Eradication of Staphylococcus aureus Biofilm Infection by Persister Drug Combination.

Staphylococcus aureus can cause a variety of infections, including persistent biofilm infections, which are difficult to eradicate with current antibiotic treatments. Here, we demonstrate that combining drugs that have robust anti-persister activity, such as clinafloxacin or oritavancin, in combination with drugs that have high activity against growing bacteria, such as vancomycin or meropenem, could completely eradicate S. aureus biofilm bacteria in vitro. In contrast, single or two drugs, including the current treatment doxycycline plus rifampin for persistent S. aureus infection, failed to kill all biofilm bacteria in vitro. In a chronic persistent skin infection mouse model, we showed that the drug combination clinafloxacin + meropenem + daptomycin which killed all biofilm bacteria in vitro completely eradicated S. aureus biofilm infection in mice while the current treatments failed to do so. The complete eradication of biofilm bacteria is attributed to the unique high anti-persister activity of clinafloxacin, which could not be replaced by other fluoroquinolones including moxifloxacin, levofloxacin, or ciprofloxacin. We also compared our persister drug combination with the current approaches for treating persistent infections, including gentamicin + fructose and ADEP4 + rifampin in the S. aureus biofilm infection mouse model, and found neither treatment could eradicate the biofilm infection. Our study demonstrates an important treatment principle, the Yin-Yang model, for persistent infections by targeting both growing and non-growing heterogeneous bacterial populations, utilizing persister drugs for the more effective eradication of persistent and biofilm infections. Our findings have implications for the improved treatment of other persistent and biofilm infections in general.

Bactericidal Efficacy of High Irradiance Ultraviolet A Photoactivation of Riboflavin Versus Standard Corneal Cross-Linking Protocol In Vitro.

PURPOSE: The purpose of this study was to compare the efficacy of high ultraviolet A (UVA) irradiance photoactivation of riboflavin (vitamin B2) versus the standard corneal cross-linking protocol on bacterial viability. METHODS: Methicillin-sensitive Staphylococcus aureus (MSSA) Newman strain and methicillin-resistant multidrug-resistant S. aureus (MDR-MRSA) USA300, CA409, CA127, GA656, and NY315 strains were exposed to a UVA energy dose of 5.4 to 6 J/cm 2 by 2 high irradiance regimens: A) 30 mW/cm 2 for 3 minutes and B) 10 mW/cm 2 for 10 minutes with B2 0.1%. Control groups included B2/UVA alone, CA409 exposed to standard B2 0.1% + UVA (3 mW/cm 2 for 30 minutes), and an untreated sample. Cell viability was assessed. Triplicate values were obtained. The Mann-Whitney test and Student t test were used for statistical analysis. RESULTS: There was no difference comparing the median bacterial load (log CFU/mL) of the untreated samples versus regimen A: Newman P = 0.7, CA409 P = 0.3, USA300 P = 0.5, CA127 P = 0.6, GA656 P = 0.1, and NY315 P = 0.2 ( P ≥ 0.1); and B: Newman P = 0.1, CA409 P = 0.3, USA300 P = 0.4, CA127 P = 0.6, GA656 P = 0.1, and NY315 P = 0.3 ( P ≥ 0.1). Standard regimen killed 100% of CA409. CONCLUSIONS: Photoactivation of B2 by high UVA irradiance does not seem to be effective for bacterial eradication in this study.

Effect of topical bovine colostrum in wound healing of corneal surface after acute ocular alkali burn in mice.

The purpose of this study was to evaluate the effect of bovine colostrum (BC) in the regeneration of corneal epithelial cells on an ocular alkali burn model. Twenty-four C57BL/6 mice were categorized into two gender/age-matched groups for treatment. Two days after inducing a corneal alkali burn in all left eyes with 4 µl of sodium hydroxide 0.15 mol/l, both eyes of group 1 were treated with BC 4 times per day, and both eyes of group 2 were treated with isotonic saline solution (SS). The epithelial defect was photographed and measured by fluorescein staining on days two, four, seven, and ten. Ocular burn damage was assessed with a pre-established classification in clock hours from the limbus. After 10 days both eyes were processed, half of the group's corneas were assessed histopathologically, and the other half was used for pro/anti-inflammatory cytokine quantification using ELISA. BC treated (Group 1) corneas revealed significantly improved fluorescein staining score for limbal involvement when compared to SS treated (Group 2) corneas at days 4 (p = 0.013), 7 (p < 0.001), and 10 (p < 0.001), respectively. No differences were noted in limbal involvement at day 2 between the two groups (p > 0.99). The overall change (difference in slope) in fluorescein staining for limbal involvement between days 2 and 10 was -0.1669 (p = 0.006). Histologic examinations and cytokine measurements of group 2 demonstrated a strong inflammatory component compared to group 1. Our data indicates that topical application of BC facilitates corneal re-epithelialization and wound healing by suppressing the inflammatory process in an ocular alkali burn model.

Infection with persister forms of Staphylococcus aureus causes a persistent skin infection with more severe lesions in mice: failure to clear the infection by the current standard of care treatment.

Staphylococcus aureus can cause persistent infections and is known to develop persister cells in vitro. However, the in vivo significance of in vitro persisters in general is largely unclear. Here, we evaluated S. aureus stationary phase cultures and biofilm bacteria enriched in persister bacteria in comparison with actively growing log phase bacteria in terms of their ability to cause disease in a mouse skin infection model. We found that mice infected with the stationary phase and biofilm bacteria, which were enriched with persisters, produced more pronounced skin lesions that took longer to heal, and had more severe skin pathology and higher bacterial load than mice infected with log phase bacteria. Using our persistent infection mouse model, we showed that the clinically recommended treatment for recurrent S. aureus skin infection, doxycycline + rifampin, was not effective in eradicating the bacteria in mice. Analogous findings were observed in a Caenorhabditis elegans model, where stationary phase S. aureus caused greater virulence or mortality than log phase bacteria as early as two days post-infection. Our findings associate in vitro persisters and biofilm bacteria with more persistent and more severe infections and emphasize the importance of quality or metabolic status of the inoculum bacteria (persister bacteria versus growing bacteria) not just the number of bacteria in causing disease. The persistent infection mouse model we developed with persister inocula should have implications for understanding the process of disease establishment and pathogenesis, for developing persistent infection animal models, and for developing more effective treatments for chronic persistent infections in general.

Ocular Emergencies: Red Eye.

"Red eye" is used as a general term to describe irritated or bloodshot eyes. It is a recognizable sign of an acute/chronic, localized/systemic underlying inflammatory condition. Conjunctival injection is most commonly caused by dryness, allergy, visual fatigue, contact lens overwear, and local infections. In some instances, red eye can represent a true ocular emergency that should be treated by an ophthalmologist. A comprehensive assessment of red eye conditions is required to preserve the patients visual function. Severe ocular pain, significant photophobia, decreased vision, and history of ocular trauma are warning signs demanding immediate ophthalmological consultation.