Detection of pulmonary blastomycosis in patients with different respiratory diseases using conventional and molecular methods
ازهار عبد الفتاح ابراهيم
Authors : Zahraa Q. Abdulrahman1*, Azhar A. F. Al-Attraqchi1 , Haider N. D. Al-Taee2, Haider F. Ghazi1
Blastomycosis is a rare but important fungal infection cause by dimorphic fungus Blastomyces dermatitidis.Pneumonia is the most common manifestation and the lung is almost always the organ initially infected. The performance of blastomycosis diagnosis using microscopic examination and culture are highly dependent on the quality of the clinical specimens and the experience of the laboratory personnel. Moreover, these classical methods have previously shown lower sensitivity in fungal detection compared to molecular methods.Determination the frequency of pulmonary blastomycosis in patients with respiratory symptoms by using conventional (Wet mounting, culture and cytology) and molecular methods (conventional PCR and real time PCR), then comparing between conventional and molecular methods concerning sensitivity and specificity.This study involved 132patients who attended Baghdad Teaching Hospital, Al-Emamein Al- Kadhimein Medical City, Al-Amal National Hospital and Chest and Respiratory Diseases Institute/ Baghdad Medical City who enrolled in this cross sectional study from September(2018) to the end of August (2019). Sputum, bronchial washings, bronchoalveolar lavage (BAL) or pleural fluid were collected from patients with different age groups of both males and females all were diagnosed clinically with pulmonary symptoms.Sample (132) were obtained from patients in dry and clean screw cupped bottles. Fresh samples were divided into two parts, for mycological detection by conventional (Wet mounting, culture and cytology) and molecular methods (conventional PCR and real time PCR).Laboratory diagnosis of blastomycosis been done using different procedures, starting with conventional methods (culture, wet mounting and cytology) followed by two molecular methods. The conventional PCR with ITS1 and ITS4 gene was showed nonspecific result with gel electrophoresis of eighteen positive samples with amplicon size of approximated 600 bp which were recorded as suspectedBlastomycesdermatitidis. It have been recorded that all suspected cases (18/18) were recovered from sputum only. According to wet mount, only one out of eighteen was matches the suspected B.dermatitidis. In case of cytology and PAD culture, none of the eighteen samples were recognized as Blastomyces spp. Concerning qPCR using BAD1 gene, it was seen there was no specific band for Blastomyces hence, these results was negative.This study found no evidence of blastomycosis in samples from Iraqipatients with respiratory symptoms based on BAD1 gene detection using real time PCR and culture, however other tests like wet mountand ITS1-4 based conventional PCR gave false positive results.

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Nov. 2020