Monitoring methicillin-resistant bacteria in river water by using meca-specific DNA probe

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Tezin Türü: Yüksek Lisans

Tezin Yürütüldüğü Kurum: Orta Doğu Teknik Üniversitesi, Fen Bilimleri Enstitüsü, Biyokimya Anabilim Dalı, Türkiye

Tezin Onay Tarihi: 2014

Tezin Dili: İngilizce

Öğrenci: ELNAZ SEYEDMONİR

Danışman: Bülent İçgen

Özet:

Aquatic ecosystems represent important vehicles for the dissemination of not only antibiotic resistant bacteria but also antibiotic resistance genes. Of particular interest are methicillin-resistant staphylococci (MRS) harbouring mecA gene that confers their resistance to β-lactams. Therefore, in this study, water samples collected from different locations of a river impacted by surrounding facilities and domestic effluents were analysed to learn more about the occurrence of MRS and mecA gene. Out of 290, 12 surface water isolates displayed resistance to both cefoxitin and oxacillin (derivatives of methicillin) antibiotics. Cefoxitin/oxacillin-resistant surface water isolates were screened for the prevalence of mecA gene by a polymerase chain reaction method. All of the cefoxitin/oxacillin-resistant surface water isolates including 4 Staphylococcus, 7 Pseudomonas, and one Aeromonas species, identified by 16S rRNA sequencing, were found out to harbor mecA. Following Sodium Dodecyl Sulfate Polyacrylamide Gel Electrophoresis (SDS-PAGE), Western Blot analysis of penicillin-binding protein (PBP2a) encoded by mecA revealed that staphylococcal PBP2a-specific antibodies were unsuccessful in detecting non-staphylococcal carriers. For this reason, mecA-specific DNA probe was also used to monitor all mecA harboring surface water isolates during the periods of two years in between 2011 and 2012 by using in situ fluorescent hybridization technique and image-analyzed microscopy. Our results indicated that the mecA-specific DNA probe might be a potential analytical tool in selecting and in situ monitoring of methicillin resistant isolates in surface waters. Once in the environment, bacteria of different origin come into physical contact and may exchange resistance genes with the indigenous bacterial population. Therefore, surface waters are not only hot spots for mecA harbouring staphylococcal isolates but also non-staphylococcal ones due to gene dissemination and require special scientific consideration.