Stomatologist № 3 (38) – 2020, pp. 8-17                                                                                                   SCIENTIFIC PUBLICATION

Preparatory measures in laboratory express diagnostics of the state of periodontal tissues

L.N. Dedovaa, P.A. Semizhonb, О.V. Каndrukevichc, E.P. Scheslenokb, E.Y. Burdeikod, S.V. Poddubnovd

aMD, PhD, DMSci, Professor, Belarusian State Medical University, Minsk, Belarus
bPhD, The Republican Research and Practical Center for Epidemiology and Microbiology, Minsk, Belarus
cMD, PhD, Associate Professor, Belarusian State Medical University, Minsk, Belarus
dThe Republican Research and Practical Center for Epidemiology and Microbiology, Minsk, Belarus

https://doi.org/10.32993/dentist.2020.3(38).10

ABSTRACT
Effective diagnosis of periodontal diseases includes the ability to determine the quantitative and qualitative composition of the oral microflora.
The aim of the study. Determine the order of preparatory measures in laboratory diagnostics of the state of periodontal tissues using PCR and assessment of trypsin-like activity of biological samples.
Objects and methods. The objects of study were the nucleotide sequences of the genomes of Treponema denticola, Porphyromonas gingivalis, Bacteroides forsythus; persons who have applied for dental care, samples of biological material (contents of pathological periodontal pocket – PPP) from patients with a clinically established diagnosis. The following research methods were used in the work: dental examination of patients, PCR diagnostics method, assessment of trypsin-like activity of pathogenic microflora of the periodontium.
Results and discussion. An algorithm for laboratory diagnosis of periodontal diseases has been developed, including the sampling of material and the detection of periopathogenic T. denticola, P. gingivalis, B. forsythus in the sample using the biochemical method; sampling of material, its conservation and keeping for research by quantitative PCR; identification of the genetic material of periopathogenic T. denticola, P. gingivalis, B. forsythus by quantitative PCR in “real time”.
Conclusion. The previously proposed method of collection, keeping and preparation of biological samples from PPP in patients with chronic generalized periodontitis for laboratory diagnostics has shown its appropriateness. BANA (N-α-benzoyl-DL-arginine-2-naphthylamide) is the most optimal and effective among other synthetic substrates in reactions to study the trypsin-like activity of biological samples. An algorithm for sample preparation and laboratory diagnosis of the state of periodontal tissues in patients with periodontitis was developed using the method PCR and a domestic express method based on the assessment of the trypsin-like activity of biological samples to conduct an effective rapid study in dental practice.

