Molecular Analysis of Resistance Genes in Multidrug-Resistant Klebsiella Pneumoniae İsolates

Abstract

Purpose: Development of antimicrobial resistance in Kelbsiella pneumoniae is a major problem in our country as well as in the whole world. Resistant K. pneumoniae is a bacterium that the World Health Organization has given critical priority in the list of the most dangerous pathogens for which antimicrobial drug development is needed. The aim of our study was to investigate carbapenem and colistin resistance genes in multidrug resistant K. pneumoniae isolates in a tertiary hospital in Turkey. Materials and Methods: Identification and antibiotic susceptibility tests were performed by Vitek-2 (bioMerieux, France). results for carbapenem and colistin were confirmed by gradient diffusion and microdilution methods. Carbapenemase and mcr genes were investigated by Polymerase Chain Reaction (PCR). Results: The blaNDM-1, blaOXA-48, blaKPC, blaVIM and blaIMP genes that cause resistance to carbapenem group antibiotics were tried to be determined by PCR in 69 isolates. As a result of the experiments, 54 (78%) of 69 isolates were positive for blaOXA-48 and 13 (19%) for blaNDM-1. KPC, VIM and IMP type beta-lactamases were not detected in any isolate. NDM-1 and OXA-48 type carbapenemases were found to be coexisting in 11 K. pneumoniae isolates. The mcr-1, mcr-2, mcr-3, mcr-4 and mcr-5 genes were not detected in any of the colistin resistant isolates. Conclusion: Colistin and carbapenem resistance mechanisms should be investigated by molecular methods in tertiary hospitals. The resistance profile data from the present study can be used as a knowledge base to understand the prevalent infection pattern in the hospital.

Keywords

• carbapenem
• carbapenemase
• colistin
• mcr

Çok İlaca Dirençli Klebsiella Pneumoniae İzolatlarında Direnç Genlerinin Moleküler Analizi

Abstract

Amaç: Klebsiella pneumoniae’da antimikrobiyal direnç gelişimi tüm dünyada olduğu gibi ülkemizde de büyük bir problemdir. Dirençli K. pneumoniae, Dünya Sağlık Örgütünün antimikrobiyal ilaç geliştirmeye ihtiyaç duyulan en tehlikeli patojenlerin listesinde kritik öncelik verdiği bir bakteridir. Bu çalışmanın amacı, Türkiye'de üçüncü basamak bir hastanede çok ilaca dirençli K. pneumoniae izolatlarında karbapenem ve colistin direnç genlerinin araştırılmasıdır. Araçlar ve Yöntem: Tanımlama ve antibiyotik duyarlılık testleri Vitek-2 (bioMerieux, Fransa) tarafından gerçekleştirildi. Karbapenem ve kolistin için sonuçlar gradyan difüzyon ve mikrodilüsyon yöntemleriyle doğrulandı. Karbapenemaz ve mcr genleri Polimeraz Zincir Reaksiyonu (PCR) ile araştırıldı. Karbapenem grubu antibiyotiklere direnç oluşumunu sağlayan blaNDM-1, blaOXA-48, blaKPC, blaVIM ve blaIMP genleri 69 izolatta çalışıldı. Bulgular: Yapılan deneyler sonucunda 69 izolatın 54’ünde (%78) blaOXA-48 ve 13’ünde (%19) blaNDM-1 pozitif olduğu tespit edildi. Hiçbir izolatta KPC, VIM ve IMP tipi beta-laktamaz belirlenemedi. Çalışmamızda 11 (%16) K. pneumoniae izolatında NDM-1 ve OXA-48 tipi karbapenemazın birlikte bulunduğu saptandı. Kolistin dirençli izolatların hiçbirinde mcr-1, mcr-2, mcr-3, mcr-4 ve mcr-5 genleri tespit edilememiştir. Sonuç: Üçüncü basamak hastanelerde kolistin ve karbapenem direnç mekanizmalarını moleküler yöntemlerle araştırılmalıdır. Mevcut çalışmadan elde edilen direnç profili verileri, hastanede yaygın olan enfeksiyon örüntüsünün anlaşılması için bir bilgi tabanı olarak kullanılabilir.

