Comparative effectiveness of molecular tests based on isothermal amplification and polymerase chain reaction for detection carbapenemase genes in hospital-acquired pneumonia

Мұқаба


Дәйексөз келтіру

Толық мәтін

Ашық рұқсат Ашық рұқсат
Рұқсат жабық Рұқсат берілді
Рұқсат жабық Рұқсат ақылы немесе тек жазылушылар үшін

Аннотация

 

BACKGROUND: Antibiotic resistance in pathogens causing hospital-acquired pneumonia (HAP), particularly to carbapenems non-susceptibility, is a serious public health problem. New rapid and sensitive diagnostic methods are required for the timely initiation of effective antimicrobial therapy. The development and implementation of molecular diagnostic tests, based on polymerase chain reaction and promising isothermal amplification techniques, will significantly reduce turnaround time and improve detection of HAP pathogens with carbapenemase genes.

AIM: The aim was to compare efficacy of a new loop-mediated isothermal amplification (LAMP) test for the detection of NDM, OXA-48, and KPC carbapenemase genes and routine laboratory tests based on real-time polymerase chain reaction in the diagnosis of hospital-acquired lower respiratory tract infections (LRTI), including hospital-acquired pneumonia.

MATERIALS AND METHODS: The study included 141 samples of lower respiratory tract biomaterial collected from 107 patients with hospital-acquired LRTI (hospital-acquired pneumonia and/or purulent tracheobronchitis). Tracheal aspirate (TA, n=78), bronchoalveolar lavage (BAL, n=29) and sputum (n=34) were used as biomaterial samples. Nucleic acids were isolated using an AmpliTest® RIBO-prep kit. NDM, OXA-48 and KPC genes were analyzed using the proposed AmpliTest® CP NDM/OXA-48/KPC LAMP reagent kit by loop isothermal amplification (LAMP). To compare results, real-time PCR was performed using the AmpliSens MDR MBL-FL and AmpliSens MDR KPC/OXA-48-FL reagent kits.

RESULTS: Concordant results were obtained for the detection of NDM, OXA-48 and KPC carbapenemase genes in lower respiratory tract biomaterial samples using a new LAMP test and PCR tests for all samples tested. Based on the time to positivity (TTP), the turnaround time for LAMP was shorter than the turnaround time for PCR. Additional PCR tests were used to identify the Gram-negative microorganisms in the samples tested (both individually and in combination).

CONCLUSION: The obtained data showed that the efficiency of detection of the study groups of carbapenemase genes by the new LAMP test is not lower than the efficiency of their detection by the PCR tests previously used in practice.

Толық мәтін

Рұқсат жабық

Авторлар туралы

Danilov Dmitry Igorevich Danilov

Centre for Strategic Planning and Management of Biomedical Health Risks

Хат алмасуға жауапты Автор.
Email: danilov.i.dmitry@gmail.com
ORCID iD: 0000-0001-5772-8498
SPIN-код: 8372-8456
Ресей, 10 bldg. 1 Pogodinskaya street, 119121 Moscow

Elizaveta Glushchenko

Centre for Strategic Planning and Management of Biomedical Health Risks

Email: Glushhenko@cspfmba.ru
ORCID iD: 0000-0002-6674-2045
SPIN-код: 7440-4930
Ресей, 10 bldg. 1 Pogodinskaya street, 119121 Moscow

Yulia Savochkina

Centre for Strategic Planning and Management of Biomedical Health Risks

Email: YSavochkina@cspmz.ru
ORCID iD: 0000-0003-2313-0521

Cand. Sci. (Biology)

Ресей, 10 bldg. 1 Pogodinskaya street, 119121 Moscow

Daria Strelkova

Sechenov First Moscow State Medical University (Sechenov University)

Email: dashastrelkova@gmail.com
ORCID iD: 0000-0002-2124-0623
SPIN-код: 9549-8053

MD, Cand. Sci. (Medicine)

Ресей, Moscow

Svetlana Rachina

Sechenov First Moscow State Medical University (Sechenov University)

