Bactericidal and Anti-biofilm Effects of Polyhexamethylene Biguanide in Models of Intracellular and Biofilm of Staphylococcus aureus Isolated from Bovine Mastitis

Abstract

Staphylococcus aureus infection is a common cause of mastitis, reducing milk yield, affecting animal welfare and causing huge economic losses within the dairy industry. In addition to the problem of acquired drug resistance, bacterial invasion into udder cells and the formation of surface biofilms are believed to reduce antibiotic efficacy, leading to treatment failure. Here, we investigated the antimicrobial activities of enrofloxacin, an antibiotic that is commonly used in mastitis therapy and polyhexamethylene biguanide (PHMB), an antimicrobial polymer. The antimicrobial activities were tested against intracellular S. aureus in infected Mac-T cells (host cells). Also, fluorescein-tagged PHMB was used to study PHMB uptake and localization with S. aureus within the infected Mac-T cells. Anti-biofilm activities were tested by treating S. aureus biofilms and measuring effects on biofilm mass in vitro. Enrofloxacin and PHMB at 15 mg/L killed between 42 to 92 and 99.9% of intracellular S. aureus, respectively. PHMB-FITC entered and colocalized with the intracellular S. aureus, suggesting direct interaction of the drug with the bacteria inside the host cells. Enrofloxacin and PHMB at 15 mg/L reduced between 10 to 27% and 28 to 37% of biofilms’ mass, respectively. The half-maximal inhibitory concentrations (IC50) obtained from a cytotoxicity assay were 345 ± 91 and 21 ± 2 mg/L for enrofloxacin and PHMB, respectively; therefore, both compounds were tolerated by the host cells at high concentrations. These findings suggest that both antimicrobials are effective against intracellular S. aureus and can disrupt biofilm structures, with PHMB being more potent against intracellular S. aureus, highlighting the potential application of PHMB in mastitis therapy.

Potential of Novel Bacterial Cellulose Dressings Chemisorbed with Antiseptics for the Treatment of Oral Biofilm Infections

“…Due to antibiotic inefficiency in the context of persistent biofilm infections, the remaining therapeutic option is the local application of antiseptic agents, which display strong, unspecific mechanisms of antimicrobial and antibiofilm action [16]. The antiseptics broadly used against bacterial biofilms are, among others, povidone-iodine, polihexanide, and chlorhexidine [17][18][19]. However, the latter one, due to a quickly growing bulk of evidence indicating increasing microbial cross-resistance following application [20][21][22], is being substituted with other antiseptic substances.…”

Polyhexamethylene Biguanide and Nadifloxacin Self-Assembled Nanoparticles: Antimicrobial Effects against Intracellular Methicillin-Resistant Staphylococcus aureus

“…To prepare the PHMB/nadifloxacin nanoparticles, both compounds were mixed in to a ratio of PHMB to nadifloxacin 1:2 ( w / w ). The ratio was determined based on the minimum inhibitory concentrations (MIC) of the individual compound against EMRSA-15, where the MIC of PHMB and nadifloxacin were 1 mg/L and 2 mg/L, respectively [33]. Formulation of PHMB and nadifloxacin produced nanoparticles with Z-average of 291.3 ± 89.6 nm (Figure 2a), with a polydispersity index (PDI) of 0.35 ± 0.04.…”

Antimicrobial Polymers: The Potential Replacement of Existing Antibiotics?

“…PHMB is particularly interesting for exhibiting broad spectrum activity against intracellular bacteria and parasites in nanoparticle form. Of note, PHMB demonstrates antimicrobial effects at low concentrations and has been shown to have lower toxicity than other agents of its class [20,150,155]. Future studies exploring the delivery options for PHMB, novel formulations with small molecule antibiotics, and interactions with ESKAPE isolates are required.…”