Hyperthermia reduces antimicrobial tolerance of Mycobacterium marinum in biofilms and ex vivo zebrafish granulomas

Abstract

The treatment of tuberculosis (TB) is slow and inefficient. One suggested reason for the treatment difficulties are drug-tolerant biofilms that have been shown to develop in TB patients and animal models. In this pioneering pilot study, we explored the potential of hyperthermia in sensitizing biofilm-associated tolerant mycobacteria to antimicrobial agents. As the starting point, we used our newly-developed in vitro biofilm minimum duration for killing (MDK) tolerance assay for screening hyperthermal treatments of bioluminescent mycobacteria. Using standard plating methods, we verify that the combination of hyperthermia at 47{degrees}C for 30 minutes and an antimicrobial drug rifampicin at 400 g/ml has positive effects against cultured Mycobacterium marinum biofilms as well as M. marinum from granulomas isolated from adult zebrafish with chronic-stage infection. These intriguing findings suggest that hyperthermia could be used to enhance conventional TB treatments. Author SummaryTuberculosis (TB) and similar infections are hard to cure because the bacteria can "hide" from antibiotics. They do this by entering a state where they tolerate drugs without becoming resistant, often forming biofilms--sticky clusters that protect them from treatment. This makes therapy long and relapse common. We asked whether heat could help break this tolerance. Using a new lab test, we combined heat at different durations and temperatures with antibiotics against Mycobacterium marinum, a close relative of the TB bacterium. Heating made the antibiotic rifampicin much more effective, both in biofilms and in infected tissue from zebrafish, which mimic TB-like lesions. We saw similar benefits in TB bacteria when heat was paired with streptomycin. These results suggest that controlled heating could work alongside standard drugs to kill stubborn bacteria faster. Because heat damages cells in a general way, not by targeting one molecule, it might also reduce the risk of antibiotic resistance