UC San Diego

The bacteria that cause tuberculosis (TB), Mycobacterium tuberculosis, is estimated to infect 10 million people annually. Though often associated with lung disease, the deadliest form of tuberculosis occurs when M. tuberculosis invades the brain, causing catastrophic inflammation and neuronal damage. Currently, much of what is known about TB meningitis relies on in vitro models that do not accurately represent the interactions between host vasculature, the brain, and the bacteria. To address this, our lab uses a novel in vivo model of mycobacterial brain infection. In this model, zebrafish larvae are injected in the tail vein with Mycobacterium marinum, a natural fish pathogen and close genetic relative of M. tuberculosis. Zebrafish larvae, which are transparent, allow for observation of natural bacterial dissemination into the brain from the tail and whole-brain imaging in living animals. Using this system, we observed that M. marinum disseminate into the brain through the blood and replicate features of the human disease, such as granuloma formation. Through live imaging, we found that M. marinum invade brain tissue through a previously undescribed mechanism. Our data suggest that extracellular mycobacteria initiate crossing by first attaching within the lumen of brain blood vessels. Following attachment, we observe a global disruption of blood flow velocity in the brain, as well as full occlusion of vessels containing attached colonies. What’s more, infected zebrafish display other hallmarks of neurovascular disease, such as oxidative stress. Interestingly, neurovascular disease is a major contributor to death in human cases of TB meningitis yet remains understudied. We observe that following attachment and vessel occlusion, zebrafish induce vessel enlargement and the reorganization of f-actin and tight-junction proteins. These events precede the formation of permeable paracellular holes in brain blood vessels that allow for mycobacterial entry into the brain parenchyma. We have further confirmed these findings by replicating these results in zebrafish infected with an M. tuberculosis auxotroph. Together, these studies will grant a better understanding of the neuropathogenesis of mycobacteria and establish an animal model of TB meningitis neurovascular disease to lay a foundation for developing more effective treatments.