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STREPTOCOCCUS MILLERI GRUP (SMG) BAKTERİLERİNİN BİYOKİMYASAL VE MOLEKÜLER YÖNTEMLERLE TANIMLANMASI

Year 2023, Volume: 6 Issue: 1, 73 - 79, 28.02.2023
https://doi.org/10.26650/JARHS2023-1164336

Abstract

Amaç: Streptococcus milleri grubu (SMG) bakterilerin, moleküler, biyokimyasal ve yarı otomatize tekniklerle tür ayırımının yapılması ve tedavide sık kullanılan antibiyotiklere karşı duyarlılık sonuçlarının araştırılması amaçlanmıştır. Bu çalışma, SMG bakteriler ile ilgili bazı önemli noktalara dikkat çekmek ve epidemiyolojik olarak önemini tespit etmek için planlanmıştır. Gereç ve Yöntem: Çalışmamıza 100 streptokok cinsi bakteri dahil edilmiştir. Tür tanımlaması için biyokimyasal testler ve API 20 STREP yarı otomatik identifikasyon kiti kullanılmıştır. Polimeraz zincir reaksiyonu (PZR) ile hedef gen bölgeleri (HYL-MIX-U, HLY-INT- D, HLY-CC-D, 16S-ANG-U, 16S-ANG-D, ILY-4DFw, ILY-wholeC Bw) çoğaltılarak tür tanımlamaları doğrulanmıştır. Bulgular: Tüm örneklerin Voges-Proskauer (VP) deneyi pozitif, hipurat hidrolizi ve pirolidonil arylamidaz (PYR) negatif olarak saptanmıştır. Tüm suşlar Lancefield grup antijeni açısından değerlendirilmiş olup, %42 G, %26 F, %5 ise C grup antijenine sahip olduğu bulunmuştur. Suşların %27’sinin ise Lancefield grup antijenleri ile antijen tipi saptanamamıştır. 100 suşun tamamı vankomisin, seftriakson, sefepim, sefotaksim, levofloksasin, linezolid antibiyotiklerine duyarlı olarak bulunmuştur. Suşlarda, %16 eritromisin, %6 klindamisin, %5 tetrasiklin, %1 kloramfenikol direnci saptanmıştır. Ampisilin ve penisilin direnci E-test yöntemi ile saptanmış ve %1 ampisilin/penisiline dirençli Streptococcus anginosus suşu, %2 ampisiline orta duyarlı S. anginosus suşu saptanmıştır. API 20 STREP sonuçlarına göre bakterilerin %41 cins bazında tanımlanmış, %45’i S. anginosus, %5’i Streptococcus constellatus, %1’i Streptococcus intermedius olarak sonuçlandırılmıştır. Multipleks polimeraz zincir reaksiyonu (PZR) sonuçları ise %56 S. anginosus, %24 S.constellatus, %8’i S. intermedius olarak saptanmıştır. Suşların %12’sinde herhangi bir bant saptanamamıştır. Sonuç: Streptococcus milleri grubun biyokimyasal identifikasyon şemasının henüz tam olarak tanımlanmadığı saptanmıştır. Antibiyotik duyarlılık sonuçlarımıza göre SMG’de direnç gelişimi ciddi boyutta değildir. Grubu tanımlamada ticari kitlerin yetersiz olabileceği ve yapılacak çalışmaların mutlaka moleküler bir yöntemle desteklenmesi gerekmektedir. Streptococcus milleri grubun biyokimyasal identifikasyon şemasının henüz tam olarak tanımlanmadığı saptanmıştır. Antibiyotik duyarlılık sonuçlarımıza göre SMG’de direnç gelişimi ciddi boyutta değildir. Grubu tanımlamada ticari kitlerin yetersiz olabileceği ve yapılacak çalışmaların mutlaka moleküler bir yöntemle desteklenmesi gerekmektedir.

Supporting Institution

İstanbul Üniversitesi Bilimsel Araştırma Projeleri Birimi (İÜ-BAP)

Project Number

17164

Thanks

İstanbul Üniversitesi Bilimsel Araştırma Projeleri Birimi’ne desteklerinden ötürü teşekkür ederiz. The University of Tokushima’dan Dr. Hideaki Nagamune ve Queen Mary, University of London’dan Dr. Robert A. Whiley’e deneylerimizde kullandığımız standart suşları (S. anginosus NCTC 10713 S. intermedius NCDO 2227 ve S. constellatus subsp. constellatus NCDO2226) temin edip bizimle bilgi alışverişinde oldukları için teşekkür ederiz.

