Acta Univ. Agric. Silvic. Mendelianae Brun. 2012, 60(6), 251-256 | DOI: 10.11118/actaun201260060251

Rep-PCR-based typing as a tool for tracking of MRSA infection origin

Ivan Manga, Marcela Vyletělová
Agrovýzkum Rapotín s. r. o., Výzkumný ústav pro chov skotu, s. r. o., Výzkumníků 267, 788 13 Vikýřovice, Česká republika

A performance of rep-PCR fingerprinting method with the (GTG)5 primer was explored in order to evaluate its practical application for confirmation the source of human staphylococci infection. A laboratory worker daily working with the MRSA and Staphylococcus aureus strains has been tested positively for MRSA nosocomial infection. The collection of nine MRSA strains held in the laboratory has been typed with the rep-PCR method. Analysis of the fingerprints with the unweighted pair-group method using arithmetic averages (UPGMA) clustering method revealed presence of six strain clusters with similarity lower than 85%. The fingerprint of MRSA strain infecting the human differed significantly from remaining fingerprints. This provided clear evidence that MRSA strain infecting the personal did not come from the laboratory strains collection. Our experimental results showed rep-PCR with the (GTG)5 primer as an effective tool applicable for differentiation of individual S. aureus strains. However, the reproducibility and discrimination power of the method depended on strict observance of the optimal PCR time and temperature profile and PCR composition as well.

Keywords: rep-PCR, fingerprinting, MRSA, infection
Grants and funding:

This research was supported by the projects No. LA10029, QH81111 and the Institutional support No. RO0312 of the Ministry of Agriculture of the Czech Republic.

Received: June 29, 2012; Prepublished online: July 19, 2013; Published: July 20, 2013  Show citation

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Manga, I., & Vyletělová, M. (2012). Rep-PCR-based typing as a tool for tracking of MRSA infection origin. Acta Universitatis Agriculturae et Silviculturae Mendelianae Brunensis60(6), 251-256. doi: 10.11118/actaun201260060251
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References

