In Vitro Efficacy of Nonantibiotic Agents Against Bacterial Biofilm From Equine Uterine Isolates

To effectively degrade and kill bacteria in a biofilm state, accurately identifying the causative organism is important. No single agent tested in vitro was effective against all isolates of the four most common equine uterine pathogens. 

1. Introduction

Biofilm has been implicated in cases of chronic or recurrent endometritis. However, it is unclear if currently recommended therapies are effective against pathogens from the equine reproductive tract.

2. Materials and Methods

Equine uterine pathogens were utilized to form a biofilm in vitro and challenged for 6 hours with 9 nonantibiotic agents (Table 1) at concentrations suggested in equine and/or human medicine. Endpoints evaluated were disruption of the biofilm and killing of the bacteria. Positive controls were included in each replicate.

3. Results

Escherichia coli (N = 10) biofilm was significantly (P < 0.05) degraded when exposed to Tris ethylenediaminetetraacetic acid (EDTA), 2-amino-2-hydroxymethyl-1,3-propanediolandiol and disodium ethylenediaminetetraacetate dehydrate^a  , acetylcysteine, dimethyl sulfoxide, and antimicrobial peptide mimic^b  , and the bacterial load was significantly reduced when treated with acetylcysteine or hydrogen peroxide. Klebsiella pneumoniae (N = 10) biofilm was significantly reduced in mass (90%) after treatment with antimicrobial peptide mimicb , and the number of bacteria was significantly reduced with hydrogen peroxide. Biofilm preformed with Pseudomonas aeruginosa (N = 10) was significantly reduced in 50% of isolates when treated with either hydrogen peroxide or antimicrobial peptide mimic^b  , and acetylcysteine significantly reduced the bacterial load. Biofilm preformed with Streptococcus zooepidemicus (N = 10) was significantly disrupted, and the bacterial load was reduced for all test agents except ozone. 
 

4. Discussion

Bacterial isolates recovered from the equine reproductive tract are capable of forming a biofilm in vitro. However, none of the routine non-antibiotic treatments currently used in clinical practice evaluated in these studies were capable of disrupting a preformed biofilm under the conditions tested in species of equine bacteria isolated from the uterus. Therefore, a critical need exists to identify the causative organism(s) of equine bacterial endometritis and determine which anti-biofilm treatments will be effective in vivo. None of the agents evaluated in this study were able to eradicate a biofilm in all species of bacteria tested.

Acknowledgments

This study was funded by the Grayson-Jockey Research Foundation and donations to the ERL Research Foundation.

Declaration of Ethics

The Authors declare that they have adhered to the Principles of Veterinary Medical Ethics of the AVMA.

Conflict of Interest

The Authors declare no conflicts of interest.

Footnotes

^a Tricide®, Sigma-Aldrich, St. Louis MO 63103. 

^bCeragyn®, Ceragyn LLC, Spanish Fork, Utah 84660.