Research Article
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Determination of Genetic Diversity and Similarity among Methicillin Resistant Staphylococcus aureus strains by RAPD-PCR

Year 2022, Volume: 3 Issue: 1, 62 - 69, 30.01.2022

Halil Bal Nurten Altanlar

https://doi.org/10.51972/tfsd.1015796

Abstract

ABSTRACT:
Purpose: The aim of this study was to determine the genetic diversity and genetic similarity of Methicillin Resistant S.aureus (MRSA) strains isolated from clinical samples.
Material and Methods: Thirty-two MRSA strains were identified by conventional methods. Methicillin resistance of strains were determined by PCR using the mecA gene primers. These strains were genetically typed by RAPD PCR using primers OLP-11 and OLP-13. Bionumerics V7.5 (Applied Maths) program was used for analysis and dendograms were generated by unweighted pair group method with arithmetic averages (UPGMA).
Results: All strains were confirmed as MRSA by PCR. Many different bands from 400 bp to 1000 bp were detected by RAPD PCR and five clusters (1-5) with OLP-11 and four clusters (1-4) were formed with OLP-13. In RAPD PCR performed with OLP-11 and OLP-13 primers, 80% (cluster 3-5) and 86% (cluster 1-4) similarities were found, respectively. MRSA strains isolated from wound samples were found to be more genetically similar to each other, with at least one in each cluster.
Conclusion: RAPD PCR was found to be an effective method for the evaluation of genetic similarity and genetic diversity of MRSA strains.