  1. DE LENCASTRE, H., 2001: MRSA: origin and evolution of a resistant gene and emergence and spread of resistant clones, In: Proceedings from the MicroBiotec 2011 conference, Braga, Portugal, 1.-3. 12. 2011, p. 11, ISBN 978-989-97478-1-4.
  2. GRISOLD, A. J., ZARFEL, G., HOENIGL, M., KRZIWANEK, K., FEIERL, G., MASOUD, L., LEITNER, E., WAGNER-EIBEL, U., BADURA, A., MARTH, E., 2010: Occurrence and genotyping using automated repetitive-sequence-based PCR of methicillin-resistant Staphylococcus aureus ST398 in Southeast Austria. Diagn. Microbiol. Infect. Dis. 66: 217-221. ISSN 0732-8893. DOI: 10.1016/j.diagmicrobio.2009.09.006 Go to original source...
  3. HEALY, M., HUONG, J., BITTNER, T., LISING, M., FRYE, S., RAZA, S., SCHROCK, R., MANRY, J., RENWICK, A., NIETO, R., WOODS, C., VERSALOVIC, J., LUPSKI, J. R., 2005: Microbial DNA typing by automated repetitive-sequence-based PCR. J. Clin. Microbiol. 43: 199-207. ISSN 0095-1137. DOI: 10.1128/JCM.43.1.199-207.2005 Go to original source...
  4. IACUMIN, L., COMI, G., CANTONI, C., COCOLIN, L., 2006: Molecular and technological characterization of Staphylococcus xylosus isolated from naturally fermented Italian sausages by RAPD, rep-PCR and Sau-PCR analysis. Meat Sci. 74: 281-288. ISSN 0309-1740. DOI: 10.1016/j.meatsci.2006.03.020 Go to original source...
  5. KOREŇOVÁ, J., LOPA©OVSKÁ, J., KUCHTA, T., 2009: Biofilm forming bacterial contaminants in small and medium-sized ewes'milk and meat processing enterprises in Slovakia. J. Food Nutr. Res. 48: 115-120. ISSN 1336-8672.
  6. MANGA, I., VYLETĚLOVÁ, M., KARPI©KOVÁ, R., ©«ÁSTKOVÁ, Z., 2011: Molecular characterization of MRSA strains isolated from livestock milk and meat in the Czech Republic, In: Book of abstracts from the SfAM Summer Conference, Dublin, Ireland, 4-7. 7. 2011, P85, 55-56.
  7. POULSEN, A. B., SKOV, R., PALLESEN, L. V., 2003: Detection of methicillin resistance in coagulase-negative staphylococci and in staphylococci directly from simulated blood cultures using the EVIGENE MRSA Detection Kit. J. Antimicrob. Chemother. 51: 419-421. ISSN 0305-7453. DOI: 10.1093/jac/dkg084 Go to original source...
  8. RANTSIOU, K., IACUMIN, L., CANTONI, C., COMI, G., COCOLIN, L., 2005: Ecology and characterization by molecular methods of Staphylococcus species isolated from fresh sausages. Int. J. Food. Microbiol. 97: 277-284. ISSN 0168-1605. DOI: 10.1016/j.ijfoodmicro.2004.02.018 Go to original source...
  9. ROSS, T. L., MERZ, W. G., FARKOSH, A., CARROLL, K. C., 2005: Comparison of an automated repetitive sequence-based PCR microbial typing system to pulsed-field gel electrophoresis for analysis of outbreaks of methicillin-resistant Staphylococcus aureus. J. Clin. Microbiol. 43: 5642-5647. ISSN 0095-1137. DOI: 10.1128/JCM.43.11.5642-5647.2005 Go to original source...
  10. SNELLING, A., GERNER-SMIDT, P., HAWKEY, P., HERITAGE, J., PARNELL, P., PORTER, C., BODENHAM, A., INGLIS, T., 1996: Validation of use of whole-cell repetitive extragenic palindromic sequence-based PCR (REP-PCR) for typing strains belonging to the Acinetobacter calcoaceticus-Acinetobacter baumannii complex and application of the method to the investigation of a hospital outbreak. J. Clin. Microbiol. 34: 1193-1202. ISSN 0095-1137. Go to original source...
  11. STEGGER, M., LINDSAY, A. J., MOODLEY, A., SKOV, R. M., BROENS, E., GUARDABASSI, L., 2011: Rapid PCR Detection of Staphylococcus aureus Clonal Complex 398 by Targeting the Restriction-Modification System Carrying sau1-hsdS1, J. Clin. Microbiol. 49: 732-734. ISSN 0095-1137. DOI: 10.1128/JCM.01970-10 Go to original source...
  12. ©TĚPÁN, J., PANTŮČEK, R., DO©KAŘ, J., 2004: Molecular diagnostics of clinically important staphylococci. Folia Microbiol. 49: 353-386. ISSN 0015-5632. Go to original source...
  13. ©VEC, P., NOVÁKOVÁ, D., ®ÁČKOVÁ, L., KUKLETOVÁ, M., SEDLÁČEK, I., 2008: Evaluation of (GTG)5-PCR for rapid identification of Streptococcus mutans. Anton. Leeuw. Int. J. G. 94: 573-579. ISSN 0003-6072. DOI: 10.1007/s10482-008-9275-6 Go to original source...
  14. ©VEC, P., PANTŮČEK, R., PETRAS, P., SEDLÁČEK I., NOVÁKOVÁ D., 2010: Identification of Staphylococcus spp. using (GTG)(5)-PCR fingerprinting. Syst. Appl. Microbiol. 33: 451-456. ISSN 0723-2020. DOI: 10.1016/j.syapm.2010.09.004 Go to original source...
  15. TE WITT, R., KANHAI, V., VAN LEEUWEN, W. B., 2009: Comparison of the DiversiLabTM system, pulsed-field gel electrophoresis and multi-locus sequence typing for the characterization of epidemic reference MRSA strains. J. Microbiol. Methods 77: 130-133. ISSN 0167-7012. DOI: 10.1016/j.mimet.2009.01.009 Go to original source...
  16. VERSALOVIC, J., SCHNEIDER, M., DE BRUIJN, F. J., LUPSKI, J. R., 1994: Genomic fingerprinting of bacteria using repetitive sequence-based polymerase chain reaction. Methods Mol. Cell. Biol. 5: 25-40. ISSN 0898-7750.
  17. WILSON, L. A., SHARP, P. M., 2006: Enterobacterial repetitive intergenic consensus (ERIC) sequences in Escherichia coli: evolution and implications for ERIC-PCR. Mol. Biol. Evol. 23: 1156-1168. ISSN 0737-4038. DOI: 10.1093/molbev/msj125 Go to original source...

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