Detection of Carbapenem Resistance Using the Genotypic and Phenotypic Methods in Klebsiella pneumoniae

Year 2024, Volume: 26 Issue: 1, 15 - 20, 30.04.2024

Mehmet Akif Durmuş , Mustafa Derya Aydin

https://doi.org/10.18678/dtfd.1383748

Cited By: 1

Abstract

Aim: This study aimed to detect the carbapenem resistance of the Klebsiella pneumoniae strains, isolated from clinical specimens with genotypic and phenotypic methods.
Material and Methods: A total of 87 Klebsiella pneumoniae strains whose carbapenem resistance was determined by disc diffusion method were included in the study. Carbapenemase was investigated using the combined disk method and polymerase chain reaction (PCR).
Results: The evaluation of the PCR results demonstrated that OXA was detected in 60 (68.9%) samples, NDM was detected in 20 (22.9%), OXA + NDM in 5 (5.7%), and KPC was detected in 1 (1.1%) out of 87 clinical samples. Carbapenemase was not detected in one specimen with the PCR method. The results were found compatible with the combined disc test results for all isolates which were detected as only OXA, NDM, and KPC type carbapenemase positive. In 5 (5.7%) strains in which the co-existence of NDM and OXA type carbapenemases was detected by PCR, the combined disc method detected only OXA type carbapenemase.
Conclusion: The combined disk method is inadequate in the presence of strains that have multiple carbapenemases, and also have OXA which is the most frequently detected carbapenemase in our hospital. EUCAST recommends verification by other methods in the presence of OXA-48. Genotypic methods can be used for confirmation testing. The detections of strains with NDM, multiple carbapenemases, and the first detection of KPC were striking in the study. Monitoring the spread of these strains in the hospital will be necessary for infection control.

Keywords

Klebsiella pneumoniae, carbapenem, carbapenemase

Klebsiella pneumoniae Suşlarında Karbapenem Direncinin Genotipik ve Fenotipik Yöntemler ile Saptanması

Abstract

Amaç: Bu çalışmada, klinik örneklerinden izole edilen Klebsiella pneumoniae suşlarında karbapenem direncinin genotipik ve fenotipik yöntemler ile saptanması amaçlanmıştır.
Gereç ve Yöntemler: Disk difüzyon yöntemi ile karbapeneme dirençli bulunan toplam 87 Klebsiella pneumoniae suşu çalışmaya dahil edilmiştir. Karbapenemaz varlığı kombine disk yöntemi ve polimeraz zincir reaksiyonu (polymerase chain reaction, PCR) ile araştırılmıştır.
Bulgular: PCR sonuçları değerlendirildiğinde, 87 klinik örnekten 60 (%68,9) örnekte OXA tipi, 20 (%22,9) örnekte NDM tipi, 5 (%5,7) örnekte OXA + NDM tipi ve 1 (%1,1) örnekte ise KPC tipi karbapenemaz saptandığı görülmektedir. Bir örnekte ise PCR yöntemi ile karbapenemaz bulunamamıştır. Tek başına OXA, NDM ve KPC tipi karbapenemaz pozitifliği saptanan izolatların tamamı için sonuçların kombine disk testi ile uyumlu olduğu bulunmuştur. PCR yöntemi ile NDM ve OXA tipi karbapenemazın birlikte olduğu 5 (%5,7) örnekte ise kombine disk yönteminde sonuç OXA tipi karbapenemaz olarak bulunmuştur.
Sonuç: Kombine disk yöntemi, aynı anda birden fazla karbapenemaz bulunduran suşların bulunması ve hastanemizde en sık saptanan karbapenemaz tipinin OXA olması nedeni ile yetersiz kalmaktadır. EUCAST, OXA-48 varlığında diğer yöntemlerle doğrulanmasını önermektedir. Doğrulama testi olarak genotipik yöntemler kullanılabilir. NDM tipi karbapenemazın artmakta olduğu, birden fazla karbapenemazı taşıyan suşların görülmeye başlaması ve ilk defa KPC tipi karbapenemazın bulunması dikkat çekicidir. Bu suşların hastanede yayılımının takip edilmesi enfeksiyon kontrolü açısından önemli olacaktır.