References

  • 1- Forbes BA, Sham DF, Weissfeld AS. Catalase-nagative, Grampositive cocci, Streptococcus, enterococcus and smilar organisms. In: Tille PM, editors. Bailey and Scott’s Diagnostic Microbiology. Missouri: Mosby; 2008.p.298-315. google scholar
  • 2- Procop GW, Church CL, Hall GS, Janda WM, Koneman EW, Schreckenberger PC, Woods GL. Isolation and identification of Streptococci and Streptococcus-Like bacteria. Koneman’s Color Atlas and Textbook of Diagnostic Microbiology. Philadelphia: Lippincott Williams&Wilkins; 2006.p.701-45. google scholar
  • 3- Ruoff KL, Bisno AL. Classification of Streptococci. In: Mandell GL, Bannett JE, Dolin R, editors. Principles and Practice of Infectious Diseases. Philadelphia: Elsevier Churchill Livingstone; 2005.p.2591-610. google scholar
  • 4- Patterson MJ. Streptococcus. In: Baron S, editor. Medical Microbiology. 4th edition. Galveston (TX): University of Texas Medical Branch at Galveston; 1996. Chapter 13. Available from: https://www.ncbi.nlm.nih.gov/books/NBK7611/ google scholar
  • 5- Hansen AP, Hucker GJ, Snyder MA. Hemolytic properties of the mastitis streptococcus. American J Public Health 1933;23(12):1262-70. google scholar
  • 6- Sherman JM. The Streptococci. Microbiol Mol Biol Reviews 1937;1(1):3-97. google scholar
  • 7- Lancefield R. A serological differentiation of human and other groups of hemolytic streptococci. J Exp Med 1933;57(4):571-95. google scholar
  • 8- Carroll KC. Biographical Feature: Rebecca Lancefield, Ph.D. J Clin Microbiol. 2019;57(8):e00728-19. doi:10.1128/JCM.00728-19 google scholar
  • 9- Hardie JM, Whiley RA. Classification and overview of the genera Streptococcus and Enterococcus. J Appl Microbiol. 1997;83(S1):1S-11S. doi:10.1046/j.1365-2672.83.s1.1.x google scholar
  • 10- Patel S, Gupta RS. Robust demarcation of fourteen different species groups within the genus Streptococcus based on genomebased phylogenies and molecular signatures. Infect Genet Evol 2018;66:130-51. google scholar
  • 11- Yao JDC, Moellering RC-JR. Antibacterial Agents. In: Murray PR, Baron EJ, Jorgensen JH, Landry ML, Pfaller MA, editors. Manual of Clinical Microbiology. Washington: ASM press; 2007.p.1077-113. google scholar
  • 12- Berkiten R. Gram pozitif koklar: Laboratuvar tanı, epidemiyoloji, korunma. İçinde: Bozkaya E, editör. Tıbbi Mikrobiyoloji-2. Ankara: Nobel Tıp Kitabevleri; 2005.sf.1-28. google scholar
  • 13- Berkiten R. Mini (pinpoint) koloni oluşturan beta-hemolitik streptokoklar: Streptococcus milleri grubu ve çeşitli özellikleri. Ankem Dergisi 2008;22(2):89-94. google scholar
  • 14- Ardıç N. B grubu Streptokoklar (Streptococcus agalactiae). Klimik Dergisi 2003;16(3):101-5. google scholar
  • 15- Wilson L.G. The early recognition of streptococci as causes of disease. Medical history. 1987;31(4):403-14. google scholar
  • 16- Biochemical Tests for the Identification of Aerobic Bacteria. Editor(s):Amy L.Leber. Clinical Microbiology Procedures Handbook. 2016; Chapter 3.17. 4th Edition. Volume 1-3. doi:10.1128/9781555818814.ch3.17.1 google scholar
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  • 18- York MK, Traylor MM, Hardy J, Henry M. Biochemical tests for identification of aerobic bacteria. In: Lynne JG, editors. Clinical Microbiology Procedures Handbook, Washington DC: ASM press;2010. doi/book/10.1128/9781683670438.CMPH google scholar
  • 19- Performance Standards for Antimicrobial Susceptibility Testing; Twenty-second Informational Supplement. CLSI document M100-S22, Wayne PA; Clinical and Laboratory Standards Institute;2012. google scholar
  • 20- Takao A, Nagamune H, Maeda N. Identification of the Anginosus group within the genus Streptococcus using polymerase chain reaction. FEMS Microbiol letters 2004;233(1):83-9. google scholar
  • 21- Goto T, Nagamune H, Miyazaki A, Kawamura Y, Ohnishi O, Hattori K, et al. Rapid identification of Streptococcus intermedius by PCR with the ily gene as a species marker gene. J Med Microbiol 2002;51(2):178-86. google scholar
  • 22- Hoshino T, Fujiwara T, Kilian M. Use of phylogenetic and phenotypic analyses to identify nonhemolitic streptococci isolated from bacteremic patients. J Clin Microbiol 2005;43(12):6073-85. google scholar
  • 23- Jiang S, Li M, Fu T, Shan F, Jiang L, Shao Z. Clinical Characteristics of Infections Caused by Streptococcus Anginosus Group. Scientific reports 2020;10(1):9032. google scholar
  • 24- Pilarczyk-Zurek M, Sitkiewicz I, Koziel J. The Clinical View on Streptococcus anginosus group-opportunistic pathogens coming out of hiding. Frontiers in microbiology 2022:13;956677. google scholar
  • 25- Lehman DC, Mahon CR, Suvarna K. Streptococcus, enterococcus and other catalase-negative Gram-positive cocci. In: Mahon CR, Lehman DC ve Manuselis G, editors. Textbook of Diagnostic Microbiology. Philadelphia; Elsevier. 2007. p.382-95. google scholar
  • 26- Topçu AW, Söyletir G, Doğanay M. Beta Hemolitik Streptokoklar. İçinde: Söyletir G, Över U, editörler. İnfeksiyon Hastalıkları ve Mikrobiyolojisi, Etkenlere Göre İnfeksiyonlar. Ankara; Nobel Tıp Kitabevleri:2002.sf.1478-97. google scholar
  • 27- Fox K, Turner J, Fox A. Role of beta-hemolytic group C streptococci in pharyngitis: Incidence and Biochemical Characteristics of Streptococcus equisimilis and Streptococcus anginosus in patients and healty controls. J Clin Microbiol 1993;31(4):804-7. google scholar
  • 28- Al Amri A, Senok AC, Al Ismaeel AY, Al-Mahmeed AE, Botta GA. Multiplex PCR for direct identification of Campylobacter spp. in human and chicken stools. J Medical Microbiol 2007;56(Pt 10):1350-55. google scholar
  • 29- Niven CF, Smiley KL, Sherman JM. The hydrolysis of arginine by streptococci. J Bacteriol 1942;43(6):651-60. google scholar
  • 30- Woo PC, Tse H, Chan KM, Lau SK, Fung AM, Yip KT, et al. Streptococcus milleri endocarditis caused by Streptococcus anginosus. Diag Microbiol and Infec Disease 2004;48(2):81-8. google scholar
  • 31- Bantar C, Fernandez Canigia L, Relloso S, Lanza A, Bianchini H, Smayevsky J. Species belonging to the Streptococcus milleri group: antimicrobial susceptibility and comparative prevalence in significant clinical specimens. American Soc for Microbiol 1996;34(8):2020-22. google scholar

