The etiology of head and neck squamous cell carcinoma (HNSCC) is not fully understood. While risk factors such as positive human papilloma virus (HPV) status, smoking and tobacco use have been identified, they do not account for all cases of the disease. We aimed to characterize the bacteriome, mycobiome and mycobiome-bacteriome interactions of oral wash in HNSCC patients and to determine if they are distinct from those of the oral wash of matched non-HNSCC patients. Oral wash samples were collected from 46 individuals with HNSCC and 46 controls for microbiome analyses. We identified three fungal phyla and eleven bacterial phyla of which Ascomycota (fungi, 72%) and Firmicutes (bacteria, 39%) were the most dominant, respectively. A number of organisms were identified as being differentially abundant between oral wash samples from patients with HNSCC and oral wash samples from those without HNSCC. Of note, strains of Candida albicans and Rothia mucilaginosa were differentially abundant and Schizophyllum commune was depleted in those with HNSCC compared to oral wash from those without HNSCC. Our results suggest that the oral cavity of HNSCC patients harbors unique differences in the mycobiome, bacteriome, and microbiome interactions when compared to those of control patients.