Keywords

mecA OLP-11 OLP-13 RAPD PCR

References

  • Binelli, G., Bucci, G. (1994). A genetic linkage map of Picea abies Karst., based on RAPD markers, as a tool in population genetics. TAG. Theoretical and applied genetics. Theoretische und angewandte Genetik, 88(3-4), 283–288. https://doi.org/10.1007/BF00223633
  • Cocconcelli, PS., Porro, D., Galandini, S., Senini, L. (1995). Development of RAPD protocol for typing of strains of lactic acid bacteria and enterococci. Lett Appl Microbiol 21:376–379.
  • Debnath, A., Chikkaswamy, B.K. (2015). RandomlyAmplified Polymorphic DNA Assay of Methicillin Resistant Staphylococcus aureus Isolated from Clinical Samples from Bengaluru, India. International Journal of Current Microbiology and Applied Sciences ISSN: 2319-7706 Volume 4 Number 11 (2015) pp. 342-355.
  • Egan, A.J.F., Cleverley, R.M., Peters, K., Lewis, R.J. and Vollmer, W. (2017), Regulation of bacterial cell wall growth. FEBS J, 284: 851-867. https://doi.org/10.1111/febs.13959.
  • EUCAST Disk Diffusion Method for Antimicrobial Susceptibility Testing (2021). - Version 9.0.
  • Fuda, C., Suvorov, M., Vakulenko, B.S., Mobashery, S. (2004). The Basis for Resistance to β-Lactam Antibiotics by Penicillin-binding Protein 2a of Methicillin-resistant Staphylococcus aureus*, Journal of Biological Chemistry,Volume 279, Issue 39, 2004, Pages 40802-40806, ISSN 0021-9258. https://doi.org/10.1074/jbc.M403589200
  • Gould, D., Chamberlaine, A. (1995). Staphylococcus aureus: a review of the literature. Journal of clinical nursing, 4(1), 5–12. https://doi.org/10.1111/j.13652702.1995.tb00004.x
  • Hakimi Alni, R., Mohammadzadeh, A., Mahmoodi, P. (2017). RAPD-PCR analysis of Staphylococcus aureus strains isolated from different sources. Comp Clin Pathol 26, 823–830. https://doi.org/10.1007/s00580-017-2453-z
  • Hennekinne, J. A., Kerouanton, A., Brisabois, A., De Buyser, M. L. (2003). Discrimination of Staphylococcus aureus biotypes by pulsed-field gel electrophoresis of DNA macro-restriction fragments. Journal of applied microbiology, 94(2),321–329. https://doi.org/10.1046/j.13652672.2003.0187.x
  • Hookey, J. V., Richardson, J. F., Cookson, B. D. (1998). Molecular typing of Staphylococcus aureus based on PCR restriction fragment length polymorphism and DNA sequence analysis of the coagulase gene. Journal of clinical microbiology, 36(4),1083–1089. https://doi.org/10.1128/JCM.36.4.10831089.1998
  • Ivbule, M., Miklaševičs, E., Čupāne, L., Bērziņa, L., Bālinš, A., Valdovska, A. (2017). Presence of Methicillin-resistant Staphylococcus Aureus in Slaughterhouse Environment, Pigs, Carcasses, and Workers. Journal of veterinary research, 61(3),267–277. https://doi.org/10.1515/jvetres-2017-0037
  • Juhász-Kaszanyitzky, É., Jánosi, S., Somogyi, P., Dán, Á., van der Graaf-van Bloois, L.,Van Duijkeren, E. (2007). MRSA transmission betweencows and humans. Emerg Infect Diseases 13:630.
  • Keshava, C., Keshava, N., Zhou, G., Whong, WZ., Ong, TM. (1999). Genetic instability in silica- and cadmium chloride-transformed BALB/c-3T3 and tumor cell lines by randomly amplified polyinorphic DNA analysis. Mutat Res 425:117–123.
  • Kloos, WE., Schleifer, KH. (1986). The genus Staphylococcus. In: Sneath PHA, Mair NS, Sarpe ME, Holt JG (ed) Bergey's Manual of Systematic Bacteriology, 2nd ed. Williams and Wilkins, Baltimore.
  • Koh, TL., Khoo, G., Fan, LQ., Phang, VPE. (1999) Genetic diversity among wild forms and cultivated varieties of discus (Symphysodon spp.) as revealed by randomly amplified polymorphic DNA (RAPD) fingerprinting. Aquaculture 173:485–497.
  • Kurlenda, J., Grinholc, M., Jasek, K., Wegrzyn, G. (2007). RAPD typing of methicillin-resistant Staphylococcus aureus: a 7-year experience in a Polish hospital. Medical science monitor : international medical journal of experimental and clinical research, 13(6), MT13–MT18.
  • Lee, J. H. (2003). Methicillin (Oxacillin)-resistant Staphylococcus aureus strains isolated from major food animals and their potential transmission to humans. Applied and environmental microbiology, 69(11), 6489–6494.https://doi.org/10.1128/AEM.69.11.64896494.2003
  • Ma, H., Cheng, J., Peng, L., Gao, Y., Zhang, G., & Luo, Z. (2020). Adjunctive rifampin for the treatment of Staphylococcus aureus bacteremia with deep infections: A meta-analysis. PloS one, 15(3), e0230383. https://doi.org/10.1371/journal.pone.0230383