Email: rachina_s_a@staff.sechenov.ru
ORCID iD: 0000-0002-3329-7846
SPIN-код: 1075-7329

MD, Dr. Sci. (Medicine), Assistant Professor

Ресей, Moscow

Diana Gasanova

Sechenov First Moscow State Medical University (Sechenov University)

Email: gasanova_556@mail.com
ORCID iD: 0009-0008-9830-7516
Ресей, Moscow

Lyudmila Fedina

Moscow City Hospital named after S.S. Yudin

Email: fedina201368@gmail.com
ORCID iD: 0000-0002-6417-9535
SPIN-код: 1961-7486
Ресей, Moscow

Igor Sychev

Moscow City Hospital named after S.S. Yudin

Email: sychevigor@mail.ru
ORCID iD: 0000-0002-2970-3442
SPIN-код: 7282-6014

MD, Cand. Sci. (Medicine), Assistant Professor

Ресей, Moscow

Egor Larin

Hospital for War Veterans No. 3

Email: dr.egorlarin@gmail.com
ORCID iD: 0000-0002-7450-6317
Ресей, Moscow

German Shipulin

Centre for Strategic Planning and Management of Biomedical Health Risks

Email: Shipulin@cspfmba.ru
ORCID iD: 0000-0002-3668-6601
SPIN-код: 1908-9098

MD, Cand. Sci. (Medicine)

