Skip to main content

Resistance to antimicrobials and biofilm formation in Staphylococcus spp. isolated from bovine mastitis in the Northeast of Brazil

Tropical Animal Health and Production volume 47, pages 511–518 (2015)Cite this article

Abstract

Mastitis is the principal disease affecting dairy herds worldwide. The aim of the present study was to characterize phenotypic and genotypic features associated with resistance to antimicrobials in Staphylococcus spp. isolated from 2064 milk samples of 525 lactating cows in the Northeast of Brazil. Of the 218 isolates analyzed, 57.8 % were characterized as Staphylococcus aureus, 28 % as coagulase-positive staphylococci other than S. aureus (oCPS), and 14.2 % as coagulase-negative staphylococci (CNS). The test for susceptibility to antimicrobials showed amoxicillin (32.6 %) to be the less effective drug in vitro, and the multi-drug resistance (MDR) rate for beta-lactams varied from 0 to 0.75. The genotypic characterization showed that 93.1 % of the samples were tested positive for the blaZ gene, while none amplified mecA. The antibiotic efflux mechanism was observed in 0.9 % of isolates. The biofilm formation was found in 3.7 and 96.3 % of samples, respectively, on Congo red agar and on the microplate adhesion test, while the icaD gene was present in 92.2 % of Staphylococcus spp. The high frequency of blaZ gene observed in this study was associated with the resistance of most Staphylococcus spp. to one or more of the beta-lactams tested, which are routinely used in Brazilian herds for mastitis treatment. The biofilm formation was also detected in the isolates analyzed being an important characteristic for pathogenicity and antimicrobial resistance of bacteria.

This is a preview of subscription content, access via your institution.

Access options

Buy single article

Instant access to the full article PDF.

43,34 €

Price includes VAT (Finland)
Tax calculation will be finalised during checkout.

References

  • Alian, F., Rahimi, E., Shakerian, A., Momtaz, H., Riahi, M. and Momeni, M., 2012. Antimicrobial resistance of Staphylococcus aureus isolated from bovine, sheep and goat raw milk, Global Veterinaria, 8, 111–114.

    Google Scholar 

  • Arciola, C.R., Baldassarri, L. and Montanaro, L., 2001. Presence of icaA and icaD genes and slime production in a collection of Staphylococcal strains from catheter-associated infections, Journal of Clinical Microbiology, 39, 2151–2156.

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Arslan, E., Çelebi, A., Açik, L. and Uçan, U.S., 2009. Characterisation of coagulase positive Staphylococcus species isolated from bovine mastitis using protein and plasmid patterns, Turkish Journal of Veterinary and Animal Sciences, 33, 493–500.

    CAS  Google Scholar 

  • Asfour, H.A.E. and Darwish, S.F., 2011. Phenotypic and genotypic detection of both mecA and blaZ genes mediated β-lactam resistance in Staphylococcus strains isolated from bovine mastitis, Global Veterinaria, 6, 39–50.

    CAS  Google Scholar 

  • Barkema, H.W., Schukken, Y.H. and Zadoks, R.N., 2006. Invited review: The role of cow, pathogen, and treatment regimen in the therapeutic success of bovine Staphylococcus aureus mastitis, Journal of Dairy Science, 89, 1877–1895.

    Article  CAS  PubMed  Google Scholar 

  • Bjorland, J., Steinum, T., Kvitle, B., Waage, S., Sunde, M. and Heir, E., 2005. Widespread distribution of disinfectant resistance genes among staphylococci of bovine and caprine origin in Norway, Journal of Clinical Microbiology, 43, 4363–4368.

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Bradley, A.J., Leach, K.A., Breen J.E., Green, L.E. and Green, M.J., 2007. Survey of the incidence and aetiology of mastitis on dairy farms in England and Wales, Veterinary Record, 160, 253–258.

    Article  CAS  PubMed  Google Scholar 

  • CLSI, 2008. Performance Standards for Antimicrobial Disk and Dilution Susceptibility Tests for Bacteria Isolated from Animals: Approved Standard - Third Edition, CLSI document M31-A3 (CLSI, Wayne).

  • CLSI, 2012. Performance Standards for Antimicrobial Susceptibility Testing: Twenty-Second Informational Supplement, CLSI document M100-S22 (CLSI, Wayne).

