Osmotic Compounds Enhance Antibiotic Efficacy against Acinetobacter baumannii Biofilm Communities.

Biofilm-associated infections are a clinical challenge, in part because a hydrated matrix protects the bacterial community from antibiotics. Herein, we evaluated how different osmotic compounds (maltodextrin, sucrose, and polyethylene glycol [PEG]) enhance antibiotic efficacy against Acinetobacter baumannii biofilm communities. Established (24-h) test tube biofilms (strain ATCC 17978) were treated with osmotic compounds in the presence or absence of 10× the MIC of different antibiotics (50 µg/ml tobramycin, 20 µg/ml ciprofloxacin, 300 µg/ml chloramphenicol, 30 µg/ml nalidixic acid, or 100 µg/ml erythromycin). Combining antibiotics with hypertonic concentrations of the osmotic compounds for 24 h reduced the number of biofilm bacteria by 5 to 7 log (P < 0.05). Increasing concentrations of osmotic compounds improved the effect, but there was a trade-off with increasing solution viscosity, whereby low-molecular-mass compounds (sucrose, 400-Da PEG) worked better than higher-mass compounds (maltodextrin, 3,350-Da PEG). Ten other A. baumannii strains were similarly treated with 400-Da PEG and tobramycin, resulting in a mean 2.7-log reduction in recoverable bacteria compared with tobramycin treatment alone. Multivariate regression models with data from different osmotic compounds and nine antibiotics demonstrated that the benefit from combining hypertonic treatments with antibiotics is a function of antibiotic mass and lipophilicity (r2 > 0.82; P < 0.002), and the relationship was generalizable for biofilms formed by A. baumannii and Escherichia coli K-12. Augmenting topical antibiotic therapies with a low-mass hypertonic treatment may enhance the efficacy of antibiotics against wound biofilms, particularly when using low-mass hydrophilic antibiotics.IMPORTANCE Biofilms form a barrier that protects bacteria from environmental insults, including exposure to antibiotics. We demonstrated that multiple osmotic compounds can enhance antibiotic efficacy against Acinetobacter baumannii biofilm communities, but viscosity is a limiting factor, and the most effective compounds have lower molecular mass. The synergism between osmotic compounds and antibiotics is also dependent on the hydrophobicity and mass of the antibiotics. The statistical models presented herein provide a basis for predicting the optimal combination of osmotic compounds and antibiotics against surface biofilms communities.

Immunization with Anaplasma centrale Msp2 HVRs Is Less Effective than the Live A. centrale Vaccine against Anaplasmosis.

Bovine anaplasmosis, caused by Anaplasma marginale, is the most prevalent tick-transmitted pathogen of livestock globally. In many parts of the world, Anaplasma centrale, a related organism, is used as a live blood-borne vaccine as it causes either no or only a mild clinical disease. Anaplasma centrale does not prevent infection with A. marginale but does prevent acute disease. Anaplasma centrale is prohibited from being used in the U.S. due to the risk of transmitting emerging pathogens. Both of these organisms encode proteins known as major surface protein 2 (Msp2), which is the most immunodominant protein for the organism. Both organisms persist in their host by evading clearance, i.e., the adaptive immune response, by recombining the hypervariable region (HVR) of msp2 with pseudogene alleles. The study goal was to test whether the Msp2 HVRs encoded by A. centrale are a sufficient source of immune stimulation to provide the clinical protection exhibited by the blood-borne vaccine. Calves were inoculated with recombinantly expressed A. centrale HVRs. Control groups were inoculated with saponin or infected with the A. centrale live vaccine and compared with the test group. A Western blot analysis demonstrated that the HVR immunizations and A. centrale live vaccine stimulated an immune response. All animals in the study became infected upon challenge with A. marginale-infected ticks. The saponin-immunized control group had a high PPE (5.4%) and larger drops in PCVs (14.6%). As expected, the A. centrale-immunized animals were protected from acute disease with lower (0.6%) parasitemia and lower drops in PCV (8.6%). The HVR-immunized group had intermediate results that were not statistically significantly different from either the negative or positive controls. This suggests that the HVR immunogen does not fully recapitulate the protective capacity of the live vaccine.

Sequential Hypertonic-Hypotonic Treatment Enhances Efficacy of Antibiotic against Acinetobacter baumannii Biofilm Communities.

Infections with bacterial biofilm communities are highly tolerant of antibiotics. This protection is attributed, in part, to a hydrated extracellular polymeric substance (EPS) that surrounds the bacterial community and that limits antibiotic diffusion. In this study, we evaluated whether it is possible to dehydrate and then re-hydrate a biofilm as a means to increase antibiotic penetration and efficacy. Acinetobacter baumannii biofilms (24 h) were exposed to hypertonic concentrations of maltodextrin, sucrose or polyethylene glycol (PEG) as the dehydration step. These biofilms were then washed with deionized water containing 10 times the concentration of antibiotics needed to kill these bacteria in broth culture (50 µg/mL tobramycin, 300 µg/mL chloramphenicol, 20 µg/mL ciprofloxacin or 100 µg/mL erythromycin) as the rehydration step. Biofilms were then harvested, and the number of viable cells was determined. Sequential treatment with PEG and tobramycin reduced cell counts 4 to 7 log (p < 0.05) relative to combining PEG and tobramycin in a single treatment, and 3 to 7 log relative to tobramycin treatment alone (p < 0.05). Results were variable for other osmotic compounds and antibiotics depending on the concentrations used, likely related to mass and hydrophobicity. Our findings support future clinical evaluation of sequential regimens of hypertonic and hypotonic solutions to enhance antibiotic efficacy against chronic biofilm infections.

Electrochemical scaffold generates localized, low concentration of hydrogen peroxide that inhibits bacterial pathogens and biofilms.

We hypothesized that low concentrations of H2O2 could be generated through the electrochemical conversion of oxygen by applying an electric potential to a conductive scaffold and produce a low, but constant, concentration of H2O2 that would be sufficient to destroy biofilms. To test our hypothesis we used a multidrug-resistant Acinetobacter baumannii strain, because this species is often implicated in difficult-to-treat biofilm infections. We used conductive carbon fabric as the scaffold material ("e-scaffold"). In vitro experiments demonstrated the production of a maximum constant concentration of ~25 µM H2O2 near the e-scaffold surface. An e-scaffold was overlaid onto an existing A. baumannii biofilm, and within 24 h there was a ~4-log reduction in viable bacteria with an ~80% decrease in biofilm surface coverage. A similar procedure was used to overlay an e-scaffold onto an existing A. baumannii biofilm that was grown on a porcine explant. After 24 h, there was a ~3-log reduction in viable bacteria from the infected porcine explants with no observable damage to the underlying mammalian tissue based on a viability assay and histology. This research establishes a novel foundation for an alternative antibiotic-free wound dressing to eliminate biofilms.