Species-specific vocalizations can act as a reproductive isolating mechanism between closely related populations. We analyzed vocal differences between 2 hybridizing species of sex-role reversed polyandrous shorebirds, the Northern Jacana (Jacana spinosa) and Wattled Jacana (J. jacana). We found that Northern Jacana calls have higher fundamental frequency and peak frequency than Wattled Jacana calls. We also compared calls between females and males, as both jacana species are sex-role reversed and females compete for male mates. Males produce calls with a higher fundamental and peak frequency and shorter notes than females. These results suggest that vocal differences between Northern and Wattled jacanas have the potential to act as a behavioral mediator of interspecific interactions, and that sex differences in vocalizations may relate to sex-role reversal in territorial defense and mate attraction.