Cystic fibrosis (CF) is the most common lethal autosomal recessive disorder in Caucasians. Patients with classic CF have two mutations in CFTR and manifest symptoms in the sweat gland, respiratory tract, gastrointestinal tract, and male reproductive tract. Non-classic CF patients manifest disease in only a subset of the organ systems affected in classic CF. Most patients with non-classic CF have two mutations in CFTR, however a subset of patients have zero or one CFTR mutation. The goal of this work was to elucidate the molecular etiology of non-classic CF in patients without two CFTR mutations. Haplotype analysis in 3 families with multiple affected siblings confirmed that there is genetic heterogeneity in CF. To identify other genes that play a role in non-classic CF, we sequenced the coding regions of five genes that play a role in epithelial fluid or electrolyte transport (SCNN1A, SCNN1B, SCNN1G, AQP5, and AQP2) in a group of non-classic CF patients without mutations in the coding region of CFTR. We identified two patients with novel mutations in the beta subunit of the epithelial sodium channel (SCNN1B). These findings suggest that βENaC may play a unique role in the airways and that mutations in SCNN1B can have unexpected pleiotropy resulting in a phenotype resembling CF. We also identified a novel AQP2 mutation in two siblings with non-classic CF. Functional analysis is ongoing to determine the contribution of this mutation to the development of non-classic CF. Additionally, we have explored the role of CFTR in the etiology of non-classic CF in patients who have one CFTR mutation identified after screening. Sequencing of the coding regions of CFTR identified a second mutation in 6 of 9 patients. Lastly, we identified two patients with one CFTR mutation identified after sequencing that have a decrease in CFTR expression from the unknown allele, suggesting that they have a 2^nd unidentified mutation in a non-coding region of CFTR. Collectively, this work illustrates that mutations in CFTR play a major role in the etiology of non-classic CF, but challenges the paradigm that CF is a monogenic disorder, suggesting that genetic heterogeneity should be considered in patients without mutations in the coding region of CFTR.