  • Costa, A.M., Kay, I. and Palladino, S., 2005. Rapid detection of mecA and nuc genes in staphylococci by real-time multiplex polymerase chain reaction, Diagnostic Microbiology and Infectious Disease, 51, 13–17.

    Article  CAS  PubMed  Google Scholar 

  • Cucarella, C., Tormo, M.A., Úbeda, C., Trotonda, M.P., Monzón, M., Peris, C., Amorena, B., Lasa, I. and Penadés, J.R., 2004. Role of biofilm-associated protein bap in the pathogenesis of bovine Staphylococcus aureus, Infection and Immunity, 72, 2177–2185.

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Gatermann, S.G., Koschinski, T. and Friedrich, S., 2007. Distribution and expression of macrolide resistance genes in coagulase-negative Staphylococci, Clinical Microbiology and Infection, 13, 777–781.

    Article  CAS  PubMed  Google Scholar 

  • Greco C., Mastronardi, C., Pagotto, F., Mack, D. and Ramirez-Arcos, S., 2008. Assessment of biofilm-forming ability of coagulase-negative staphylococci isolated from contaminated platelet preparations in Canada, Transfusion, 48, 969–977.

    CAS  PubMed  Google Scholar 

  • Guérin Faublée, V., Carret, G. and Houffschmitt, P., 2003. In vitro activity of 10 antimicrobial agents against bacteria isolated from cows with clinical mastitis, Veterinary Record, 152, 466–471.

    Article  PubMed  Google Scholar 

  • Haftu, R., Taddele, H., Gugsa, G. and Kalayou, S., 2012. Prevalence, bacterial causes, and antimicrobial susceptibility profile of mastitis isolates from cows in large-scale dairy farms of Northern Ethiopia. Tropical Animal Health and Production, 44, 1765–1771.

    Article  PubMed  Google Scholar 

  • Haveri, M., Suominen, S., Rantala, L., Honkanen-Buzalski, T. and Pyörälä, S., 2005. Comparison of phenotypic and genotypic detection of penicillin G resistance of Staphylococcus aureus isolated from bovine intramammary infection, Veterinary Microbiology, 106, 97–102.

    Article  CAS  PubMed  Google Scholar 

  • Kateete, D.P., Kimani, C.N., Katabazi, F.A., Okeng, A., Okee, M.S., Nanteza, A., Joloba, M.L. and Najjuca, F.C., 2010. Identification of Staphylococcus aureus: DNAse and mannitol salt agar improve the efficiency of the tube coagulase test, Annals of Clinical Microbiology and Antimicrobials, 9, 23.

    Article  PubMed Central  PubMed  Google Scholar 

  • Keane, O.M., Budd, K.E., Flynn, J. and McCoy, F., 2013. Pathogen profile of clinical mastitis in Irish milk-recording herds reveals a complex aetiology, The Veterinary Record, 173, 17.

    Article  CAS  PubMed  Google Scholar 

  • Knobloch, J.K.M., Horstkotte, M.A., Rohde, H. and Mack, D., 2002. Evaluation of different detection methods of biofilm formation in Staphylococcus aureus, Medical Microbiology and Immunology, 191, 101–106.

    Article  CAS  PubMed  Google Scholar 

  • Kumar, R., Yadav, B.R. and Singh, R.S., 2010. Genetic determinants of antibiotic resistance in Staphylococcus aureus isolates from milk of mastitic crossbred cattle, Current Microbiology, 60, 379–386.

    Article  CAS  PubMed  Google Scholar 

  • Lewis, K., 2001. Riddle of biofilm resistance, Antimicrobial Agents and Chemotherapy, 45, 999–1007.

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Loncarevic, S., Jorgensen, H.J., Lovseth, A., Mathisen, T. and Rorvik, L.M., 2005. Diversity of Staphylococcus aureus enterotoxin types within single samples of raw milk and raw milk products, Journal of Applied Microbiology, 98, 344–350.

    Article  CAS  PubMed  Google Scholar 

  • Medeiros, E.S., França, C.A., Krewer, C.C., Peixoto, R.M., Souza Júnior, A.F., Cavalcante, M.B., Costa, M.M. and Mota, R.A., 2011. Antimicrobial resistance of Staphylococcus spp. isolates from cases of mastitis in buffalo in Brazil, Journal of Veterinary Diagnostic Investigation, 23, 793–796.