  • Maes, N., Magdalena, J., Rottiers, S., De Gheldre, Y., Struelens, M. J. (2002). Evaluation of a triplex PCR assay to discriminate Staphylococcus aureus from coagulase-negative Staphylococci and determine methicillin resistance from blood cultures. Journal of clinical microbiology, 40(4), 1514–1517. https://doi.org/10.1128/JCM.40.4.15141517.2002.
  • McGuinness, W. A., Malachowa, N., & DeLeo, F. R. (2017). Vancomycin Resistance in Staphylococcus aureus. The Yale journal of biology and medicine, 90(2), 269–281.
  • Morandi, S., Brasca, M., Lodi, R., Brusetti, L., Andrighetto, C., Lombardi, A. (2010). Biochemical profiles, restriction fragment length polymorphism (RFLP), random amplified polymorphic DNA (RAPD) and multilocus variable number tandem repeat analysis (MLVA) for typing Staphylococcus aureus isolated from dairy products, Research in Veterinary Science, Volume 88, Issue 3, Pages 427-435, ISSN 0034-5288, https://doi.org/10.1016/j.rvsc.2009.10.005.
  • Nikbakht, M., Nahaei M.R., Akhi, M.T., Asgharzadeh, M., Nikvash, S. (2008). Molecular fingerprinting ofmeticillinresistant Staphylococcus aureus strains isolated from patients and staff of two Iranian hospitals. J HospInfect.; 69:46-55.
  • Nnachi, AU., Emele F.E., Ukaegbu, C.O., Agah, M.V., Udu-Ibiam, O.E., Chukwu, O.S. (2014). Prevalence of methicillin-resistant Staphylococcus aureus (MRSA) in raw meat and meat handlers in Onitsha, Nigeria. Eur J Prev Med 2:9–15.
  • Onasanya, A., Mignouna, H.D., Thottappilly, G. (2003). Genetic fingerprinting and phylogenetic diversity of Staphylococcus aureus isolates from Nigeria. Afr. J.Biotechnol. 2 (8), 246–250.
  • Rabello, R. F., Moreira, B. M., Lopes, R., Teixeira, L. M., Riley, L. W., & Castro, A. (2007). Multilocus sequence typing of Staphylococcus aureus isolates recovered from cows with mastitis in Brazilian dairy herds. Journal of medical microbiology, 56(Pt 11), 1505–1511. https://doi.org/10.1099/jmm.0.47357-0
  • Rađenović, M., Ašanin, J., Aksentijević, K., & Mišić, D. (2016). Prisustvo sojeva stafilokoka rezistentnih na meticilin kod studenata Fakulteta veterinarske medicine Univerziteta u Beogradu. Arhiv veterinarske medicine Naučni institut za veterinarstvo "Novi Sad", Novi Sad., 9(2), 17-28.
  • Reinoso, E., Bettera, S., Frigerio, C., DiRenzo, M., Calzolari, A., & Bogni, C. (2004). RAPD-PCR analysis of Staphylococcus aureus strains isolated from bovine and human hosts. Microbiological research, 159(3), 245–255. https://doi.org/10.1016/j.micres.2004.04.002
  • Rocco, L., Izzo, A., Zito, G., Peluso, C., Stingo, V . (2011) Genotoxicity in zebrafish (Danio rerio) exposed to two pharmacological products from an impacted Italian river. J Environ Anal Toxicol 1:103. doi:10.4172/2161-0525.1000103
  • Sabat, A., Krzyszton-Russjan, J., Strzalka, W., Filipek, R., Kosowska, K., Hryniewicz, W., Travis, J., & Potempa, J. (2003). New method for typing Staphylococcus aureus strains: multiple-locus variable-number tandem repeat analysis of polymorphism and genetic relationships of clinical isolates. Journal of clinical microbiology, 41(4), 1801–1804. https://doi.org/10.1128/jcm.41.4.18011804.2003
  • Sauvage, E., Kerff, F., Terrak, M., Ayala, J.A., Charlier, P. (2008). The penicillin-binding proteins: structure and role in peptidoglycan biosynthesis, FEMS Microbiology Reviews, Volume 32, Issue 2, Pages 234–258, https://doi.org/10.1111/j.15746976.2008.00105.x.
  • Shoen, H., Rose, S. J., Ramsey, S. A., de Morais, H., & Bermudez, L. (2019). Analysis of Staphylococcus infections in a veterinary teaching hospital from 2012 to 2015. Comparative immunology, microbiology and infectious diseases, 66, 101332. https://doi.org/10.1016/j.cimid.2019.101332
  • Tambic, A., Power, E.G., Talsania, H., Anthony, R.M., French, G.L. (1997). Analysis of an outbreak of non-phage-typeable methicillin-resistant Staphylococcus aureus by usingrandomly amplified polymorphic DNA assay. Vet. Microbiol. 82, 61–67.
  • Typas, A., Banzhaf, M., Gross, C. A., & Vollmer, W. (2011). From the regulation of peptidoglycan synthesis to bacterial growth and morphology. Nature reviews. Microbiology, 10(2), 123–136. https://doi.org/10.1038/nrmicro2677.
  • Williams, J. G., Kubelik, A. R., Livak, K. J., Rafalski, J. A., & Tingey, S. V. (1990). DNA polymorphisms amplified by arbitrary primers are useful as genetic markers. Nucleic acids research, 18(22), 6531–6535. https://doi.org/10.1093/nar/18.22.6531
  • Zare, S., Derakhshandeh, A., Haghkhah, M., Naziri, Z., & Broujeni, A. M. (2019). Molecular typing of Staphylococcus aureus from different sources by RAPD-PCR analysis. Heliyon, 5(8), e02231. https://doi.org/10.1016/j.heliyon.2019.e02231.