IDENTIFICATION OF STREPTOCOCCUS MILLERI GROUP (SMG) BACTERIA BY BIOCHEMICAL AND MOLECULAR METHODS

Year 2023, Volume: 6 Issue: 1, 73 - 79, 28.02.2023
https://doi.org/10.26650/JARHS2023-1164336

Abstract

Objective: The aim of this study was to differentiate the strains of Streptococcus milleri group (SMG) bacteria using molecular, biochemical and semi-automated techniques and to investigate the results of susceptibility to commonly used antibiotics in treatment. It was anticipated that the study would draw attention to some important points about SMG bacteria and to determine their epidemiological importance. Materials and method: A total of 100 streptococcal bacteria were included in our study. Biochemical tests and the API 20 STREP semiautomatic identification kit were used for species identification. Species identifications were confirmed by amplification of the target gene regions (HYL-MIX-U, HLY-INT-D, HLY-CC-D, 16S-ANG-U, 16S-ANG-D, ILY-4DFw, ILYwholeC Bw) by polymerase chain reaction (PCR). Results: Voges-Proskauer tests of all strains were positive, whereas hippurate hydrolysis and pyrrolidinyl arylamidase (PYR) tests were negative. All strains were evaluated in terms of Lancefield group antigen, and it was found that 42% were G, 26% were F, and 5% were C group antigen. It was not possible to determine antigen type with Lancefield group antigens in 27% of the strains. All of the isolates were found to be susceptible to vancomycin, ceftriaxone, cefepime, cefotaxime, levofloxacin, and linezolid. Erythromycin, clindamycin, tetracycline, and chloramphenicol resistance was found in 16%, 6%, 5%, and %1 of the isolates, respectively. Ampicillin and penicillin resistance were determined by the E-test method. One Streptococcus anginosus strain was resistant to ampicillin and penicillin, and two S. anginosus isolates were found to be intermediately susceptible to ampicillin. According to API 20 STREP results, 41% of the isolates were identified as genus level, 45% were S. anginosus, 5% were Streptococcus constellatus, and 1% were Streptococcus intermedius. Multiplex polymerase chain reaction (PCR) results were found to be 56% S. anginosus, 24% S. constellatus, and 8% S. intermedius. No band was detected in 12% of the strains. It has been determined that the biochemical identification scheme of the Streptococcus milleri group has not yet been fully defined. According to the antibiotic susceptibility results, the development of resistance in SMG is not critical. Commercial kits are unsatisfactory in identifying this group, and the studies should be supported with molecular methods.