    Article  PubMed  Google Scholar 

  • Melchior, M.B., Vaarkamp, H. and Fink-Gremmels, J., 2006. Biofilms: a role in recurrent mastitis infection?, Veterinary Journal, 171, 398–407.

    Article  CAS  Google Scholar 

  • Melchior, M.B., van Osch, M.H.J., Graat, R.M., van Duijkeren, E., Mevius, D.J., Nielen, M., Gaastra, W. and Fink-Gremmels, J., 2009. Biofilm formation and genotyping of Staphylococcus aureus bovine mastitis isolates: evidence for lack of penicillin-resistance in Agr-type II strains. Veterinary Microbiology, 137, 83–89.

    Article  CAS  PubMed  Google Scholar 

  • Merino, N., Toledo-Arana, A., Vergara-Irigaray, M., Valle, J., Solano, C., Calvo, E., Lopez, J.A., Foster, T.J., Penadés, J.R. and Lasa, I., 2009. Protein A-mediated multicellular behavior in Staphylococcus aureus, Journal of Bacteriology, 191, 832–843.

    Article  PubMed Central  CAS  PubMed  Google Scholar 

  • Moon, J.S., Lee, A.R., Kang, H.M., Lee, E.S., Kim, M.N., Paik, Y.H., Park, Y.H., Joo, Y.S. and Koo, H.C., 2007. Phenotypic and genetic antibiogram of methicillin-resistant Staphylococci isolated from bovine mastitis in Korea, Journal of Dairy Science, 90, 1176–1185.

    Article  CAS  PubMed  Google Scholar 

  • Murakami, K., Minamide, W., Wada, K., Nakamura, E., Teraoka, H. and Watanabe, S., 1991. Identification of methicillin-resistant strains of staphylococci by polymerase chain reaction, Journal of Clinical Microbiology, 29, 2240–2244.

    PubMed Central  CAS  PubMed  Google Scholar 

  • National Mastitis Council, 1999. Laboratory Handbook and Bovine Mastitis, (The National Mastitis Council, Arlington).

  • Olsen, J.E., Christensen, H., and Aarestrup, F.M., 2006. Diversity and evolution of blaZ from Staphylococcus aureus and coagulase-negative staphylococci, The Journal of Antimicrobial Chemotherapy, 57, 450–460.

    Article  CAS  PubMed  Google Scholar 

  • Olson, M.E., Ceri, H., Morck, D.W., Buret, A.G. and Read, R.R., 2002. Biofilm bacteria: formation and comparative susceptibility to antibiotics, Canadian Journal of Veterinary Research, 66, 86–92.

    PubMed Central  PubMed  Google Scholar 

  • Palomares, C., Torres, M.J., Torres, A., Aznar, J. and Palomares, J.C., 2003. Rapid detection and identification of Staphylococcus aureus from blood culture specimens using real-time fluorescence PCR, Diagnostic Microbiology and Infectious Disease, 45, 183–189.

    Article  CAS  PubMed  Google Scholar 

  • Quinn, P.J., Carter, M.E., Markey, B. and Carter, G.R., 1994. Clinical Veterinary Microbiology, (Wolfe, London).

    Google Scholar 

  • Rajala-Schultz, P.J., Smith, K.L., Hogan, J.S. and Love, B.C., 2004. Antimicrobial susceptibility of mastitis pathogens from first lactation and older cows, Veterinary Microbiology, 102, 33–42.

    Article  CAS  PubMed  Google Scholar 

  • Sawant, A.A., Gillespie, S.P. and Oliver, S.P., 2009. Antimicrobial susceptibility of coagulase-negative Staphylococcus species isolated from bovine milk, Veterinary Microbiology, 134, 73–81.

    Article  CAS  PubMed  Google Scholar 

  • Schalm, O.W. and Noorlander, D.O., 1957. Experiments and observations leading to development of the California mastitis test, Journal of the American Veterinary Medical Association, 130, 199–204.

    CAS  PubMed  Google Scholar 

  • Simojoki, H., Hyvönen, P., Ferrer, C.P., Taponen, S., and Pyörälä, S., 2012. Is the biofilm formation and slime producing ability of coagulase-negative staphylococci associated with the persistence and severity of intramammary infection?, Veterinary Microbiology, 158, 344–352.