Year 2022, Volume: 3 Issue: 1, 62 - 69, 30.01.2022

Halil Bal Nurten Altanlar

https://doi.org/10.51972/tfsd.1015796

Abstract

References

  • Binelli, G., Bucci, G. (1994). A genetic linkage map of Picea abies Karst., based on RAPD markers, as a tool in population genetics. TAG. Theoretical and applied genetics. Theoretische und angewandte Genetik, 88(3-4), 283–288. https://doi.org/10.1007/BF00223633
  • Cocconcelli, PS., Porro, D., Galandini, S., Senini, L. (1995). Development of RAPD protocol for typing of strains of lactic acid bacteria and enterococci. Lett Appl Microbiol 21:376–379.
  • Debnath, A., Chikkaswamy, B.K. (2015). RandomlyAmplified Polymorphic DNA Assay of Methicillin Resistant Staphylococcus aureus Isolated from Clinical Samples from Bengaluru, India. International Journal of Current Microbiology and Applied Sciences ISSN: 2319-7706 Volume 4 Number 11 (2015) pp. 342-355.
  • Egan, A.J.F., Cleverley, R.M., Peters, K., Lewis, R.J. and Vollmer, W. (2017), Regulation of bacterial cell wall growth. FEBS J, 284: 851-867. https://doi.org/10.1111/febs.13959.
  • EUCAST Disk Diffusion Method for Antimicrobial Susceptibility Testing (2021). - Version 9.0.
  • Fuda, C., Suvorov, M., Vakulenko, B.S., Mobashery, S. (2004). The Basis for Resistance to β-Lactam Antibiotics by Penicillin-binding Protein 2a of Methicillin-resistant Staphylococcus aureus*, Journal of Biological Chemistry,Volume 279, Issue 39, 2004, Pages 40802-40806, ISSN 0021-9258. https://doi.org/10.1074/jbc.M403589200
  • Gould, D., Chamberlaine, A. (1995). Staphylococcus aureus: a review of the literature. Journal of clinical nursing, 4(1), 5–12. https://doi.org/10.1111/j.13652702.1995.tb00004.x
  • Hakimi Alni, R., Mohammadzadeh, A., Mahmoodi, P. (2017). RAPD-PCR analysis of Staphylococcus aureus strains isolated from different sources. Comp Clin Pathol 26, 823–830. https://doi.org/10.1007/s00580-017-2453-z
  • Hennekinne, J. A., Kerouanton, A., Brisabois, A., De Buyser, M. L. (2003). Discrimination of Staphylococcus aureus biotypes by pulsed-field gel electrophoresis of DNA macro-restriction fragments. Journal of applied microbiology, 94(2),321–329. https://doi.org/10.1046/j.13652672.2003.0187.x
  • Hookey, J. V., Richardson, J. F., Cookson, B. D. (1998). Molecular typing of Staphylococcus aureus based on PCR restriction fragment length polymorphism and DNA sequence analysis of the coagulase gene. Journal of clinical microbiology, 36(4),1083–1089. https://doi.org/10.1128/JCM.36.4.10831089.1998
  • Ivbule, M., Miklaševičs, E., Čupāne, L., Bērziņa, L., Bālinš, A., Valdovska, A. (2017). Presence of Methicillin-resistant Staphylococcus Aureus in Slaughterhouse Environment, Pigs, Carcasses, and Workers. Journal of veterinary research, 61(3),267–277. https://doi.org/10.1515/jvetres-2017-0037
  • Juhász-Kaszanyitzky, É., Jánosi, S., Somogyi, P., Dán, Á., van der Graaf-van Bloois, L.,Van Duijkeren, E. (2007). MRSA transmission betweencows and humans. Emerg Infect Diseases 13:630.
  • Keshava, C., Keshava, N., Zhou, G., Whong, WZ., Ong, TM. (1999). Genetic instability in silica- and cadmium chloride-transformed BALB/c-3T3 and tumor cell lines by randomly amplified polyinorphic DNA analysis. Mutat Res 425:117–123.