Keywords

• karbapenem
• karbapenemaz
• kolistin
• mcr

References

1. 1. Wyres KL, Lam MMC, Holt KE. Population genomics of Klebsiella pneumoniae. Nat Rev Microbiol. 2020;18(6):344-359.
2. 2. WHO. World Health Organization. WHO publishes list of bacteria for which new antibiotics are urgently needed. https://www.who.int/news/item/27-02-2017-who-publishes-list-of-bacteria-for-which-new-antibiotics-are-urgently-needed Accessed 23 September, 2024.
3. 3. Ling Z, Yin W, Shen Z, et al. Epidemiology of mobile colistin resistance genes mcr-1to mcr-9. J Antimicrob Chemother. 2020;75(11):3087-3095.
4. 4. EUCAST The European Committee on Antimicrobial SusceptibilitynTesting (2023) Breakpoint tables for interpretation of MICs and zone diameters. Version 13.0. http://www.eucast.org. Accessed 1 January, 2024.
5. 5. Poirel L, Heritier C, Tolun V, Nordmann P. Emergence of oxacillinase-mediated resistance to imipenem in Klebsiella pneumoniae. Antimicrob Agents Chemother. 2004;48(1):15-22.
6. 6. Schechner V, Straus-Robinson K, Schwartz D, et al. Evaluation of PCR-based testing for surveillance of KPC-producing carbapenem-resistant members of the Enterobacteriaceae family. J Clin Microbiol. 2009;47(10):3261-3265.
7. 7. Jeon BC, Jeong SH, Bae IK, et al. Investigation of a nosocomial outbreak of imipenem-resistant Acinetobacter baumannii producing the OXA-23 β-lactamase in Korea. J Clin Microbiol. 2005;43(5):2241-2245.
8. 8. Rita RA, Valeria B, Jette SK, et al. Multiplex PCR for detection of plasmid-mediated colistin resistance determinants, mcr-1, mcr-2, mcr-3, mcr-4 and mcr-5 for surveillance purposes. Euro Surveill. 2018;23(6):17-672.