Keywords

Klebsiella pneumoniae, karbapenem, karbapenemaz

References

• Collignon PJ. 11: Antibiotic resistance. Med J Aust. 2002;177(6):325-9.
• Livermore DM. Minimising antibiotic resistance. Lancet Infect Dis. 2005;5(7):450-9.
• Queenan AM, Bush K. Carbapenemases: The versatile β-lactamases. Clin Microbiol Rev. 2007;20(3):440-58.
• Santajit S, Indrawattana N. Mechanisms of antimicrobial resistance in ESKAPE pathogens. Biomed Res Int. 2016;2016:2475067.
• Cohen Stuart J, Leverstein-Van Hall MA; Dutch Working Party on the Detection of Highly Resistant Microorganisms. Guideline for phenotypic screening and confirmation of carbapenemases in Enterobacteriaceae. Int J Antimicrob Agents. 2010;36(3):205-10.
• Ambler RP. The structure of beta-lactamases. Philos Trans R Soc Lond B Biol Sci. 1980;289(1036):321-31.
• Nordmann P, Naas T, Poirel L. Global spread of carbapenemase-producing Enterobacteriaceae. Emerg Infect Dis. 2011;17(10):1791-8.
• Bradford PA, Bratu S, Urban C, Visalli M, Mariano N, Landman D, et al. Emergence of carbapenem-resistant Klebsiella species possessing the class A carbapenem-hydrolyzing KPC-2 and inhibitor-resistant TEM-30 beta-lactamases in New York City. Clinical Infectious Diseases. 2004;39(1):55-60.
• Bratu S, Landman D, Haag R, Recco R, Eramo A, Alam M, et al. Rapid spread of carbapenem-resistant Klebsiella pneumoniae in New York City: A new threat to our antibiotic armamentarium. Arch Intern Med. 2005;165(12):1430-5.
• Bennett JE, Dolin R, Blaser MJ, editors. Mandell, Douglas, and Bennett’s principles and practice of infectious diseases. Philadelphia, PA: Elsevier Saunders; 2015.
• Nordmann P, Poirel L, Walsh TR, Livermore DM. The emerging NDM carbapenemases. Trends Microbiol. 2011;19(12):588-95.
• Yong D, Toleman MA, Giske CG, Cho HS, Sundman K, Lee K, et al. Characterization of a new metallo-β-lactamase gene, bla NDM-1, and a novel erythromycin esterase gene carried on a unique genetic structure in Klebsiella pneumoniae sequence type 14 from India. Antimicrob Agents Chemother. 2009;53(12):5046-54.
• Poirel L, Héritier C, Tolün V, Nordmann P. Emergence of oxacillinase-mediated resistance to imipenem in Klebsiella pneumoniae. Antimicrob Agents Chemother. 2004;48(1):15-22.
• Nordmann P, Poirel L. The difficult-to-control spread of carbapenemase producers among Enterobacteriaceae worldwide. Clin Microbiol Infect. 2014;20(9):821-30.
• Doyle D, Peirano G, Lascols C, Lloyd T, Church DL, Pitouta JDD. Laboratory detection of Enterobacteriaceae that produce carbapenemases. J Clin Microbiol. 2012;50(12):3877-80.
• Clinical and Laboratory Standards Institute. Performance standards for antimicrobial susceptibility testing. CLSI supplement M100. 28th ed. Wayne, PA: Clinical and Laboratory Standards Institute; 2018.
• Saito R, Koyano S, Dorin M, Higurashi Y, Misawa Y, Nagano N, et al. Evaluation of a simple phenotypic method for the detection of carbapenemase-producing Enterobacteriaceae. J Microbiol Methods. 2015;108:45-8.
