Anton Christanto1, Jaka Widada2, Venzha3
1.Department of Otolaryngology Head and Neck Surgery, Faculty of Medicine, University of Gadjah Mada/DR. Sarjito Hospital, Yogyakarta
2. Departement of Biotechnology Study Program, University of Gadjah Mada, Yogyakarta
3. The House of Natural Fiber, Yogyakarta New Media Art Laboratory, Yogyakarta |
ABSTRACT
Background: Active benign chronic suppurative otitis media (ABCSOM) is a potentially serious disorder, since there is a risk of permanent hearing lost, and mainly caused by bacterial infections. Traditionally, the identification and enumeration of bacterial species has depended entirely upon pure-culture techniques. However, the difficulty with which some types of organisms are cultured, particularly fastidious and/or anaerobic bacteria means that the more easily grown species in a mixed microbial community are likely to be overrepresented by cultivation and plate counts. Bacterial molecular-analyses have provided a new perspective on bacterial analysis by allowing the detection and identification of constituent microorganisms in the absence of cultivation.
Objective: In this study we established a culture-independent by using ribosomal intergenic spacer analysis (RISA) to survey bacteria present in the active benign chronic suppurative otitis media without fastidious cultivation.
Methods: Middle ear fluid discharge samples were collected from 5 individuals of ABCSOM using sterile syringe equipped with intravenous catheter. Samples collected in the laboratory were preserved with isopropanol and immediately frozen and stored at -20°C prior to DNA extraction. Total DNA was direct extracted from the samples, and the bacterial constituents were analyzed by RISA.
Results: Several bacterial species were detected using RISA, whereas culture dependent examination based approach (plate method) revealed only one type of bacterium from all samples.
Conclusion: The results showed that culture-independent approach (RISA) is the robust method to determine bacterial constituent middle ear fluid discharge. Further investigation such sequencing analysis is needed to determine phylogenetic affiliation of bacterial species residing in middle ear fluid discharge.
Key words: ABCSOM, ribosomal intergenic spacer analysis, culture independent, bacterial community
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