Project Number

17164

References

  • 1- Forbes BA, Sham DF, Weissfeld AS. Catalase-nagative, Grampositive cocci, Streptococcus, enterococcus and smilar organisms. In: Tille PM, editors. Bailey and Scott’s Diagnostic Microbiology. Missouri: Mosby; 2008.p.298-315. google scholar
  • 2- Procop GW, Church CL, Hall GS, Janda WM, Koneman EW, Schreckenberger PC, Woods GL. Isolation and identification of Streptococci and Streptococcus-Like bacteria. Koneman’s Color Atlas and Textbook of Diagnostic Microbiology. Philadelphia: Lippincott Williams&Wilkins; 2006.p.701-45. google scholar
  • 3- Ruoff KL, Bisno AL. Classification of Streptococci. In: Mandell GL, Bannett JE, Dolin R, editors. Principles and Practice of Infectious Diseases. Philadelphia: Elsevier Churchill Livingstone; 2005.p.2591-610. google scholar
  • 4- Patterson MJ. Streptococcus. In: Baron S, editor. Medical Microbiology. 4th edition. Galveston (TX): University of Texas Medical Branch at Galveston; 1996. Chapter 13. Available from: https://www.ncbi.nlm.nih.gov/books/NBK7611/ google scholar
  • 5- Hansen AP, Hucker GJ, Snyder MA. Hemolytic properties of the mastitis streptococcus. American J Public Health 1933;23(12):1262-70. google scholar
  • 6- Sherman JM. The Streptococci. Microbiol Mol Biol Reviews 1937;1(1):3-97. google scholar
  • 7- Lancefield R. A serological differentiation of human and other groups of hemolytic streptococci. J Exp Med 1933;57(4):571-95. google scholar
  • 8- Carroll KC. Biographical Feature: Rebecca Lancefield, Ph.D. J Clin Microbiol. 2019;57(8):e00728-19. doi:10.1128/JCM.00728-19 google scholar
  • 9- Hardie JM, Whiley RA. Classification and overview of the genera Streptococcus and Enterococcus. J Appl Microbiol. 1997;83(S1):1S-11S. doi:10.1046/j.1365-2672.83.s1.1.x google scholar
  • 10- Patel S, Gupta RS. Robust demarcation of fourteen different species groups within the genus Streptococcus based on genomebased phylogenies and molecular signatures. Infect Genet Evol 2018;66:130-51. google scholar
  • 11- Yao JDC, Moellering RC-JR. Antibacterial Agents. In: Murray PR, Baron EJ, Jorgensen JH, Landry ML, Pfaller MA, editors. Manual of Clinical Microbiology. Washington: ASM press; 2007.p.1077-113. google scholar
  • 12- Berkiten R. Gram pozitif koklar: Laboratuvar tanı, epidemiyoloji, korunma. İçinde: Bozkaya E, editör. Tıbbi Mikrobiyoloji-2. Ankara: Nobel Tıp Kitabevleri; 2005.sf.1-28. google scholar
  • 13- Berkiten R. Mini (pinpoint) koloni oluşturan beta-hemolitik streptokoklar: Streptococcus milleri grubu ve çeşitli özellikleri. Ankem Dergisi 2008;22(2):89-94. google scholar
  • 14- Ardıç N. B grubu Streptokoklar (Streptococcus agalactiae). Klimik Dergisi 2003;16(3):101-5. google scholar
  • 15- Wilson L.G. The early recognition of streptococci as causes of disease. Medical history. 1987;31(4):403-14. google scholar
  • 16- Biochemical Tests for the Identification of Aerobic Bacteria. Editor(s):Amy L.Leber. Clinical Microbiology Procedures Handbook. 2016; Chapter 3.17. 4th Edition. Volume 1-3. doi:10.1128/9781555818814.ch3.17.1 google scholar
  • 17- Bilgehan H. Klinik Mikrobiyolojik Tanı. Barış Yayınları: İzmir; 2002:495-521. google scholar
  • 18- York MK, Traylor MM, Hardy J, Henry M. Biochemical tests for identification of aerobic bacteria. In: Lynne JG, editors. Clinical Microbiology Procedures Handbook, Washington DC: ASM press;2010. doi/book/10.1128/9781683670438.CMPH google scholar
