Comparison of methods to evaluate bacterial contact-killing materials

Marieke van de lagemaat, Arjen Grotenhuis, Betsy van de Belt-Gritter, Steven Roest, Ton J. A. Loontjens, Henk J. Busscher, Henny C. van der Mei*, Yijin Ren

*Corresponding author for this work

Research output: Contribution to journalArticleAcademicpeer-review

40 Citations (Scopus)
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Cationic surfaces with alkylated quaternary-ammonium groups kill adhering bacteria upon contact by membrane disruption and are considered increasingly promising as a non-antibiotic based way to eradicate bacteria adhering to surfaces. However, reliable in vitro evaluation methods for bacterial contact-killing surfaces do not yet exist. More importantly, results of different evaluation methods are often conflicting. Therefore, we compared five methods to evaluate contact-killing surfaces. To this end, we have copolymerized quaternary-ammonium groups into diurethane dimethacrylate/glycerol dimethacrylate (UDMA/GDMA) and determined contact-killing efficacies against five different Gram-positive and Gram-negative strains. Spray-coating bacteria from an aerosol onto contact-killing surfaces followed by air-drying as well as ASTM E2149-13a (American Society for Testing and Materials) were found unsuitable, while the Petrifilm (R) system and JIS Z 2801 (Japanese Industrial Standards) were found to be excellent methods to evaluate bacterial contact-killing surfaces. It is recommended however, that these methods be used in combination with a zone of inhibition on agar assay to exclude that leakage of antimicrobials from the material interferes with the contact-killing ability of the surface.

Statement of Significance

Bacterial adhesion to surfaces of biomaterials implants can be life-threatening. Antimicrobials to treat biomaterial-associated infections often fail due to the bacterial biofilm-mode-of-growth or are ineffective due to antibiotic-resistance of causative organisms. Positively-charged, quaternized surfaces can kill bacteria upon contact and are promising as a non-antibiotic-based treatment of biomaterial-associated infections. Reliable methods to determine efficacies of contact-killing surfaces are lacking, however. Here, we show that three out of five methods compared, including an established ASTM, are unsuitable. Methods found suitable should be used in combination with a zone-of-inhibition-assay to establish absence of antimicrobial leaching, potentially interfering with contact-killing. Identification of suitable assays for evaluating bacterial contact-killing will greatly assist this emerging field as an alternative for antibiotic-based treatment of biomaterial-associated-infections. (C) 2017 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Original languageEnglish
Pages (from-to)139-147
Number of pages9
JournalActa Biomaterialia
Publication statusPublished - 1-Sep-2017


  • Quaternary ammonium compounds
  • Positively-charged surface
  • JIS Z 2801
  • Leaching coating
  • Bacterial contact-killing surface
  • Antibiotic resistance

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