Keywords: laboratory diagnostics periodontology, microbiology

References 

  1. Teles R.P., Haffajee A.D., Socransky S.S. Microbiological goals of periodontal therapy. Periodontol. 2000. 2006, vol. 42, pp. 180–218.
  2. Ellen R.P., Galimanas V.B. Spirochetes at the forefront of periodontal infections. Periodontol. 2000. 2005, vol. 38, pp. 13–32.
  3. Dedova L.N. [et al.] Terapevticheskaja stomatologija. Bolezni periodonta: ucheb. posobie pod red. L.N. Dedovoj [Therapeutic dentistry. Periodontal disease: textbook. manual, ed. L.N. Dedova]. Minsk: Jekoperspektiva, 2016. 268 p.
  4. König J., Holtfreter B., Kocher T. Periodontal health in Europe: future trends based on treatment needs and the provision of periodontal services – position paper 1. Eur. J. Dent. Educ. 2010, vol. 14, pp. 4–24.
  5. Dedova L.N., Rubnikovich S.P., Denisova Ju.L., Kandrukevich O.V., Solomevich A.S., Rosenik N.I. Rasprostranennost’ stomatologicheskih zabolevanij v Respublike Belarus’ [Prevalence of dental diseases in the Republic of Belarus]. Stomatologija. Jestetika. Innovacii. – Dentistry. Aesthetics. Innovation. 2017, no. 2, pp. 193–202.
  6. Dedova L.N. [et al.] Sostojanie tkanej periodonta i kariesa poverhnosti kornja po dannym jepidemiologicheskogo obsledovanija 35–54-letnih zhitelej Respubliki Belarus’ [Condition of periodontal tissues and caries of the root surface according to the data of an epidemiological survey of 35–54-year-old residents of the Republic of Belarus]. Stomatologicheskij zhurnal. – Dental journal. 2006, no. 4, pp. 322–323.
  7. Kinane D.F., Stathopoulou P.G., Papapanou P.N. Periodontal diseases. Nat. Rev. Dis. Primers. 2017, vol. 22, no. 3, pp. 1–14.
  8. Suchett-Kaye G., Morrier J.J., Barsotti O. Clinical usefulness of microbiological diagnostic tools in the management of periodontal disease. Res. Microbiol. 2001, vol. 152, pp. 631–639.
  9. Loesche W.J., Syed S.A., Stoll J. Trypsin-like activity in subgingival plaque. A diagnostic marker for spirochetes and periodontal disease? J. Periodontol. 1987, vol. 58, no. 4, pp. 266–273.
  10. Ohta K., Makinen K.K., Loesche W.J. Purification and characterization of an enzyme produced by Treponema denticolacapable of hydrolyzing synthetic trypsin substrates. Infect. Immunity. 1986, vol. 53, no. 1, pp. 213–220.
  11. Tsutsui H. [et al.] Purification and characterization of a protease from Bacteroides gingivalis 381. Infect. Immunity. 1987, vol. 55, no. 2, pp. 420–427.
  12. Yoshimura F. [et al.] Characterization of a trypsin-like protease from the bacterium bacteroides gingivalis isolated from human dental plaque. Arch. Oral. Biol. 1984, vol. 29, no. 7, pp. 559–564.
  13. Hockensmith K. [et al.] Identification and characterization of a chymotrypsin-like serine protease from periodontal pathogen, Tannerella forsythia. Microb. Pathog. 2016, vol. 100, pp. 37–42.
  14. Loesche W.J. [et al.] Development of a diagnostic test for anaerobic periodontal infections based on plaque hydrolysis of benzoyl-DLarginine-naphthylamide. J. Clin. Microbiol. 1990, vol. 28, no. 7, pp. 1551–1559.
  15. Loesche W.J., Kazor C.E., Taylor G.W. The optimization of the BANA test as a screening instrument for gingivitis among subjects seeking dental treatment. J. Clin. Periodontol. 1997, vol. 24, pp. 718–726.
  16. Loesche W.J. [et al.] Comparison of the benzoyl-DL-arginine-naphthylamide (BANA) test, DNA probes, and immunological reagents for ability to detect anaerobic periodontal infections due to Porphyromonas gingivalis, Treponema denticola, and Bacteroides forsythus. J. Clin. Microbiol. 1992, vol. 30, no. 2, pp. 427–433.
  17. Bretz W.A. [et al.] The use of a rapid enzymatic assay in the field for the detection of infections associated with adult periodontitis. J. Public Health Dentistry. 1993, vol. 53, no. 4, pp. 235–240.
  18. Grisi M.F. [et al.] Correlation between the CPITN score and anaerobic periodontal infections assessed by BANA assay. Braz. Dent. J. 1999, vol. 10, pp. 93–97.
  19. Andrade J.A. [et al.] The ability of the BANA test to detect Porphyromonas gingivalis, Treponema denticola and Tannerella forsythia. Braz. Oral. Res. 2010, vol. 24, no. 2, pp. 224–230.
  20. Haffajee A.D., Socransky S.S. Microbial etiological agents of destructive periodontal disease. Periodontol. 2000. 1994, vol. 5, pp. 78–111.
  21. Loesche W.J., Giordano J., Hujoel P.P. The utility of the BANA test for monitoring anaerobic infections due to spirochetes (Treponema denticola) in periodontal disease. J. Dent. Res. 1990, vol. 69, pp. 1696–1990.
  22. Loesche W.J. [et al.] A logistic regression model for the decision to perform access surgery. J. Clin. Periodontol. 1997, vol. 24, pp. 171–179.
  23. Loesche W.J. [et al.] The non-surgical treatment of periodontal patients. Oral. Med. Oral. Surg. Oral. Path. 1996, vol. 81, pp. 533–544.
  24. Cortelli J.R., Barbosa M.D.S., Westphal M.A. Halitosis: a review of associated factors and therapeutic approach. Braz. Oral. Res. 2008, vol. 22, pp. 44–54.
  25. De Boever, E.H. Assessing the contribution of anaerobic microflora of the tongue to oral malodor / E.H. De Boever, W.J. Loesche // J. Am. Dent. Assoc. – 1995. – Vol. 126. – P. 1384–1393.
  26. Mattila K.J. [et al.] Dental infection and the risk of new coronary events: prospective study of patients with documented coronary artery disease. Clin. Infect. Dis. 1995, vol. 20, no. 3, pp. 588–592.
  27. Loesche W.J. [et al.] Assessing the relationship between dental disease and coronary heart disease in elderly U.S. veterans. J. Am. Dent. Assoc. 1998, vol. 129, no. 3, pp. 301–311.
  28. Desvarieux M. [et al.] Periodontal microbiota and carotid intima-media thickness. Circulation. 2005, vol. 111, pp. 576–582.
  29. Bretz W.A. [et al.] Systemic inflammatory disease markers, periodontal disease and periodontal infections in an elderly population. J. Am. Geritric Soc. 2005, vol. 53, no. 9, pp. 1532–1537.
  30. Seinost G. [et al.] Periodontal treatment improves endothelial dysfunction in patients with severe periodontitis. Am. Heart J. 2005, vol. 149, no. 6, pp. 1050–1054.
  31. Goldenberg R.L. [et al.] Epidemiology and causes of preterm birth. Lancet. 2008, vol. 371, pp. 75–84.
  32. Chan H.C. [et al.] Periodontal disease activity measured by the benzoyl-DL-arginine-naphthylamide test is associated with preterm births. J. Periodontol. 2010, vol. 81, pp. 982–991.
  33. Turton M.S., Henkel R. R., Africa C.W.J. A simple point of care test can indicate the need for periodontal therapy to reduce the risk for adverse pregnancy outcomes in mothers attending antenatal clinics. Biomarkers. 2017, vol. 22, no. 8, pp. 740–746.
  34. Jeffcoat M. [et al.] Periodontal infection and preterm birth: successful periodontal therapy reduces the risk of preterm birth. BJOG. 2011, vol. 118, no. 2, pp. 250–256.
  35. Dedova L.N. [et al.] Osobennosti kliniko-laboratornoj diagnostiki v periodontologii [Features of clinical and laboratory diagnostics in periodontology]. Stomatolog. Minsk. – Stomatologist. Minsk. 2019, no. 3, pp. 22–28.

Correspondence to:  Е-mail: Dedova.bsmu@mail.ru