OPTIMIZATION OF A METHOD OF ISOTHERMAL AMPLIFICATION (LAMP) FOR DIAGNOSIS OF WHOOPING COUGH

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Aim. Optimization of the accelerated whooping cough method of isothermal amplification for DNA Bordetela pertussis. Materials and methods. The research was conducted on 35 standard collection strains and 169 strains of Bordetella allocated in bacteriological laboratories of territorial subjects of the Russian Federation. The research included 329 clinical samples received from patients with whooping cough and the persons, contact with them, hospitalized in IDCH No. 1 DZM. Chromosomal DNA was extracted with a standard method of boiling from strains, from clinical samples by means of commercial sets. Identification of causative agents of whooping cough were performed with use of the АмплиСенс® Bordetella multi-FL. Results. We performed optimization method of a diagnostics of whooping cough by LAMP with detection by means of an electrophoresis and with naked-eye inspection under normal light is developed. The developed method allows to detect a DNA of B.pertussis within 4 - 5 hours in clinical material. The analytical sensitivity was 102 GE/ml. Assessment of validity showed that the developed method possesses 99,6% sensitivity and 98,7% specificity; predictive value positiveness and negative result was 99,6% and 98,7%, respectively; the index of accuracy (diagnostic efficiency) - 99,4%; the likelihood ratio of positive and negative result - 76,6 and 0,004, respectively. Assessment of analytical reliability in 100% of cases showed convergence and reproducibility of a technique. Conclusion. Diagnostic test on DNA of B.pertussis identification by LAMP method will allow to increase efficiency of laboratory diagnosis of whooping cough.

About the authors

A. S. Pimenova

Gabrichevsky Moscow Research Institute of Epidemiology and Microbiology

Author for correspondence.
Email: noemail@neicon.ru
Russian Federation

O. Yu. Borisova

Gabrichevsky Moscow Research Institute of Epidemiology and Microbiology, Pirogov Russian National Research Medical University

Email: noemail@neicon.ru
Russian Federation

M. S. Petrova

Gabrichevsky Moscow Research Institute of Epidemiology and Microbiology

Email: noemail@neicon.ru
Russian Federation

E. V. Vlasov

Infectious Diseases Clinical Hospital No. 1 of the Moscow Department of Healthcare