Ресей, 10 bldg. 1 Pogodinskaya street, 119121 Moscow

Әдебиет тізімі

  1. Xu E, Pérez-Torres D, Fragkou PC, Zahar JR, Koulenti D. Nosocomial pneumonia in the era of multidrug-resistance: Updates in diagnosis and management. Microorganisms. 2021;9(3):534. doi: 10.3390/microorganisms9030534
  2. Ramadan RA, Bedawy AM, Negm EM, et al. Carbapenem-resistant Klebsiella pneumoniae among patients with ventilator-associated pneumonia: Evaluation of antibiotic combinations and susceptibility to new antibiotics. Infect Drug Resist. doi: 10.2147/IDR.S371248
  3. Doi Y. Treatment Options for Carbapenem-resistant Gram-negative Bacterial Infections. Clin Infect Dis. 2019;69:S565–S575. doi: 10.1093/cid/ciz830
  4. Erfani Y, Rasti A, Janani L. Prevalence of Gram-negative bacteria in ventilator-associated pneumonia in neonatal intensive care units: a systematic review and meta-analysis protocol. BMJ Open. 2016;6(10):e012298. doi: 10.1136/bmjopen-2016-012298
  5. Kaboré B, Ouédraogo GA, Ouédraogo HS, et al. Identification and phylogenetic analysis of carbapenemase genes from clinical strains of Klebsiella pneumoniae. J Infect Dev Ctries. 2024;18(09):1387–1393. doi: 10.3855/jidc.17519
  6. Kuzmenkov AYu, Vinogradova AG, Trushin IV, et al. AMRmap – antibiotic resistance surveillance system in Russia. Clin Microbiol Antimicrob Chemother. 2021;23(2):198–204. (In Russ.) doi: 10.36488/cmac.2021.2.198-204
  7. Yakovlev S, Suvorova M, Bykov A. Infections Caused by Carbapenem-Resistant Enterobacterales: Epidemiology, Clinical Significance, and Possibilities for Antibiotic Therapy Optimization. Antibiot Chemother. 2020;65(5–6):41–69. (In Russ.) doi: 10.37489/0235-2990-2020-65-5-6-41-69
  8. Yakovlev S, Suvorova M, Beloborodov V, et al. Nosocomial infections in surgical wards in Russian emergency hospitals: ERGINI study. Infections in surgery. 2016;61(5–6):32–42. (In Russ.) EDN: OJZPKJ
  9. Brkic S, Cirkovic I. Carbapenem-Resistant Enterobacterales in the Western Balkans: Addressing Gaps in European AMR Surveillance Map. Antibiotics. 2024;13(9):895. doi: 10.3390/antibiotics13090895
  10. Piotrowski M, Alekseeva I, Arnet U, Yücel E. Insights into the Rising Threat of Carbapenem-Resistant Enterobacterales and Pseudomonas aeruginosa Epidemic Infections in Eastern Europe: A Systematic Literature Review. Antibiotics. 2024;13(10):978. doi: 10.3390/antibiotics13100978
  11. Rachina SА, Fedina LV, Sukhorukova MV, et al. Diagnosis and antibiotic therapy of nosocomial pneumonia in adults: from recommendations to real practice. A review. Ter Arkh. 2023;95(11):996–1003. (In Russ.) doi: 10.26442/00403660.2023.11.202467
  12. Beloborodov V, Gusarov V, Dekhnich A, et al. Guidelines of the Association of Anesthesiologists-Intensivists, the Interregional Non-Governmental Organization Alliance of Clinical Chemotherapists and Microbiologists, the Interregional Association for Clinical Microbiology and Antimicrobial Chemotherapy (IACMAC), and NGO Russian Sepsis Forum Diagnostics and antimicrobial therapy of the infections caused by multiresistant microorganisms. Messenger Anesthesiol Resusc. 2020;16:52–83. doi: 10.21292/2078-5658-2020-16-1-52-83
  13. Sukhorukova M, Edelstein M, Ivanchik N, et al. Antimicrobial resistance of nosocomial Enterobacterales isolates in Russia: results of multicenter epidemiological study “MARATHON 2015–2016”. Clinical Microbiology and Antimicrobial Chemotherapy. 2019;21(2):147–159. doi: 10.36488/cmac.2019.2.147-159
  14. Wu W, Feng Y, Tang G, et al. NDM Metallo-β-Lactamases and Their Bacterial Producers in Health Care Settings. Clin Microbiol Rev. 2019;32(2):10–1128. doi: 10.1128/CMR.00115-18
  15. Li X, Zhao D, Li W, Sun J, Zhang X. Enzyme Inhibitors: The Best Strategy to Tackle Superbug NDM-1 and Its Variants. Int J Mol Sci. 2021;23(1):197. doi: 10.3390/ijms23010197
  16. Pitout JD, Peirano G, Kock MM, Strydom KA, Matsumura Y. The Global Ascendency of OXA-48-Type Carbapenemases. Clin Microbiol Rev. 2019;33(1):10–1128. doi: 10.1128/CMR.00102-19
  17. Hirvonen VH, Spencer J, Van Der Kamp MW. Antimicrobial Resistance Conferred by OXA-48 β-Lactamases: Towards a Detailed Mechanistic Understanding. Antimicrob Agents Chemother. 2021;65(6):10–1128. doi: 10.1128/AAC.00184-21
  18. Yakovlev SV. Clinical Efficacy of Ceftazidime-Avibactam in the Treatment of Infections Caused by Carbapenem–Resistant Gram-Negative Bacteria. Antibiot Chemother. 2021;66(7–8):67–82. doi: 10.37489/0235-2990-2021-66-7-8-67-82
  19. Emelyanov VN, Zorya AI, Glushkov AA. Epidemiological features of antibiotic resistance of clinically significant pathogenic microorganisms using the example of bacteria of the genus Serratia. Medicine. 2024;12(3):118–129. doi: 10.29234/2308-9113-2024-12-3-118-129
  20. Boutal H, Vogel A, Bernabeu S, et al. A multiplex lateral flow immunoassay for the rapid identification of NDM-, KPC-, IMP-and VIM-type and OXA-48-like carbapenemase-producing Enterobacteriaceae. J Antimicrob Chemother. 2018;73(4):909–915. doi: 10.1093/jac/dkx521
  21. Sękowska A, Bogiel T. The Evaluation of Eazyplex® SuperBug CRE Assay Usefulness for the Detection of ESBLs and Carbapenemases Genes Directly from Urine Samples and Positive Blood Cultures. Antibiotics. 2022;11(2):138. doi: 10.3390/antibiotics11020138
  22. Isler B, Aslan AT, Akova M, Harris P, Paterson DL. Treatment strategies for OXA-48-like and NDM producing Klebsiella pneumoniae infections. Expert Rev Anti Infect Ther. 2022;20(11):1389–1400. doi: 10.1080/14787210.2022.2128764
  23. Suvorova MP, Sychev IN, Ignatenko OV, et al. The First Experience of Combined Use of Cefepime/Sulbactam and Aztreonam in ICU Patients with Nosocomial Infections Caused by Carbapenem-Resistant Gram-Negative Microorganisms Producing Class B and D Carbapenemases. Antibiot Chemother. 2023;67(11–12):36–45. doi: 10.37489/0235-2990-2022-67-11-12-36-45
  24. Porreca AM, Sullivan KV, Gallagher JC. The epidemiology, evolution, and treatment of KPC-producing organisms. Curr Infect Dis Rep. 2018;20(13):1–12. doi: 10.1007/s11908-018-0617-x
  25. Gu D, Yan Z, Cai C, et al. Comparison of the NG-Test Carba 5, Colloidal Gold Immunoassay (CGI) Test, and Xpert Carba-R for the Rapid Detection of Carbapenemases in Carbapenemase-Producing Organisms. Antibiotics. 2023;12(2):300. doi: 10.3390/antibiotics12020300
  26. Poirier AC, Kuang D, Siedler BS, et al. Development of Loop-Mediated Isothermal Amplification Rapid Diagnostic Assays for the Detection of Klebsiella pneumoniae and Carbapenemase Genes in Clinical Samples. Front Mol Biosci. 2022;8:794961. doi: 10.3389/fmolb.2021.794961
  27. Seymour CW, Gesten F, Prescott HC, et al. Time to Treatment and Mortality during Mandated Emergency Care for Sepsis. N Engl J Med. 2017;376(23):2235–2244. doi: 10.1056/NEJMoa1703058
  28. Shajdullina E, Edelstein M, Skleenova EY, Sukhorukova M, Kozlov R. Antimicrobal resistance of nosocomial carbapenemase-producing Enterobacterales in Russia: results of surveillance, 2014-2016. Clinical Microbiology and Antimicrobial Chemotherapy. 2018;20(4):362–369. doi: 10.36488/cmac.2018.4.362-369

