Anuran vocalization is species-specific and bioacoustic analyses are important tools for species delimitation. The present study describes the vocalization of Sphaenorhynchus mirim, recognizing two different types of call, here referred to as call type A and call type B. We analyzed a total of 86 calls of type A and 15 calls of type B. Call type A consists of a single multipulsed note with duration of 34.12–101.00 ms, with 9–25 pulses/note, pulse rate ranges from 240.40–338.21 pulses/sec and the dominant frequency is between 3085.51–3398.86 Hz. We arbitrarily subdivided the type B call into two patterns: Bd (with well-defined notes) and Bnd (with non-defined notes). Call type B with well-defined notes (n = 4) has 3–5 multipulsed notes, duration of 180.12–262.68 ms, note duration of 15.62–100.75 ms, interval between notes with 7.93–23.12 ms, 2–22 pulses/note and dominant frequency of 3067.77–3180.11 Hz. Call type B with non-defined notes (n = 11) has duration of 143.75–-218.12 ms; 26–32 pulses/ call and a dominant frequency of 3180.11–3375.22 Hz. Sphaenorhynchus mirim vocalization differs from all other species in the genus by its higher dominant frequency.