and the viability and on the community structure of the lichen-associated bacteria. We found that the mycobiont and photobiont highly reacted to different antibiotics, independently of temperature exposure. We did not find major differences in bacterial community composition or alpha diversity between antibiotic treatments and controls. For these reasons, the upregulation of stress-related genes in antibiotic-treated samples could be caused by genotoxicity in L. pulmonaria and its photobiont caused by exposure to antibiotics, and the observed stress responses are reactions of the symbiotic partners to reduce damage to their cells. Our study is of great interest for the community of researchers studying symbiotic organisms as it represents one of the first steps to understanding gene expression in an endangered lichen in response to exposure to toxic environments, along with dynamics in its associated bacterial communities.