• Çakar A, Akyön Y, Gür D, Karatuna O, Öǧünç D, Özhak Baysan B, et al. Investigation of carbapenemases in carbapenem-resistant Escherichia coli and Klebsiella pneumoniae strains isolated in 2014 in Turkey. Mikrobiyol Bul. 2016;50(1):21-33. Turkish.
• Alp E, Perçin D, Colakoǧlu S, Durmaz S, Kürkcü CA, Ekincioǧlu P, et al. Molecular characterization of carbapenem-resistant Klebsiella pneumoniae in a tertiary university hospital in Turkey. J Hosp Infect. 2013;84(2):178-80.
• Gümüş HH, Köksal F. Carbapenem-resistant Klebsiella pneumoniae: Resistance mechanisms, epidemiology, and mortality. Flora. 2023;28(2):131-43.
• Süzük Yıldız S, Şimşek H, Bakkaloğlu Z, Numanoğlu Çevik Y, Hekimoğlu CH, Kılıç S, et al. The epidemiology of carbapenemases in Escherichia coli and Klebsiella pneumoniae isolated in 2019 in Turkey. Mikrobiyol Bul. 2021;55(1):1-16.
• Eser OK, Altun Uludağ H, Ergin A, Boral B, Sener B, Hasçelik G. Carbapenem resistance in ESBL positive Enterobacteriaceae isolates causing invasive infections. Mikrobiyol Bul. 2014;48(1):59-69. Turkish.
• Alkan Bilik Ö, Bayraktar M, Özcan N, Gül K, Akpolat N. Dissemination of bla OXA-48-like, bla NDM, bla KPC, bla IMP-1, bla VIM genes among carbapenem-resistant Escherichia coli and Klebsiella pneumoniae strains in Southeastern Turkey: first report of Klebsiella pneumoniae co-producing bla OXA-48-like, bla VIM and bla IMP-1 genes. Rev Med Microbiol. 2021;32(4):205-10.
• Demir HK, Nakipoglu Y. Investigation of the prevalence of carbapenem resistance genes in faecal carriage of carbapenem resistant Klebsiella spp. isolates by multiplex real-time PCR method. J Infect Dev Ctries. 2023;17(11):1606-12.
• Kalayci-Yuksek F, Gumus D, Uyanik-Ocal A, Gun G, Bayirli-Turan D, Macunluoglu AC, et al. Carbapenem and colistin resistance, integrons and plasmid replicon types in multi-drug resistant Klebsiella strains isolated in Turkey. Clin Lab. 2023;69(3):509-15.
• Labarca J, Poirel L, Özdamar M, Turkoglü S, Hakko E, Nordmann P. KPC-producing Klebsiella pneumoniae, finally targeting Turkey. New Microbes New Infect. 2014;2(2):50-1.
• Cizmeci Z, Aktas E, Otlu B, Acikgoz O, Ordekci S. Molecular characterization of carbapenem- resistant Enterobacteriaceae yields increasing rates of NDM-1 carbapenemases and colistin resistance in an OXA-48- endemic area. J Chemother. 2017;29(6):344-50.
• Samasti M, Koçoğlu ME, Davarcı İ, Vahaboğlu H, Çaşkurlu H. Investigation of carbapenemase genes and clonal relationship in carbapenem resistant Klebsiella pneumoniae strains. Bezmialem Science. 2019;7(3):186-90.
• Kilic A, Baysallar M. The first Klebsiella pneumoniae isolate co-producing OXA-48 and NDM-1 in Turkey. Ann Lab Med. 2015;35(3):382-3.
• Isler B, Özer B, Çınar G, Aslan AT, Vatansever C, Falconer C, et al. Characteristics and outcomes of carbapenemase harbouring carbapenem-resistant Klebsiella spp. bloodstream infections: a multicentre prospective cohort study in an OXA-48 endemic setting. Eur J Clin Microbiol Infect Dis. 2022;41(5):841-7.
