DOI: https://doi.org/10.22263/2312-4156.2019.5.69
Ziamko V.Y.1, Okulich V.K.1, Dzyadzko A.M.2, Kolchanova N.E.1, Kornilov A.V.1, Senkovich S.A.1
The role of DNA content in the composition of microbial biofilm in the pathogenesis of severe respiratory infections and the disease outcome modelling
1Vitebsk State Order of Peoples’ Friendship Medical University, Vitebsk, Republic of Belarus
2Minsk Scientific and Practical Center of Surgery, Transplantation and Hematology, Minsk, Republic of Belarus
Vestnik VGMU. 2019;18(5):69-76.
Abstract.
Objectives. To develop a method for determining deoxyribonucleic acid (DNA) percentage in the biofilm composition and to ascertain its significance in the pathogenesis of severe respiratory infections and their outcome modelling.
Material and methods. A total of 238 isolates taken from the sputum and pharynx of 175 patients treated at Vitebsk Regional Clinical Hospital and outpatient clinics during 2016-2019 were studied. Patients were divided into two groups: the 1st group consisting of 139 people (79.4%) had severe respiratory infections, the 2nd group composed of 36 persons (20.6%) suffered from respiratory infections of moderate severity.
Results. The method for determining DNA percentage in the composition of microbial community using 4’6-diamidino-2-phenylindole dihydrochloride was developed. The average age of the 1st group was higher than that of the second group (p<0.05). P. aeruginosa had the largest mass of the biofilm formed and DNA percentage in group 1, making up 48.25 [30.5-70.1] µg/ml and 5.21 [2.17-7.67] %, p=0.04. A strong relationship was found between DNA percentage in P. aeruginosa biofilm and the disease severity, r=0.73, and a weak one – in Acinetobacter spp., r=0.35, p<0.05. The incidence of adverse outcomes in case of antibiotic-resistant isolates was higher than that in case of antibiotic-sensitive ones (p<0.05). The analysis of the results made it possible to prognosticate to a high degree probability fatal outcome when the microbial biofilm mass was >47.5 μg/well and DNA percentage was >2.33%, (р<0,01).
Conclusions. The method for determining DNA percentage in the microbial community has been proposed allowing to judge the aggressiveness of the microorganism and the severity of the infection caused by it, which enables the providing of a personalized approach to the choice of antibiotic therapy based on the ability of the drug to penetrate through the biofilm.
Key words: antibiotic resistance, biofilm, deoxyribonucleic acid, 4’6-diamidino-2-phenylindole dihydrochloride.
The investigation was conducted within the frames of the scientific research work theme «Systemic inflammatory reaction in severe bacterial pneumonia: molecular-genetic mechanisms of the pathogenesis and their prognostic role in the evaluation of the antibiotic therapy efficacy», agreement with Belarusian Republican Fund of Fundamental Research № М19М-028 dated 02.05.2019
References
1. Lyamin AV, Botkin EA, Zhestkov AV. Methods for identifying biofilms in medicine: opportunities and prospects. Klin Mikrobiologiia Antimikrob Khimioterapiia. 2012;14(1):17-22. (In Russ.)
2. Tets VV, Tets GV. Microbial biofilms and problems of antibiotic therapy. Prakt Pul’monologiia. 2013;(4):60-4. (In Russ.)
3. Chebotar’ IV, Mayanskiy AN, Konchakova ED, Lazareva AV, Chistyakova VP. Antibiotic resistance of biofilm bacteria. Klin Mikrobiologiia Antimikrob Khimioterapiia. 2012;14(1):51-8. (In Russ.)
4. Das T, Sharma PK, Busscher HJ, van der Mei HC, Krom BP. Role of extracellular DNA in initial bacterial adhesion and surface aggregation. Appl Environ Microbiol. 2010 May;76(10):3405-8. doi: http://dx.doi.org/10.1128/AEM.03119-09
5. Goeres DM, Loetterle LR, Hamilton MA, Murga R, Kirby DW, Donlan RM. Statistical assessment of a laboratory method for growing biofilms. Microbiology. 2005 Mar;151(Pt 3):757-62. doi: http://dx.doi.org/10.1099/mic.0.27709-0
6. Ormerod MG, Tribukait B, Giaretti W. Consensus report of the task force on standardisation of DNA flow cytometry in clinical pathology. Anal Cell Pathol. 1998;17(2):103-10.
