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Amphibians,byvirtueoftheirphylogeneticposition,provideinvaluableinsightsonnervoussystemevolution, development, and remodeling. The genetic toolkit for amphibians, however, remains limited. Recombinant adeno-associated viral vectors (AAVs) are a powerful alternative to transgenesis for labeling and manipulating neurons. Although successful in mammals,AAVshaveneverbeenshowntotransduceamphibiancells efficiently. We screened AAVs in three amphibian species—the frogs Xenopus laevis and Pelophylax bedriagae andthesalamanderPleurodeles waltl—and identified at least two AAV serotypes per speciesthat transduceneurons.Indevelopingamphibians,AAVslabeledgroupsofneuronsgeneratedatthesametimeduring development. In the mature brain, AAVrg retrogradely traced long-range projections. Our study introduces AAVs as a tool for amphibian research, establishes a generalizable workflow for AAV screening in new species, and expands opportunities for cross-species comparisons of nervous system development, function, and evolution.