Қосымша файлдар

Қосымша файлдар
Әрекет
1. JATS XML
Жүктеу
2. Fig. 1. Examples of fluorescence signal graphs for the detection of carbapenemase genes in control samples with target DNA concentrations of 1x10^5 copies/mL, 1x10^4 copies/mL, and 1x10^3 copies/mL, as well as the detection of the internal control sample: a) detection of MBL genes from the NDM group (signal in the HEX channel in the first reaction mixture); b) detection of IC DNA (signal in the ROX channel in the first mixture); c) detection of carbapenemase genes from the KPC group (signal in the HEX channel in the second reaction mixture); d) detection of carbapenemase genes from the OXA-48 group (signal in the ROX channel in the second reaction mixture).

Жүктеу (421KB)
Метадеректер
3. Fig. 2. TTP diagram for PCR-RT and LAMP methods in the analysis of lower respiratory tract clinical samples within the study: a) PCR-RT TTP for the OXA-48, NDM, and KPC target genes; b) LAMP TTP for the OXA-48, NDM, and KPC target genes.

Жүктеу (78KB)
Метадеректер

© Danilov D.I., Glushchenko E.E., Savochkina Y.A., Strelkova D.A., Rachina S.A., Gasanova D.R., Fedina L.V., Sychev I.N., Larin E.S., Shipulin G.A., 2025

Creative Commons License
Бұл мақала лицензия бойынша қол жетімді Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International License.
СМИ зарегистрировано Федеральной службой по надзору в сфере связи, информационных технологий и массовых коммуникаций (Роскомнадзор).
Регистрационный номер и дата принятия решения о регистрации СМИ:  ПИ № ФС 77 - 86785 от 05.02.2024 (ранее — ПИ № ФС 77 - 59057 от 22.08.2014).