  • Kloos, WE., Schleifer, KH. (1986). The genus Staphylococcus. In: Sneath PHA, Mair NS, Sarpe ME, Holt JG (ed) Bergey's Manual of Systematic Bacteriology, 2nd ed. Williams and Wilkins, Baltimore.
  • Koh, TL., Khoo, G., Fan, LQ., Phang, VPE. (1999) Genetic diversity among wild forms and cultivated varieties of discus (Symphysodon spp.) as revealed by randomly amplified polymorphic DNA (RAPD) fingerprinting. Aquaculture 173:485–497.
  • Kurlenda, J., Grinholc, M., Jasek, K., Wegrzyn, G. (2007). RAPD typing of methicillin-resistant Staphylococcus aureus: a 7-year experience in a Polish hospital. Medical science monitor : international medical journal of experimental and clinical research, 13(6), MT13–MT18.
  • Lee, J. H. (2003). Methicillin (Oxacillin)-resistant Staphylococcus aureus strains isolated from major food animals and their potential transmission to humans. Applied and environmental microbiology, 69(11), 6489–6494.https://doi.org/10.1128/AEM.69.11.64896494.2003
  • Ma, H., Cheng, J., Peng, L., Gao, Y., Zhang, G., & Luo, Z. (2020). Adjunctive rifampin for the treatment of Staphylococcus aureus bacteremia with deep infections: A meta-analysis. PloS one, 15(3), e0230383. https://doi.org/10.1371/journal.pone.0230383
  • Maes, N., Magdalena, J., Rottiers, S., De Gheldre, Y., Struelens, M. J. (2002). Evaluation of a triplex PCR assay to discriminate Staphylococcus aureus from coagulase-negative Staphylococci and determine methicillin resistance from blood cultures. Journal of clinical microbiology, 40(4), 1514–1517. https://doi.org/10.1128/JCM.40.4.15141517.2002.
  • McGuinness, W. A., Malachowa, N., & DeLeo, F. R. (2017). Vancomycin Resistance in Staphylococcus aureus. The Yale journal of biology and medicine, 90(2), 269–281.
  • Morandi, S., Brasca, M., Lodi, R., Brusetti, L., Andrighetto, C., Lombardi, A. (2010). Biochemical profiles, restriction fragment length polymorphism (RFLP), random amplified polymorphic DNA (RAPD) and multilocus variable number tandem repeat analysis (MLVA) for typing Staphylococcus aureus isolated from dairy products, Research in Veterinary Science, Volume 88, Issue 3, Pages 427-435, ISSN 0034-5288, https://doi.org/10.1016/j.rvsc.2009.10.005.
  • Nikbakht, M., Nahaei M.R., Akhi, M.T., Asgharzadeh, M., Nikvash, S. (2008). Molecular fingerprinting ofmeticillinresistant Staphylococcus aureus strains isolated from patients and staff of two Iranian hospitals. J HospInfect.; 69:46-55.
  • Nnachi, AU., Emele F.E., Ukaegbu, C.O., Agah, M.V., Udu-Ibiam, O.E., Chukwu, O.S. (2014). Prevalence of methicillin-resistant Staphylococcus aureus (MRSA) in raw meat and meat handlers in Onitsha, Nigeria. Eur J Prev Med 2:9–15.
  • Onasanya, A., Mignouna, H.D., Thottappilly, G. (2003). Genetic fingerprinting and phylogenetic diversity of Staphylococcus aureus isolates from Nigeria. Afr. J.Biotechnol. 2 (8), 246–250.
  • Rabello, R. F., Moreira, B. M., Lopes, R., Teixeira, L. M., Riley, L. W., & Castro, A. (2007). Multilocus sequence typing of Staphylococcus aureus isolates recovered from cows with mastitis in Brazilian dairy herds. Journal of medical microbiology, 56(Pt 11), 1505–1511. https://doi.org/10.1099/jmm.0.47357-0
