Molecular Typing with PFGE Method of Methicillin-resistant Staphylococcus aureus Isolates from in a University Hospital

Öz

Objectives: Staphylococcus aureus is a pathogen that causes many infections in humans, and the emergence of methicillin-resistant (MRSA) isolates in the last fifty years poses a threat to society and hospitals. Preventing the spread of MRSA strains between and within hospitals can only be achieved with a correct typing. The PFGE method is considered to be the gold standard among molecular typing methods. Methods: In this study, a clonal relationship between nine methicillin-resistant S. aureus isolates isolated from Van Yüzüncü Yıl University hospital was determined .Pfge analysis was done with the Bionumerics program. The similarity index was created at a tolerance value of 1% using the “Dice” coefficient. The clustering relationship between isolates was demonstrated using the UPGMA (unweighted pair group method of arithmetic averages) method. The principles set out by Tenover et al. were adopted for the evaluation of genotypic association between isolates Results: As a result of the analysis 4 independent genotypes were identified. It was determined that there were 2 related isolates in 3 genotypes. Conclusion: In our study, it was determined that there was not a dominant MRSA clone in different years in the hospital studied, since samples from different years were studied. However, it is thought that it would be beneficial to increase the precautions in order to prevent the distribution of infection in all hospitals and to control the infection in all areas.

Anahtar Kelimeler

• Methicillin-resistant Staphyococcus aureus
• Molecular typing
• Pulsed Field Gel Electrophoresis
• molecular epidemiology

Bir Üniversite Hastanesinden Izole Edilen Metisilin Dirençli Stapylococcus Aureus Izolatlarının PFGE Yöntemi ile Moleküler Tiplendirilmesi

Öz

Amaç Staphylococcus aureus insanlarda birçok enfeksiyona neden olan bir patojen olup, son elli yılda metisiline dirençli (MRSA) izolatların ortaya çıkması toplum ve hastaneler için tehdit oluşturmaktadır. MRSA suşlarının hastaneler arasında ve hastaneler içinde yayılmasının önlenmesi ancak doğru bir tipleme ile sağlanabilir. PFGE yöntemi, moleküler tipleme yöntemleri arasında altın standart olarak kabul edilmektedir. Gereç ve Yöntem Bu çalışmada, Van Yüzüncü Yıl Üniversitesi hastanesinden izole edilen dokuz metisiline dirençli S. aureus izolatı arasında klonal bir ilişki belirlendi. Bionumerics programı ile PFGE analizi yapıldı. Benzerlik indeksi “Dice” katsayısı kullanılarak %1 tolerans değerinde oluşturuldu. İzolatlar arasındaki kümeleme ilişkisi, UPGMA (ağırlıksız çift grup aritmatik ortalamalar yöntemi) yöntemi kullanılarak gösterildi. Tenover ve arkadaşlarının kriterleri izolatlar arasındaki genotipik ilişkinin değerlendirilmesi için kabul edildi. Bulgular Analiz sonucunda 4 bağımsız genotip tespit edilmiştir. 3 genotipte 2 ilişkili izolat olduğu belirlendi.SonuçÇalışmamızda, farklı yıllardan örnekler çalışıldığı için çalışılan hastanede farklı yıllarda baskın bir MRSA klonu olmadığı belirlendi. Ancak enfeksiyonun tüm hastanelerde yayılmasını önlemek ve enfeksiyonun her alanda kontrol altına alınması için tedbirlerin artırılmasının faydalı olacağı düşünülmektedir.

Anahtar Kelimeler

• Metisilin Dirençli Staphyococcus aureus
• Moleküler Tiplendirme
• Değişken Alanlı Jel Elektroforezi
• Moleküler Epidemiyoloji