  • 19- Performance Standards for Antimicrobial Susceptibility Testing; Twenty-second Informational Supplement. CLSI document M100-S22, Wayne PA; Clinical and Laboratory Standards Institute;2012. google scholar
  • 20- Takao A, Nagamune H, Maeda N. Identification of the Anginosus group within the genus Streptococcus using polymerase chain reaction. FEMS Microbiol letters 2004;233(1):83-9. google scholar
  • 21- Goto T, Nagamune H, Miyazaki A, Kawamura Y, Ohnishi O, Hattori K, et al. Rapid identification of Streptococcus intermedius by PCR with the ily gene as a species marker gene. J Med Microbiol 2002;51(2):178-86. google scholar
  • 22- Hoshino T, Fujiwara T, Kilian M. Use of phylogenetic and phenotypic analyses to identify nonhemolitic streptococci isolated from bacteremic patients. J Clin Microbiol 2005;43(12):6073-85. google scholar
  • 23- Jiang S, Li M, Fu T, Shan F, Jiang L, Shao Z. Clinical Characteristics of Infections Caused by Streptococcus Anginosus Group. Scientific reports 2020;10(1):9032. google scholar
  • 24- Pilarczyk-Zurek M, Sitkiewicz I, Koziel J. The Clinical View on Streptococcus anginosus group-opportunistic pathogens coming out of hiding. Frontiers in microbiology 2022:13;956677. google scholar
  • 25- Lehman DC, Mahon CR, Suvarna K. Streptococcus, enterococcus and other catalase-negative Gram-positive cocci. In: Mahon CR, Lehman DC ve Manuselis G, editors. Textbook of Diagnostic Microbiology. Philadelphia; Elsevier. 2007. p.382-95. google scholar
  • 26- Topçu AW, Söyletir G, Doğanay M. Beta Hemolitik Streptokoklar. İçinde: Söyletir G, Över U, editörler. İnfeksiyon Hastalıkları ve Mikrobiyolojisi, Etkenlere Göre İnfeksiyonlar. Ankara; Nobel Tıp Kitabevleri:2002.sf.1478-97. google scholar
  • 27- Fox K, Turner J, Fox A. Role of beta-hemolytic group C streptococci in pharyngitis: Incidence and Biochemical Characteristics of Streptococcus equisimilis and Streptococcus anginosus in patients and healty controls. J Clin Microbiol 1993;31(4):804-7. google scholar
  • 28- Al Amri A, Senok AC, Al Ismaeel AY, Al-Mahmeed AE, Botta GA. Multiplex PCR for direct identification of Campylobacter spp. in human and chicken stools. J Medical Microbiol 2007;56(Pt 10):1350-55. google scholar
  • 29- Niven CF, Smiley KL, Sherman JM. The hydrolysis of arginine by streptococci. J Bacteriol 1942;43(6):651-60. google scholar
  • 30- Woo PC, Tse H, Chan KM, Lau SK, Fung AM, Yip KT, et al. Streptococcus milleri endocarditis caused by Streptococcus anginosus. Diag Microbiol and Infec Disease 2004;48(2):81-8. google scholar
  • 31- Bantar C, Fernandez Canigia L, Relloso S, Lanza A, Bianchini H, Smayevsky J. Species belonging to the Streptococcus milleri group: antimicrobial susceptibility and comparative prevalence in significant clinical specimens. American Soc for Microbiol 1996;34(8):2020-22. google scholar
There are 31 citations in total.

Details

Primary Language Turkish
Subjects Clinical Sciences
Journal Section Research Articles
Authors

Gülşen Günel 0000-0003-1574-0231

Bülent Gürler 0000-0002-6433-5198

Project Number 17164
Publication Date February 28, 2023
Submission Date September 19, 2022
Published in Issue Year 2023 Volume: 6 Issue: 1

Cite

MLA Günel, Gülşen and Bülent Gürler. “STREPTOCOCCUS MILLERI GRUP (SMG) BAKTERİLERİNİN BİYOKİMYASAL VE MOLEKÜLER YÖNTEMLERLE TANIMLANMASI”. Sağlık Bilimlerinde İleri Araştırmalar Dergisi, vol. 6, no. 1, 2023, pp. 73-79, doi:10.26650/JARHS2023-1164336.