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A lethal combination Although many human activities have clear negative effects on the natural world, there are also unforeseen consequences. Bald eagle mass death events in the southeastern United States may be one such downstream effect of human activity. After considerable effort, Breinlinger et al. identified the cause of these events as an insidious combination of factors. Colonization of waterways by an invasive, introduced plant provided a substrate for the growth of a previously unidentified cyanobacterium. Exposure of this cyanobacterium to bromide, typically anthropogenic in origin, resulted in the production of a neurotoxin that both causes neuropathy in animals that prey on the plants and also bioaccumulates to kill predators such as bald eagles. Science, this issue p. eaax9050 A deadly toxin that kills bald eagles originated from an environmental interaction between an invasive plant, a cyanobacterium, and bromide. INTRODUCTION Vacuolar myelinopathy (VM) is a neurological disease characterized by widespread vacuolization in the white matter of the brain. First diagnosed in 1994 in bald eagles, it has since spread throughout the southeastern United States. In addition to avian species such as waterfowl and birds of prey, VM has also been found to affect amphibians, reptiles, and fish. Despite intense research efforts, the cause of this mysterious disease has been elusive. Neither contagious agents nor xenobiotics were detected in deceased animals, but field and laboratory studies demonstrated that VM can be transferred through the food chain from herbivorous fish and wildlife to birds of prey. RATIONALE Occurrence of VM has been linked to a cyanobacterium (Aetokthonos hydrillicola) growing on an invasive plant (Hydrilla verticillata) in man-made water bodies. Cyanobacteria are known to produce potent toxins, so we hypothesized that a neurotoxin produced by the epiphytic cyanobacterium causes VM. RESULTS Field studies in the southeastern United States confirmed that H. verticillata was colonized with A. hydrillicola in more than half of the watersheds. Wildlife VM deaths occurred only in reservoirs with dense H. verticillata and A. hydrillicola populations. Laboratory bioassays confirmed the neurotoxicity of crude extracts of A. hydrillicola–H. verticillata biomass collected during VM outbreaks, but neurotoxicity was not detected in samples from VM-free sites. Laboratory cultures of the cyanobacterium, however, did not elicit VM. A. hydrillicola growing on H. verticillata collected at VM-positive reservoirs was then analyzed by mass spectrometry imaging, which revealed that cyanobacterial colonies were colocalized with a brominated metabolite. Supplementation of an A. hydrillicola laboratory culture with potassium bromide resulted in pronounced biosynthesis of this metabolite. H. verticillata hyperaccumulates bromide from the environment, potentially supplying the cyanobacterium with this biosynthesis precursor. Isolation and structure elucidation of the metabolite revealed a structurally unusual pentabrominated biindole alkaloid, which we called aetokthonotoxin (AETX). Genome sequencing of A. hydrillicola allowed the identification of the AETX biosynthetic gene cluster. Biochemical characterization of a halogenase detected in the cluster demonstrated that it brominates tryptophan with the expected substitution pattern. AETX is highly toxic to the nematode Caenorhabditis elegans [median lethal concentration (LC50) 40 nM] and zebrafish (Danio rerio; LC50 275 nM). Leghorn chickens (Gallus gallus) gavaged with AETX developed brain lesions characteristic of VM, whereas no lesions were observed in control chickens. VM diagnosis in treated chickens was verified using transmission electron microscopy of brain tissue. CONCLUSION We confirmed that AETX is the causative agent of VM. AETX biosynthesis relies on the availability of bromide. Seasonal environmental conditions promoting toxin production of A. hydrillicola are watershed specific. The consequences of elevated bromide from geologic and anthropogenic sources (e.g., water treatment and power plants) on VM should be further investigated. Notably, integrated chemical plant management plans to control H. verticillata should avoid the use of bromide-containing chemicals (e.g., diquat dibromide). AETX is lipophilic with the potential for bioaccumulation during transfer through food webs, so mammals may also be at risk. Increased monitoring and public awareness should be implemented for A. hydrillicola and AETX to protect both wildlife and human health. From the cyanobacterium to the bald eagle—toxin transmission through the food chain. A. hydrillicola, growing in colonies on aquatic vegetation, produces the neurotoxin AETX. Waterbirds, tadpoles, aquatic turtles, snails, and fish consume this contaminated vegetation and develop VM. Predators develop VM when they consume animals that have been grazing on A. hydrillicola–covered plants. IMAGE CREDITS: GREENFROG TADPOLE, B. GRATWICKE; AMERICAN COOT, G. S. SEGLER; GRASS CARP, R. HAGERTY; SNAIL KITE, SIRKFISH; PAINTED TURTLE, U.S. FISH AND WILDLIFE SERVICE; BALD EAGLE, W. H. MAJOROS. IMAGES ARE ALL UNDER THE CREATIVE COMMONS ATTRIBUTION GENERIC LICENSE Vacuolar myelinopathy is a fatal neurological disease that was initially discovered during a mysterious mass mortality of bald eagles in Arkansas in the United States. The cause of this wildlife disease has eluded scientists for decades while its occurrence has continued to spread throughout freshwater reservoirs in the southeastern United States. Recent studies have demonstrated that vacuolar myelinopathy is induced by consumption of the epiphytic cyanobacterial species Aetokthonos hydrillicola growing on aquatic vegetation, primarily the invasive Hydrilla verticillata. Here, we describe the identification, biosynthetic gene cluster, and biological activity of aetokthonotoxin, a pentabrominated biindole alkaloid that is produced by the cyanobacterium A. hydrillicola. We identify this cyanobacterial neurotoxin as the causal agent of vacuolar myelinopathy and discuss environmental factors—especially bromide availability—that promote toxin production.