Kaynakça

1. Yilmaz S, Kiliç A, Karagöz A, Bedi̇r O, üsküdar Güçlü A, Başustaoglu CA, et al. Investigation of Various Virulence Factors Among the Hospital and Community-Acquired Staphylococcus aureus Isolates by Real-Time PCR Method. Microbiol. Bul.2021;11.
2. Chung S, Yi J, Jang MH, Joo SI, Ra EK, Kim SY, et al. Comparison of modified multiple-locus variable-number tandem-repeat fingerprinting with pulsed-field gel electrophoresis for typing clinical isolates of Staphylococcus aureus. Ann Lab Med. 2012;32(1):50–6.
3. Gulbandilar A. Investigation of Staphylococcus aureus Carriage and Antibiotic Sensitivity in Nose Mucous Membrane in Kutapha Province.T.R. Kutahya Dumlupınar University Journal of The Institute Of Science and Technology. 2009; 18: 1-5.
4. CDC, Ncird. Laboratory Methods for the Diagnosis of Meningitis - CHAPTER 12: Characterization of Neisseria meningitidis, Haemophilus influenzae, and Streptococcus pneumoniae by Molecular Typing Methods. http://www.chem.qmul.ac.uk/iupac/.
5. Brehony, C., K. A. Jolley, and M. C. J. Maiden. 2007. Multilocus sequence typing for global surveillance of meningococcal disease. FEMS Microbiology Reviews 31:15-26.
6. Dowson, C. G., A. Hutchison and BGS. Extensive re-modelling of the transpeptidase domain of penicillin-binding protein 2B of a penicillin-resistant South African isolate of Streptococcus pneumoniae. Mol Microbiol. 1989;3:95–6.
7. Enright, M. C. and BGS. Multilocus sequence typing. Trends Microbiol. 1999;7:482–7.
8. Xilong Kang, Chuang Meng ZP. Molecular typing of Salmonella by Pulsed-Field Gel Electrophoresis. 2020. 187–196 p.

9. Acar S, Levent B, Atalan E. A molecular epidemiological investigation of multistate outbreaks of Salmonella Enteritidis from clinical and environmental samples in Turkey, 2000–2010. Turkish J Med Sci. 2015;45(1):76–83.
10. Strandén A, Frei R, Widmer AF. Molecular typing of methicillin-resistant Staphylococcus aureus: Can PCR replace pulsed-field gel electrophoresis? J Clin Microbiol. 2003; 41(7):3181–6.
11. Golding GR, Campbell J, Spreitzer D, Chui L. Pulsed-field gel electrophoresis of Staphylococcus aureus. Methods Mol Biol. 2015;1301:85–93.
12. Clinical and Laboratory Standards Institute. Performance Standards for Antimicrobial Susceptibility Testing. Document M100-S15. 5th Informational Supplement. Wayne, PA: CLSI. In 2005. p. Document M100-S15. 5th Informational.
13. Sci TJM. Molecular typing of methicillin-resistant Staphylococcus aureus isolated from bloodstream infections in a university hospital. 2009;39(6):959–68.
14. Maslow JN, Slutsky AM, Arbeit RD. In: Persing HD ST, Eds TF. Diagnostic Application of pulsed-field gel electrophoresis to molecular epidemiology. Am Soc Microbiol. 1993;(562–72).
15. Tenover FC, Arbeit RD, Goering RV, Mickelsen PA MB, Al. PD et. Interpreting chromosomal DNA restriction patterns produced by pulsed-field gel electrophoresis: criteria for bacterial strain typing. J Clin Microbiol. 1995;33:2233–9.
16. Tekeli̇ A, Koyuncu E, Dolapçi İ, Arikan Akan Ö, Ceren KARAHAN Z, A. Molecular Characteristics of Methicillin-Resistant Staphylococcus aureus Strains Isolated From Blood Cultures Between 2002-2005 in Ankara University Hospital.Mikrobiyol Bul. 2009;43:1–10.
17. Harmsen D, Claus H, Witte W et al. Typing of methicillin-resistant Staphylococcus aureus in a university hospital setting by using novel software for spa repeat determination and database management. J Clin Microbiol. 2003;41(12):5442–8.
18. Peacock SJ, de Silva GD, Justice A et al. Comparison of multilocus sequence typing and pulsed-field gel electrophoresis as tools for typing Staphylococcus aureus isolates in a microepidemiological setting. J Clin Microbiol. 2002;40(10):3764–70.
19. Wisplinghoff H, Ewertz B, Wisplinghoff S et al. Molecular evolution of methicillin-resistant Staphylococcus aureus in the metropolitan area of Cologne, Germany, from 1984 to 1998. J Clin Microbiol. 2005;43(11):5445–51.
20. Robinson DA EM. Evolutionary models of the emergence of methicillin-resistant Staphylococcus aureus. Antimicrob Agents Chemother. 2003;47(12):3926–34.
21. Olive M BP. Principles and applications of methods for DNA-based typing of microbial organisms. J Clin Microbiol. 1999;37:1661–9.
22. Staphylococcus aureus as source of catheter-related bloodstream infection evaluated by PFGE and rep-PCR typing in a Brazilian hospital. APMIS. 2008;116:953–60.