There is significant evidence that the etiology of chronic otorhinolaryngology infections such as chronic rhinosinusitis, adenotonsillitis, and otitis media depends on biofilms. As biofilm-forming bacteria can be resistant to the immune system, antibiotics, and other treatments, biofilm infections are often chronic. To identify the genus and species of the clinical isolates obtained from the swabs collected from the patients with chronic infections of the nasal and paranasal sinus, nasopharynx, and oropharynx and to evaluate phenotypic and genotypic methods for the detection of biofilms and antimicrobial resistance among the isolated organisms. A total of 100 patients with chronic rhinosinusitis and adenotonsillitis participated in this study. Various clinical samples from the nasal cavity, nasopharynx, and oropharynx were obtained and subjected to microbiological analysis and biofilm-forming capacity by three methods: tube methods, Congo red staining, and microtiter plate method. The various specific genes were amplified by polymerase chain reaction. The amplified gene products were separated by gel electrophoresis. This was a prospective cohort study conducted on a total of 100 patients with chronic rhinosinusitis and adenotonsillitis. The age of the study participants was between 7 and 53years with a mean age of 29.22 ± 15.03. This study included 54 (54%) nasal tissue samples and 46 (46%) adenotonsillar tissue. The frequently cultured organisms are coagulase-negative staphylococci (17%), E. coli (10%), Citrobacter (10%), and Klebsiella (7%). Staphylococcus aureus (4), and Methicillin-resistant Staphylococcus aureus (3) produced strong biofilm. Acenobacter (3), Citrobacter (4), and E. coli (4) showed moderate biofilm production. Coagulase-negative Staphylococcus aureus (11), E. coli (6), and Klebsiella (7) showed weak biofilm formation. Citrobacter (6), and Coagulase negative Staphylococcus aureus (6) were negative for biofilm production. Staphylococcus aureus expressed mecA gene (3) and Panton-Valentine Leukocidin gene (2), Pseudomonas expressed mucA gene (2), Citrobacter expressed blaCARB-2 (4) qnrA gene (2), E. coli expressed bla SHV (2) and bla TEM1 gene (2) and Klebsiella expressed Kfu (2) and uge (1). Acenobacter was negative for blaIMP1, blaVIM2 genes. This study adds to the information on the common pathogens-forming biofilms in various nasal pathologies and adenotonsillitis. The knowledge that a particular organism has a higher biofilm-forming capacity will help to sensitize the physician that factors such as biofilms may be at play and take appropriate measures.