  • Rađenović, M., Ašanin, J., Aksentijević, K., & Mišić, D. (2016). Prisustvo sojeva stafilokoka rezistentnih na meticilin kod studenata Fakulteta veterinarske medicine Univerziteta u Beogradu. Arhiv veterinarske medicine Naučni institut za veterinarstvo "Novi Sad", Novi Sad., 9(2), 17-28.
  • Reinoso, E., Bettera, S., Frigerio, C., DiRenzo, M., Calzolari, A., & Bogni, C. (2004). RAPD-PCR analysis of Staphylococcus aureus strains isolated from bovine and human hosts. Microbiological research, 159(3), 245–255. https://doi.org/10.1016/j.micres.2004.04.002
  • Rocco, L., Izzo, A., Zito, G., Peluso, C., Stingo, V . (2011) Genotoxicity in zebrafish (Danio rerio) exposed to two pharmacological products from an impacted Italian river. J Environ Anal Toxicol 1:103. doi:10.4172/2161-0525.1000103
  • Sabat, A., Krzyszton-Russjan, J., Strzalka, W., Filipek, R., Kosowska, K., Hryniewicz, W., Travis, J., & Potempa, J. (2003). New method for typing Staphylococcus aureus strains: multiple-locus variable-number tandem repeat analysis of polymorphism and genetic relationships of clinical isolates. Journal of clinical microbiology, 41(4), 1801–1804. https://doi.org/10.1128/jcm.41.4.18011804.2003
  • Sauvage, E., Kerff, F., Terrak, M., Ayala, J.A., Charlier, P. (2008). The penicillin-binding proteins: structure and role in peptidoglycan biosynthesis, FEMS Microbiology Reviews, Volume 32, Issue 2, Pages 234–258, https://doi.org/10.1111/j.15746976.2008.00105.x.
  • Shoen, H., Rose, S. J., Ramsey, S. A., de Morais, H., & Bermudez, L. (2019). Analysis of Staphylococcus infections in a veterinary teaching hospital from 2012 to 2015. Comparative immunology, microbiology and infectious diseases, 66, 101332. https://doi.org/10.1016/j.cimid.2019.101332
  • Tambic, A., Power, E.G., Talsania, H., Anthony, R.M., French, G.L. (1997). Analysis of an outbreak of non-phage-typeable methicillin-resistant Staphylococcus aureus by usingrandomly amplified polymorphic DNA assay. Vet. Microbiol. 82, 61–67.
  • Typas, A., Banzhaf, M., Gross, C. A., & Vollmer, W. (2011). From the regulation of peptidoglycan synthesis to bacterial growth and morphology. Nature reviews. Microbiology, 10(2), 123–136. https://doi.org/10.1038/nrmicro2677.
  • Williams, J. G., Kubelik, A. R., Livak, K. J., Rafalski, J. A., & Tingey, S. V. (1990). DNA polymorphisms amplified by arbitrary primers are useful as genetic markers. Nucleic acids research, 18(22), 6531–6535. https://doi.org/10.1093/nar/18.22.6531
  • Zare, S., Derakhshandeh, A., Haghkhah, M., Naziri, Z., & Broujeni, A. M. (2019). Molecular typing of Staphylococcus aureus from different sources by RAPD-PCR analysis. Heliyon, 5(8), e02231. https://doi.org/10.1016/j.heliyon.2019.e02231.
There are 35 citations in total.

Details

Primary Language English
Subjects Health Care Administration
Journal Section Articles
Authors

Halil Bal 0000-0002-0017-3425

Nurten Altanlar 0000-0003-2977-2269

Publication Date January 30, 2022
Submission Date November 1, 2021
Acceptance Date January 25, 2022
Published in Issue Year 2022 Volume: 3 Issue: 1

Cite

APA Bal, H., & Altanlar, N. (2022). Determination of Genetic Diversity and Similarity among Methicillin Resistant Staphylococcus aureus strains by RAPD-PCR. Turkish Journal of Science and Health, 3(1), 62-69. https://doi.org/10.51972/tfsd.1015796
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