7. Sackoet M, Delacotte N, Bah M, Conti M, inventors; No voste Corporation, assignee. Method for assaying nucleic acids by fluorescence. WO patent 2008017948 A2, GOlN 21164, C12Q 1/68. 2008 Feb 14.
8. Smith RA, M’ikanatha NM, Read AF. Antibiotic Resistance: a primer and call to action. Health Commun. 2015;30(3):309-14. doi: http://dx.doi.org/10.1080/10410236.2014.943634
9. Høiby N, Bjarnsholt T, Givskov M, Molin S, Ciofu O. Antibiotic resistance of bacterial biofilms. Int J Antimicrob Agents. 2010 Apr;35(4):322-32. doi: http://dx.doi.org/10.1016/j.ijantimicag.2009.12.011
10. Microbiological methods for the study of biological material: instruktsiia po primeneniiu № 075-0210: utv M-vom zdravookhraneniia Resp Belarus’ 13 marta 2010 g. Minsk, RB; 2010. 75 р. (In Russ.)
11. The European Committee on Antimicrobial Susceptibility Testing. Breakpoint tables for interpretation of MICs and zone diameters. Version 9.0, valid from 2019-01-01. Available from: http://www.eucast.org/clinical_breakpoints/. [Accessed 08th Oct 2019].
12. Okulich VK, Kabanova AA, Plotnikov FV. Microbial biofilms in clinical microbiology and antibiotic therapy. Vitebsk, RB: VGMU; 2017. 300 р. (In Russ.)
13. Okulich VK, Kolchanova NE, Kornilov AV, Zemko VYu; zaiavitel’ i patentoobladatel’ Viteb gos med un-t. The method for determining the percentage of DNA in the composition of the microbial biofilm: pat Resp Belarus’, MKP S 12Q 1/04. № a 20180266; zaiavl 18.06.18; opubl 06.08.2018. (In Russ.)
Information about authors:
Ziamko V.Y. – postgraduate of the Chair of Anesthesiology and Resuscitation with the course of the Faculty for Advanced Training & Retraining, Vitebsk State Order of Peoples’ Friendship Medical University,
ORCID: https://orcid.org/0000-0002-6753-2074
Okulich V.K. – Candidate of Medical Sciences, associate professor of the Chair of Clinical Microbiology, Vitebsk State Order of Peoples’ Friendship Medical University,
ORCID: https://orcid.org/0000-0002-8226-6405
Dzyadzko A.M. – Doctor of Medical Sciences, head of the Department of Anesthesiology and Resuscitation, Minsk Scientific and Practical Center of Surgery, Transplantation and Hematology,
ORCID: https://orcid.org/0000-0003-1965-1850
Kolchanova N.E. – Candidate of Medical Sciences, associate professor of the Chair of Restorative Dentistry with the course of the Faculty for Advanced Training & Retraining, Vitebsk State Order of Peoples’ Friendship Medical University;
Kornilov A.V. – postgraduate of the Chair of Surgery of the Faculty for Advanced Training & Retraining, Vitebsk State Order of Peoples’ Friendship Medical University,
ORCID: https://orcid.org/0000-0001-8709-6639
Senkovich S.A. – Candidate of Medical Sciences, associate professor of the Chair of Clinical Microbiology, Vitebsk State Order of Peoples’ Friendship Medical University.
Correspondence address: Republic of Belarus, 210009, Vitebsk, 27 Frunze ave., Vitebsk State Order of Peoples’ Friendship Medical University, Chair of Anesthesiology and Resuscitation with the course of the Faculty for Advanced Training & Retraining. E-mail: Этот адрес электронной почты защищён от спам-ботов. У вас должен быть включен JavaScript для просмотра. – Viktoriya Y. Ziamko.