Email: noemail@neicon.ru
Russian Federation

I. S. Voronina

Gabrichevsky Moscow Research Institute of Epidemiology and Microbiology

Email: noemail@neicon.ru
Russian Federation

A. B. Borisova

Pirogov Russian National Research Medical University

Email: noemail@neicon.ru
Russian Federation

S. S. Afanasiev

Gabrichevsky Moscow Research Institute of Epidemiology and Microbiology

Email: noemail@neicon.ru
Russian Federation

E. E. Donskich

Pirogov Russian National Research Medical University

Email: noemail@neicon.ru
Russian Federation

L. I. Kafarskaya

Pirogov Russian National Research Medical University

Email: noemail@neicon.ru
Russian Federation

V. A. Aleshkin

Gabrichevsky Moscow Research Institute of Epidemiology and Microbiology

Email: noemail@neicon.ru
Russian Federation

A. V. Aleshkin

Gabrichevsky Moscow Research Institute of Epidemiology and Microbiology

Email: noemail@neicon.ru
Russian Federation

M. S. Afanasiev

Sechenov First Moscow State Medical University

Email: noemail@neicon.ru
Russian Federation

A. V. Karaulov

Sechenov First Moscow State Medical University

Email: noemail@neicon.ru
Russian Federation

References

  1. Бабаченко И.В., Харит С.М., Курова Н.Н. и др. Коклюш у детей: монография. М., Комментарий, 2014.
  2. Басов А.А., Цвиркун О.В., Герасимова А.Г. и др. Особенности распространения коклюша в организованном коллективе с высоким уровнем привитости против этой инфекции. Жизнь без опасностей. Здоровье. Профилактика. Долголетие. 2013, 8(4): 60-64.
  3. Борисова О.Ю., Петрова М.С., Гадуа Н.Т. и др. Прямой ускоренный метод выявления возбудителя коклюша. Клиническая лабораторная диагностика. 2010, (5): 53-55.
  4. ГОСТ Р 53022.3-2008. Требования к качеству клинических лабораторных исследований. Часть 3. Правила оценки клинической информативности лабораторных тестов. Введ. 2010-01-01. М.: Стандартинформ, 2009.
  5. Каратаев Г.И., Синяшина Л.Н., Медкова А.Ю. и др. Инсерционная инактивация оперона вирулентности в популяции персистирующих бактерий Bordetella pertussis. Генетика. 2016, 52(4): 422.
  6. Медкова А.Ю., Аляпкина Ю.С., Синяшина Л.Н. и др. Распространенность стертых форм коклюша и анализ фазовых состояний бактерий Bordetella pertussis. Детские инфекции. 2010, 9(4): 19-22.
  7. Онищенко Г.Г. Эпидемиологическое благополучие населения России. Журн. микробиол. 2013, (1): 42-51.
  8. Петри А., Сэбин К. Наглядная медицинская статистика: учебное пособие для вузов. М., ГЭОТАР-Медиа, 2010.
  9. Петрова М.С., Попова О.П., Борисова О.Ю. и др. Коклюш у детей раннего возраста. Эпидемиология и инфекционные болезни. 2012, (6): 12-24.
  10. Платонов А.Е. Статистический анализ в медицине и биологии: задачи, терминология, логика, компьютерные методы. М., РАМН, 2000.
  11. Прадед М.Н., Яцышина С.Б., Селезнева Т.С. и др. ПЦР-диагностика инфекций, вызванных B.pertussis, B.parapertussis и B.bronchiseptica. Клиническая лабораторная диагностика. 2013, (1): 53-56.
  12. Brotons P., de Paz H.D., Esteva C. et al. Validation of a loop-mediated isothermal amplification assay for rapid diagnosis of pertussis infection in nasopharyngeal samples. Expert Rev. Mol. Diagn. 2016, 16(1): 125-130. doi: 10.1586/14737159.2016.1112741.
  13. Douglas E., Coote J.G., Parton R. et al. Identification of Bordetella pertussis in nasopharyngeal swabs by PCR amplification of a region of the adenylate cyclase gene. J. Med. Microbiol. 1993, 38(2): 140-144. doi: 10.1099/00222615-38-2-140.
  14. Grimprel E.P., Begue P., Anjak I. et al. Comparison of polymerase chain reaction, culture and Western immunoblot serology for diagnosis of Bordetella pertussis infections. J. Clin. Microbiol. 1993, 31(10): 2745-2750.
  15. Kamachi K., Toyoizumi-Ajisaka H., Toda K. et al. Development and evaluation of a loop-mediated isothermal amplification method for rapid diagnosis of Bordetella pertussis infection. J. Clin. Microbiol. 2006, 44(5): 1899-1902. doi: 10.1128/JCM.44.5.1899-1902.2006.
  16. Kamachi K., Yoshino S., Katsukawa C. et al. Laboratory-based surveillance of pertussis using multitarget real-time PCR in Japan: evidence for Bordetella pertussis infection in preteens and teens. New Microbes New Infect. 2015, (8): 70-74. doi: 10.1016/j.nmni.2015.10.001.
  17. Lanotte Ph., Plouzeau C, Burucoa C. et al. Evaluation of four commercial Real-Time PCR assays for detection of Bordetella spp. in nasopharyngeal aspirates. J. Clin. Microbiol. 2011, 49(11): 3943-3946. doi: 10.1128/JCM.00335-11.
  18. Litt D.J., Jauneikaite E., Tchipeva D. et al. Direct molecular typing of Bordetella pertussis from clinical specimens submitted for diagnostic quantitative (real-time) PCR. J. Med. Microbiol. 2012, 61(12): 1662-1668. doi: 10.1099/jmm.0.049585-0.
  19. Loeffelholz M. Towards improved accuracy of Bordetella pertussis nucleic acid amplification tests. J. Clin. Microbiol. 2012, 50(7): 2186-2190. doi: 10.1128/JCM.00612-12.
  20. Notomi T., Okayama H., Masubuchi H. et al. Loop-mediated isothermal amplification of DNA. Nucleic Acids Res. 2000, 28(12): E63.
  21. Notomi T., Mori Y., Tomita N. et al. Loop-mediated isothermal amplification (LAMP): principle, features, and future prospects. J. Microbiol. 2015, 53(1): 1-5. doi: 10.1007/s12275-015-4656-9.
  22. Qin X., Galanakis E., Martin E.T. et al. Multitarget PCR for diagnosis of pertussis and its clinical implications. J. Clin. Microbiol. 2007, 45(2): 506-511. doi: 10.1128/JCM.02042-06.
  23. Qin X. Resurgence of Pertussis апс1 its Moratory diagnоsis. Clin. Microbiol. Newsletter. 2015, 37(9): 69-76. doi: 10.1016/j.clinmicnews.2015.04.001.
  24. Stone B.L., Daly J., Srivastava R. Duration of Bordetelta pertussis polymerase duiin reactiоn positivity in confirmed pertussis illness. J. Pediatric Infect.Dis. Soc. 2014, 3(4): 347-349. doi: 10.1093/jpids/piu004.

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Copyright (c) 2018 Pimenova A.S., Borisova O.Y., Petrova M.S., Vlasov E.V., Voronina I.S., Borisova A.B., Afanasiev S.S., Donskich E.E., Kafarskaya L.I., Aleshkin V.A., Aleshkin A.V., Afanasiev M.S., Karaulov A.V.

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