9. 9. Kalem F, Ergun AG, Ertuğrul O, et al. Colistin resistance in Carbapenem-resistant Klebsiella pneumoniae strains. Biomed Res. 2016;27(2):368-372.
10. 10. Gümüş HH, Köksal F. Carbapenem-resistant Klebsiella pneumoniae: Resistance mechanisms, epidemiology, and mortality. FLORA. 2023;28(2):131-143.
11. 11. Zarakolu P, Eser OK, Aladag E, et al. Epidemiology of carbapenem-resistant Klebsiella pneumoniae colonization: a surveillance study at a Turkish university hospital from 2009 to 2013. Diagn Microbiol Infect Dis. 2016;85(4):466-470.
12. 12. Alp E, Perçin D, Colakoğlu S, et al. Molecular characterization of carbapenem-resistant Klebsiella pneumoniae in a tertiary university hospital in Turkey. J Hosp Infect. 2013;84(2):178-180.
13. 13. Sağıroğlu P, Hasdemir U, Altınkanat Gelmez G, Aksu B, Karatuna O, Söyletir G. Performance of “RESIST-3 O.K.N. K-SeT” immunochromatographic assay for the detection of OXA-48 like, KPC, and NDM carbapenemases in Klebsiella pneumoniae Turkey. Braz J Microbiol. 2018;49(4):885-890.
14. 14. Süzük Yıldız S, Şimşek H, Bakkaloğlu Z, et al. Türkiye’de 2019 yılı içinde izole edilen Escherichia coli ve Klebsiella pneumoniae izolatlarında karbapenemaz epidemiyolojisi. Mikrobiyol Bul. 2021;55(1):1-16.
15. 15. Köle M, Sesli Çetin E, Şirin MC, Arıdoğan BC. Seftazidim- avibaktam, meropenem ve kolistinin tek başına ve ikili kombinasyonlarının çeşitli klinik örneklerden izole edilen karbapenem dirençli Klebsiella pneumoniae suşlarına karşı in-vitro etkinliğinin araştırılması. Mikrobiyol Bul. 2022;56(2):230-250.
16. 16. Yanat B, Machuca J, Yahia RD, Touati A, Pascual Á, Rodríguez-Martínez JM. First report of the plasmid- mediated colistin resistance gene mcr-1 in a clinical Escherichia coli isolate in Algeria. Int J Antimicrob Agents. 2016;48(6):760-761.
17. 17. Liu YY, Wang Y, Walsh TR, et al. Emergence of plasmid-mediated colistin resistance mechanism mcr-1 in animals and human beings in China: a microbiological and molecular biological study. Lancet Infect Dis. 2016;16(2):161-168.
18. 18. Richter SE, Miller L, Uslan DZ, et al. Risk factors for colistin resistance among Gram-negative rods and Klebsiella pneumoniae isolates. J Clin Microbiol. 2018;56(9):10-1128.
19. 19. Arabacı Ç, Dal T, Başyiğit T, Genişel N, Durmaz R. Investigation of carbapenemase and mcr-1 genes in carbapenem-resistant Klebsiella pneumoniae isolates. J Infect Dev Ctries. 2019;13(6):504-509.
20. 20. Özkaya E, Buruk CK, Tosun İ, Toraman B, Kaklıkkaya N, Aydın F. Investigation of plasmid mediated mcr colistin resistance gene in clinical Enterobacterales ısolates. Mikrobiyol Bul. 2020;54(2):191-202.
21. 21. Al-Tawfiq JA, Laxminarayan R, Mendelson M. How should we respond to the emergence of plasmid-mediated colistin resistance in humans and animals? Int J Infect Dis. 2017;54:77-84.
22. 22. Karki D, Dhungel B, Bhandari S, et al. Antibiotic resistance and detection of plasmid mediated colistin resistance mcr-1 gene among Escherichia coli and Klebsiella pneumoniae isolated from clinical samples. Gut Pathog. 2021;13(1):45.
23. 23. Liassine N, Aaaouvie L, Descombes MC, et al. Very low prevalence of MCR-1/MCR-2 plasmid-mediated colistin-resistance in urinary tract Enterobacteriaceae in Switzerland. Inter J Infect Dis. 2016;51:4-5.
24. 24. Castanheira M, Griffin MA, Deshpande LM, Mendes RE, Jones RN, Flamm RK. Detection of mcr-1 among Escherichia coli clinical isolates collected worldwide as part of the SENTRY antimicrobial surveillance program in 2014 and 2015. Antimicrob Agents Chemother. 2016;60(9):5623-5624.
25. 25. Sarı AN, Süzük S, Karatuna O, et al. Results of a multicenter study investigating plasmid mediated colistin resistance genes (mcr-1 and mcr-2) in clinical Enterobacteriaceae ısolates from Turkey. Bull Microbiol. 2017;51(3):299-303.
26. 26. Manohar P, Shanthini T, Ayyanar R, et al. The distribution of carbapenem- and colistin- resistance in Gram-negative bacteria from the Tamil Nadu region in India. J Med Microbiol. 2017;66(7):874-883.
27. 27. Eiamphungporn W, Yainoy S, Jumderm C, Tan-arsuwongkul R, Tiengrim S, Thamlikitkul V. Prevalence of the colistin resistance gene mcr-1 in colistin- resistant Escherichia coli and Klebsiella pneumoniae isolated from humans in Thailand. J Global Antimicrob Res. 2018;2018(15):32-35.
28. 28. Moosavian M, Emam N. The first report of emerging mobilized colistin resistance (mcr) genes and ERIC-PCR typing in Escherichia coli and Klebsiella pneumoniae clinical isolates in southwest Iran. Infect Drug Resist. 2019;12:1001-1010.
29. 29. Tekin R, Dal T, Pirinccioglu H, Oygucu SH. A 4-years surveillance of device-associated nosocomial infections in a neonatal intensive care unit. Pediatr Neonatol. 2013;54(5):303-308.