Noninvasive molecular assays allow the investigation of elusive, rare and/or endangered organisms, and are useful for estimates of population parameters. Sex determination of noninvasive samples provides important information of sex ratio, social structure and dispersal patterns. Still, such assays remain unavailable for most carnivore species, precluding assessments of their population biology. Here we report a standardized protocol for molecular sexing of Neotropical otter noninvasive samples collected in the field. A duplex PCR assay was employed using a multiple-tube approach, resulting in reliable gender determination for 95% of the samples and presenting a low rate (<4%) of misassignment. This assay should foster the development of molecular ecological studies of this little-known species, and likely also contribute to similar endeavors on related otters.

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