Presurgical acetone and chlorhexidine aseptic preparation of the equine hoof

Presurgical acetone and chlorhexidine aseptic preparation of the equine hoof lowers bacterial counts below surgically acceptable threshold

J Am Vet Med Assoc. 2026 Apr 15;264(9):1-8. doi: 10.2460/javma.25.12.0822. Print 2026 Sep 1.
Authors
Gabriel A Gonzalez, Caitlin M O’Shea, Ben Nabors, Jake Eldridge, Robert Wills, Natalie Stilwell

PMID: 41985521
DOI: 10.2460/javma.25.12.0822

Free article
Abstract

Objective: The objectives were to determine whether topical applications of acetone and 4% chlorhexidine following removal of the superficial hoof wall reduces tissue bacterial load below 5 log10 microorganisms per gram (CFUs/g) of tissue and whether Dey-Engley neutralizing broth (DENB) neutralizes residual chlorhexidine in vitro within equine hoof tissue samples.

Methods: In a clinical prospective study with a total of 24 hooves, the superficial hoof was mechanically removed followed by disinfection with acetone and 5-minute rough and aseptic preparations with 4% chlorhexidine. Dorsal wall, frog, and sole samples were collected aseptically before and after disinfection, minced, incubated in DENB or saline, serially diluted, and plated on blood agar for 48-hour aerobic bacterial colony counts. Linear and logistic regression mixed models assessed the effects of disinfection and neutralization on log10 CFUs/g of tissue for each site.

Results: Acetone and chlorhexidine reduced tissue bacterial load when adjusted for site and site-disinfection interaction. Adjusted log10 mean CFUs/g of tissue ranged from 3.24 to 4.76 for sites, and the proportion of samples below the log10 CFUs/g threshold of 5 was 66.7% (dorsal), 62.5% (frog), and 25% (sole). Hoof samples incubated in DENB or saline yielded similar CFUs/g of tissue values before disinfection, but DENB-incubated samples had higher CFUs/g of tissue values than non-neutralized samples following disinfection when adjusted for site and site-DENB interaction.

Conclusions: This protocol significantly reduced hoof bacterial loads in many samples, and DENB was an effective chlorhexidine-neutralizing agent.

Clinical relevance: Based on aerobic colony counts, the chlorhexidine/acetone protocol reduces hoof bacterial loads.

Keywords: DENB; chlorhexidine; disinfection; equine; hoof.