• Yigit H, Queenan AM, Anderson GJ, Domenech-Sanchez A, Biddle JW, Steward CD, et al. Novel carbapenem-hydrolyzing β-lactamase, KPC-1, from a carbapenem-resistant strain of Klebsiella pneumoniae. Antimicrob Agents Chemother. 2001;45(4):1151-61.
• Nordmann P, Cuzon G, Naas T. The real threat of Klebsiella pneumoniae carbapenemase-producing bacteria. Lancet Infect Dis. 2009;9(4):228-36.
• Tängdén T, Giske CG. Global dissemination of extensively drug-resistant carbapenemase-producing Enterobacteriaceae: Clinical perspectives on detection, treatment and infection control. J Intern Med. 2015;277(5):501-12.
• Iraz M, Özad Düzgün A, Sandalli C, Doymaz MZ, Akkoyunlu Y, Saral A, et al. Distribution of β-lactamase genes among carbapenem-resistant Klebsiella pneumoniae strains isolated from patients in Turkey. Ann Lab Med. 2015;35(6):595-601.
• Gergin B, Akpolat N, Özcan N, Alkan Bilik Ö. Determination of carbapenemase production by BD phoenix CPO method in Karbapenem resistant Klebsiella pneumoniae and Escherichia coli isolates. Sakarya Med J. 2022;12(2):273-82. Turkish.
• Aslan AT, Kırbaş E, Sancak B, Tanrıverdi ES, Otlu B, Gürsoy NC, et al. A retrospective observational cohort study of the clinical epidemiology of bloodstream infections due to carbapenem-resistant Klebsiella pneumoniae in an OXA-48 endemic setting. Int J Antimicrob Agents. 2022;59(4):106554.
• Sahin K, Tekin A, Ozdas S, Akin D, Yapislar H, Dilek AR, et al. Evaluation of carbapenem resistance using phenotypic and genotypic techniques in Enterobacteriaceae isolates. Ann Clin Microbiol Antimicrob. 2015;14:44.
• Gomez E, Urban C, Mariano N, Colon-Urban R, Eng RH, Huang DB, et al. Phenotypic and genotypic screening and clonal analysis of carbapenem-resistant Klebsiella pneumoniae at a single hospital. Microb Drug Resist. 2011;17(2):251-7.
• Doumith M, Ellington MJ, Livermore DM, Woodford N. Molecular mechanisms disrupting porin expression in ertapenem-resistant Klebsiella and Enterobacter spp. clinical isolates from the UK. J Antimicrob Chemother. 2009;63(4):659-67.
• Tekeli A, Dolapci İ, Evren E, Oguzman E, Karahan ZC. Characterization of Klebsiella pneumoniae coproducing KPC and NDM-1 carbapenemases from Turkey. Microb Drug Resist. 2020;26(2):118-25.
• Nordmann P, Gniadkowski M, Giske CG, Poirel L, Woodford N, Miriagou V, et al. Identification and screening of carbapenemase-producing Enterobacteriaceae. Clin Microbiol Infect. 2012;18(5):432-8.
• Walther-Rasmussen J, Høiby N. OXA-type carbapenemases. J Antimicrob Chemother. 2006;57(3):373-83.
• Bercot B, Poirel L, Nordmann P. Updated multiplex polymerase chain reaction for detection of 16S rRNA methylases: High prevalence among NDM-1 producers. Diagn Microbiol Infect Dis. 2011;71(4):442-5.
• Giske CG, Martinez-Martinez L, Cantón R, Stefani S, Skov R, Glupczynski Y, et al. editors. EUCAST guideline for the detection of resistance mechanisms and specific resistances of clinical and/or epidemiological importance. 2nd ed. Sweden: European Committee on Antimicrobial Susceptibility Testing; 2017.