Mutations in the SLC26A4 gene can cause both Pendred syndrome and nonsyndromic enlargement of the vestibular aqueduct, two conditions associated with sensorineural hearing loss. We analyzed the SLC26A4 gene in 44 hearing-impaired patients by nested polymerase chain reaction followed by high-resolution melt analysis. We also used this approach to scan for mutations in KCNJ10 and FOXI1, two genes reported to play a role in the pathogenesis of Pendred syndrome and enlarged vestibular aqueduct. Seven patients with known SLC26A4 mutations were included as controls. All previously identified mutations were detected by high-resolution melt analysis. Of the patients with no known mutations, we detected two SLC26A4 mutations in 5 probands (12%), one mutation in 9 probands (21%), and no mutations in 29 probands (67%). We identified two novel SLC26A4 mutations, p.T485M and p.F718S, and found no evidence of a digenic contribution of KCNJ10 and FOXI1 mutations.