In the 1970s, researchers began to observe drastic amphibian declines in Brazil's Atlantic Forest, a global hotspot of frog biodiversity. For decades, these disappearances remained a mystery. Now, new research from scientists at Penn State and the Smithsonian's National Zoo and Conservation Biology Institute's (NZCBI) Center for Conservation Genomics published this week in the journal Molecular Ecology points to amphibian chytridiomycosis, a devastating infectious disease, as the likely culprit.
Chytridiomycosis, commonly known as chytrid, is caused by the microscopic fungal pathogen Batrachochytrium dendrobatidis. Believed to have originated in Asia, the chytrid fungus likely swept the globe in the 1970s and 1980s, but scientists did not formally describe the pathogen until 1999. At that point, amphibian populations around the world had suffered catastrophic declines, with hundreds of species pushed to the brink of extinction or beyond. Because many of these declines occurred before the pathogen's discovery, however, establishing whether the chytrid fungus was responsible has remained a challenge.
"After almost 50 years, we still aren't confident what caused many historical amphibian die-offs, making these declines some of the most enduring cold cases in wildlife epidemiology," said Jack Boyette, an NZCBI graduate fellow and doctoral candidate in biology in the Penn State Eberly College of Science, who led the study. "In Brazil's Atlantic Forest, amphibian die-offs at the Boracéia Biological Station between 1979 and 1982 have been attributed by some scientists to climate change or extreme weather events. While environmental factors could have played a role, our findings most directly implicate the chytrid fungus, revealing that regional invasion of the pathogen coincided with these unexplained amphibian disappearances."
To crack the case, Boyette and his colleagues turned to the Smithsonian's National Museum of Natural History, which holds the collections of William Ronald Heyer, a former curator of amphibians and reptiles. Heyer collected specimens at Boracéia across five field campaigns: two in the 1970s, prior to the sudden amphibian declines, and three in the 1980s, during the aftermath of the die-offs. These collections offered a rare opportunity to trace the pathogen's arrival across a critical period of amphibian declines. NZCBI researchers collected samples from more than 370 museum specimens representing 17 species to determine when the chytrid fungus first appeared at Boracéia and how infection prevalence varied through time.
The results were stark - specimens collected prior to 1979 showed no evidence of chytrid infection, while more than half of the 1980s specimens tested positive. Surprisingly, the species that suffered the most dramatic declines had no detectable infections, while the species that never declined were highly infected. Although these infection patterns may seem counterintuitive, the researchers hypothesize that they reflect natural patterns of disease tolerance among the amphibian community.
"It's important to keep in mind that not all species suffered catastrophic losses at Boracéia," Boyette said. "The species that didn't decline may have naturally tolerated the fungus, allowing them to become infected without suffering mass mortality. By contrast, the species that declined most dramatically may have been so susceptible that infected frogs died before Heyer could collect them."
Together, the researchers said, these findings suggest the chytrid fungus spread rapidly through the amphibian community at Boracéia in the 1980s, shedding light on how quickly this emerging disease can sweep through a community of naive hosts. The use of museum collections is a game changer for researchers seeking to reconstruct historical disease impacts, according to Michael Campana, a genomicist at NZCBI and co-author of the study.
"Museum collectors could not have anticipated how important their collections would be to our modern conservation challenges, but without them, we would never be able to detect pathogens that went unnoticed for decades and reconstruct disease outbreaks that would otherwise remain a mystery," Campana said.
Reconstructing historical disease dynamics may also help researchers anticipate future disease outcomes. The team said this is especially relevant as the emergence of another fungal pathogen, Batrachochytrium salamandrivorans, threatens salamanders in Europe and could pose a serious risk to salamander populations in North America.
At Boracéia, however, the story today is one of resilience. The team returned to Boracéia between 2022 and 2024, alongside researchers from the University of São Paulo in Brazil, to sample modern amphibian populations. Of the 17 species represented in Heyer's collection, the researchers could only find 12 at the site. The fate of the other five species remains uncertain, although their continued absence raises concerns that they may have been lost locally, the research team said. Reassuringly, however, most amphibian species appear to have recovered since the early 1980s. While the chytrid fungus still infects frogs at Boracéia, infection patterns suggest the evolution of widespread pathogen tolerance among species that historically declined, likely promoting their persistence and recovery in the decades since the declines.
"Brazil is one of the world's great biodiversity hotspots, home to thousands of animals and plants found nowhere else on Earth, many of which are now facing serious declines," said Gui Becker, associate professor of biology at Penn State and senior author of the paper. "Amphibians are the most threatened group of vertebrates, yet the history of amphibian declines in Brazil has remained remarkably nebulous. For decades, studies produced mixed evidence about whether the chytrid fungus was actually involved in these declines. Our study provides the clearest evidence yet that chytrid was tightly linked to amphibian losses in the region where those declines were most severe."
While the researchers acknowledge the site-specificity of the study means the results cannot be extrapolated to the broader Atlantic Forest, they do solve the mystery of Boracéia's disappearing frogs and provide a blueprint for other researchers to conduct similar disease retrospectives in the future.
The work is far from over, the team said. Researchers are now using DNA collected from museum samples to investigate the genomic mechanisms that shaped historical amphibian responses to chytrid, while other ongoing projects aim to fill gaps in historical specimen records using ancient environmental DNA. Together, these efforts could reveal a more detailed picture of chytrid emergence, spread and impacts over the past century, deepening understanding of its historical toll on biodiversity and helping to anticipate future impacts in a world where fungal pathogens remain a major threat.
In addition to Boyette, Campana and Becker, the research team included postdoctoral researcher Shannon M. Buttimer, research technologist Karen A. Paniagua Torres and graduate student in anthropology William C. Vuyk at Penn State; Tamilie Carvalho at the University of Michigan; David Rodriguez and Wesley J. Neely at Texas State University; Daniel Medina at the Center for Tropical Island Biodiversity Studies in Panama; Renato A. Martins at the Universidade Estadual Paulista in Brazil; Robert J. Ossiboff and Zaira Gomez at the University of Florida; Ananda Brito de Assis at the Universidade Estadual de São Paulo in Brazil; Carlos A. Navas at the Universidade de São Paulo in Brazil; Carly R. Muletz-Wolz at NZCBI; and Corinne L. Richards-Zawacki at the University of Pittsburgh.
This research was funded by the U.S. National Science Foundation under award numbers DEB-2227340, DEB-2413542, BII-2120084, IOS-2131060 and DGE-1255832 and the Smithsonian Institution Fellowship Program. This content is solely the responsibility of the authors and does not necessarily represent the views of the funders.
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