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Background Recently, biofilms have been implicated in the pathogenesis of recalcitrant chronic rhinosinusitis (CRS). We sought to determine the prevalence of biofilm-forming cultures obtained from patients with CRS and clinical factors that may contribute to biofilm formation.Methods Endoscopically guided sinonasal cultures were obtained in duplicate from CRS patients with evidence of mucopurulence. Bacterial swabs were sent for microbiological characterization and were simultaneously evaluated for biofilm-forming capacity by a modified Calgary Biofilm Detection Assay. Biofilm formation was based on concomitant values of biofilm-forming Pseudomonas aeruginosa O1 (PAO1) (positive control) and non-biofilm-forming mutants sad-31 (type IV pili) and sad-36 (flagella K; negative control). Samples, with growth greater than the sad-31 mutant, were designated as biofilm formers.Results Sinonasal cultures were obtained from 157 consecutive patients (83 female patients) over a 4-month period. Forty-five samples (28.6%) showed biofilm formation. Among patients with a prior history of functional endoscopic sinus surgery (FESS), 30.7% (n = 42) showed biofilm growth. For patients naive to surgical intervention (n = 20), only 15% showed biofilm formation. A positive, statistically significant correlation existed between biofilm formation and number of prior FESS procedures. Polymicrobial cultures, Pseudomonas aeruginosa, and/or Staphylococcus aureus comprised 71% of samples. Chi-squared analysis showed an association with prior infections, but not with any pharmacologic therapy or comorbidies.Conclusion We show a high percentage of CRS patients (28.6%) whose sinonasal mucopurulence has biofilm-forming capacity. Postsurgical patients had a high prevalence of biofilm-forming bacteria, a possible reflection of the severe nature of their disease. Additional studies are warranted.