# Polyhexamethylene Biguanide Nanoparticles Inhibit Biofilm Formation by Mastitis-Causing Staphylococcus aureus

## TL;DR

This study explores the use of PHMB nanoparticles to inhibit biofilm formation by Staphylococcus aureus, a major cause of bovine mastitis, showing promising results for improved udder health in dairy cows.

## Contribution

The study introduces PHMB nanoparticles as a novel, low-toxicity approach to combat S. aureus biofilms in bovine mastitis.

## Key findings

- PHMB nanoparticles significantly reduced S. aureus biofilm formation by 50% at concentrations ≥1 µg/mL.
- PHMB nanoparticles showed low toxicity to bovine mammary cells at four times the effective concentration.
- In an excised teat model, PHMB nanoparticles reduced S. aureus by 37.57% compared to traditional disinfectants.

## Abstract

This manuscript addresses the persistent challenge of Staphylococcus aureus, a primary agent of bovine mastitis that compromises animal welfare and results in substantial economic losses in dairy production. The difficulty in treating S. aureus lies in its resistance to multiple antibiotics and its capacity to form resilient biofilms within mammary gland cells, which hampers the effectiveness of conventional therapies. In response, the study evaluates a novel strategy involving polyhexamethylene biguanide (PHMB) nanoparticles, known for their antimicrobial properties. Two main experiments were conducted. The first assessed the cytotoxicity of PHMB nanoparticles and their ability to disrupt S. aureus biofilms in vitro. The findings reveal that these nanoparticles were well tolerated by bovine mammary cells and significantly inhibited biofilm formation at effective concentrations. The second experiment compared PHMB nanoparticles to commonly used post-milking disinfectants. While the nanoparticles did not outperform traditional treatments, they still reduced bacterial load, with results possibly influenced by limited exposure time during application. Despite this, the study highlights the potential of PHMB nanoparticles as an alternative approach for controlling mastitis caused by S. aureus, especially in biofilm-associated infections. This innovative method may contribute to improved udder health and enhanced productivity in the dairy industry, warranting further investigation under practical conditions.

Staphylococcus aureus is a mastitis pathogen that compromises cow health and causes significant economic losses in the dairy industry. High antimicrobial resistance and biofilm formation by S. aureus limit the efficacy of conventional treatments. This study evaluated the potential of polyhexamethylene biguanide nanoparticles (PHMB NPs) against mastitis-causing S. aureus. PHMB NPs showed low toxicity to bovine mammary epithelial cells (MAC-T cells) at concentrations up to four times higher than the minimum inhibitory concentration (1 µg/mL) against S. aureus. In Experiment 1, PHMB NPs significantly reduced biofilm formation by S. aureus by 50% at concentrations ≥1 µg/mL, though they showed limited efficacy against preformed biofilms. In Experiment 2, using an excised teat model, PHMB NPs reduced S. aureus concentrations by 37.57% compared to conventional disinfectants (chlorhexidine gluconate, povidone–iodine, and sodium dichloroisocyanurate), though limited by short contact time. These findings highlight the potential of PHMB NPs for the control of S. aureus growth and biofilm formation.