Molecular Diagnosis of Fungal Sinusitis Using Limit of Detection

Abstract

Background: As an inflammatory process that involves the paranasal sinuses, chronic sinusitis (CS) is one of the most prevalent chronic illnesses that affects all age groups. Parasitic fungi are involved in sinusitis infections. Objective: This study is aimed at the molecular detection of sinusitis caused by such fungi. Methods: Seventy-two samples were collected from the secretions of maxillary and frontal sinuses of patients from Rasoul-e Akram (PbUH) Hospital in Tehran during sinus operation. Fungal genomic DNA was extracted by a DNP kit. The detection of fungi was carried out by employing a sequence-specific target, namely mt cyte b gene locus, and using primers. Polymerase chain reaction (PCR) was optimized, and the limit of detection (LOD) and specificity tests were performed. The amplicon was cloned by the T/A cloning method, which was used for sequencing and positive control. Results: The 430-bp PCR product underwent appropriate propagation before being amplified and was observed on 1.5% electrophoreses gel. The evaluation of the selected primers with seven DNA constructs from another microorganisms demonstrated 100% specificity. The limit of detection of the optimized test was evaluated up to 50 fungi. Out of 72 samples, 9.7% were positive for fungi existence. Conclusions: This study indicated that molecular diagnosis of the target mt cyte b gene using LOD enhances clinical laboratory detection of fungal sinusitis.