Does Hamamelitannin Disrupt Biofilm Development by Common Orthopaedic Pathogens?

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Authors: Wood W, McLaren A, Calara F, McLemore R
Banner Good Samaritan Medical Center, Phoenix, AZ

Title: Does Hamamelitannin Disrupt Biofilm Development by Common Orthopaedic Pathogens?

Background: Bacteria in biofilms have decreased susceptibility to antimicrobials and the microbial pathogens are have deceased host immune response, leading to resistance to treatment. Quorum Sensing Inhibitors (QSI) are intended to disrupt biofilm formation. Hamamelitannin (HAM) is a QSI with limited data on effectiveness for disruption of biofilm formation or recognized orthopaedic pathogens.

Hypothesis/Purpose: Does HAM disrupt the growth of biofilms by common orthopaedic pathogens?

Methods: S aureus (ATCC 49230, UAMS-1) and S epidermidis (ATCC 35984) were grown in Tryptic Soy Broth (TSB) supplemented to 1% glucose in 96 well tissue culture plates. Plates were seeded in groups of 8 wells with 1/100 dilution of a turbid overnight culture, and total volume of 200µL per well. Groups included: media (control-), bacteria (control+), bacteria with 4g/mL gentamicin added at 0hrs or 24hrs, bacteria with 60µg/mL HAM at 0hrs with or without 4µg/mL gentamicin at 24hrs. Plates were stained with 0.1% Crystal Violet and imaged 48hrs after initial seeding (Fig 1).. Crystal Violet was solubilized with 30% acetic acid solution and absorbance was measured at 600nm to quantify biofilm growth. Differences between groups were determined with t-test.

Results: Biofilm associated with staph epidermidis was greater than biofilm formation by S aureus (p < 0.001). Biofilm growth by S aureus in the presence of HAM was not reduced (p >0.05). Biofilm growth by staph epidermidis in the presence of HAM was not reduced (p>0.05). Gentamicin did not have a synergistic effect with HAM (p>0.05).

Discussion: S epidermidis is more robust at creating biofilm at 48 hours than S aureus. HAM at previously described concentration is not an effective QSI lacking a consistent or meaningful effect on disrupting biofilm formation of orthopaedic pathogens. Growth and biofilm formation in the presence of 4µg/mL gentamicin is consistent with the MIC for these strains in biofilm being higher than 4µg/mL. Higher concentrations of HAM must be studied before HAM can be dismissed as a QSI.

Conclusion: S aureus and S epidermidis biofilm growth is not altered by 60µg/mL hamamelitannin.

Does Hamamelitannin Disrupt Biofilm Development by Common Orthopaedic Pathogens?