    Article  CAS  PubMed  Google Scholar 

  • Sommerhäuser, J., Kloppert, B., Wolter, W., Zschöck, M., Sobiraj, A. and Failing, K., 2003. The epidemiology of Staphylococcus aureus infectious from subclinical mastitis in dairy cows during a control programme, Veterinary Microbiology, 96, 91–102.

    Article  PubMed  Google Scholar 

  • Stepanovic, S., Vukovic, D., Dakic, I., Savic, B. and Vlahovic, M.S., 2000. A modified microtiter-plate test for quantification of Staphylococcus biofilm formation, Journal of Microbiological Methods, 40, 175–179.

    Article  CAS  PubMed  Google Scholar 

  • Taponen, S. and Pyörälä, S., 2009. Coagulase-negative staphylococci as cause of bovine mastitis - not so different from Staphylococcus aureus?, Veterinary Microbiology, 134, 29–36.

    Article  CAS  PubMed  Google Scholar 

  • Vanderhaeghen, W., Cerpentier, T., Adriaensen, C., Vicca, J., Hermans, K., and Butaye, P., 2010. Methicillin-resistant Staphylococcus aureus (MRSA) ST398 associated with clinical and subclinical mastitis in Belgian cows, Veterinary Microbiology, 144, 166–171.

    Article  PubMed  Google Scholar 

  • Vasudevan, P., Nair, M.K., Annamalai, T. and Venkitanarayanan, K.S., 2003. Phenotypic and genotypic characterization of bovine mastitis isolates of Staphylococcus aureus for biofilm formation, Veterinary Microbiology, 92, 179–185.

    Article  CAS  PubMed  Google Scholar 

  • Vintov, J., Aarestrup, F.M., Zinn, C.E. and Olsen, J.E., 2003. Association between phage types and antimicrobial resistance among bovine Staphylococcus aureus from 10 countries, Veterinary Microbiology, 95, 133–147.

    Article  CAS  PubMed  Google Scholar 

  • Wade, K.A., Pounder, J.I., Cloud, J.L., Woods, G.L., 2005. Comparison of six methods of extracting Mycobacterium tuberculosis DNA from processed Sputum for testing by quantitative real-time PCR, Journal of Clinical Microbiology, 43, 2461–2473.

    Google Scholar 

  • Waller, K.P., Aspán, A., Nyman, A., Persson, Y. and Andersson, U.G., 2011. CNS species and antimicrobial resistance in clinical and subclinical bovine mastitis, Veterinary Microbiology, 152, 112–116.

    Article  PubMed  Google Scholar 

  • Webber, M.A. and Piddock, L.J., 2003. The importance of efflux pumps in bacterial antibiotic resistance, Journal Antimicrobial Chemotherapy, 51, 9–11.

    Article  CAS  Google Scholar 

Download references

Acknowledgments

This study was funded by the Fundação de Amparo à Ciência e Tecnologia do Estado de Pernambuco (FACEPE), Brazil (grant IBPG 0950-5.05/10).

Conflict of interest

The authors declare that they have no conflict of interest.

Author information

Affiliations

  1. Departamento de Medicina Veterinária, Universidade Federal Rural de Pernambuco, Rua Dom Manoel de Medeiros s/n, Dois Irmãos, 51171-900, Recife, Pernambuco, Brazil

    Carina da Costa Krewer & Rinaldo Aparecido Mota

  2. Laboratório de Microbiologia e Imunologia Animal, Universidade Federal do Vale do São Francisco, Rodovia BR-407 Km 12 Lote 543, Petrolina, 56300-990, Pernambuco, Brazil

    Evandro Santos Amanso, Gisele Veneroni Gouveia, Renata de Lima Souza & Mateus Matiuzzi da Costa

Authors
  1. Carina da Costa Krewer
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Evandro Santos Amanso
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Gisele Veneroni Gouveia
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Renata de Lima Souza
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Mateus Matiuzzi da Costa
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Rinaldo Aparecido Mota
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Carina da Costa Krewer.

Rights and permissions

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

da Costa Krewer, C., Santos Amanso, E., Veneroni Gouveia, G. et al. Resistance to antimicrobials and biofilm formation in Staphylococcus spp. isolated from bovine mastitis in the Northeast of Brazil. Trop Anim Health Prod 47, 511–518 (2015). https://doi.org/10.1007/s11250-014-0752-9

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s11250-014-0752-9

Keywords

  • Biofilm
  • Mastitis
  • Resistance
  • Staphylococcus spp.