Attachment and optimization of Staphylococcus aureus, Staphylococcus epidermidis, and Pseudomonas aeruginosa biofilms to a 3D printed lattice.

A lattice was designed and fabricated using three-dimensional (3D) printing that allows for the facile transfer of biofilms formed from either Staphylococcus aureus, Staphylococcus epidermidis, or Pseudomonas aeruginosa into a fresh cell culture flask. To enhance biofilm production onto the filaments, three protein-based treatments were compared: fetal bovine serum (FBS), bovine serum albumin (BSA), and fibrinogen (Fb). Protein treatments included either supplementing the growth broths or pre-coating the lattice prior to immersion into the broth. S. aureus and P. aeruginosa biofilms were observed on all tested filaments that contained the supplement Fb. S. epidermidis required BSA to form biofilm. Ultimately, polycarbonate (PC) was chosen as the optimal material for lattice creation since it can be autoclaved without warping key design features. In addition, this 3D printed design may facilitate biofilm transfer from the bacterial culture to different cell culture platforms.

Antimicrobial Activity of Microgels with an Immobilized Copper(II) Complex Linked to Cross-Linking and Composition.

The resistance of many bacteria against currently available antimicrobial agents is increasing worldwide at an alarming pace. The described structure-activity relationship study was prompted by the extraordinary ability of water-dispersed microgels to hydrolyze glycosidic bonds similar to building blocks of the peptidoglycan layer of Gram-positive bacteria. The results establish polyacrylate microgels with embedded copper(II) complex as antimicrobial agents. The systematic study reveals that Staphylococcus aureus is susceptible to the microgels, while common commercial agents are found intermediate or resistant. In particular, a microgel with 60 mol % of cross-linking, Cu2LP60%, shows intriguing bactericidal activity at 1 µg/mL, while vancomycin requires a 4-fold higher dose, i.e., 4 µg/mL, for the same effect. The minimum inhibitory concentration of Cu2LP60% was determined as low as 0.64 µg/mL. Excellent stability of the poly(acrylate) microgels was observed by negative zeta potentials in nanopure water and aqueous sodium dodecyl sulfate solution. The composition of the microgel matrix with embedded binuclear metal complex was shown to be responsible for the antimicrobial activity, while the aqueous buffer-surfactant solution is not.

Microdialysis Sampling of Quorum Sensing Homoserine Lactones during Biofilm Formation.

Bacteria communicate chemically through a system called quorum sensing. In this work, microdialysis sampling procedures were optimized to collect quorum sensing molecules produced during in situ biofilm formation directly on the polymeric semipermeable membrane of the microdialysis probe. V. harveyi, a Gram-negative bacterium, was used as the model organism and releases variable chain length acylhomoserine lactones (AHLs) and acyl-oxohomoserine lactones (AOHLs) as signaling molecules during quorum sensing. Eliciting biofilm formation required coating fetal bovine serum onto the poly(ether sulfone) microdialysis membrane. Dialysates were collected in different experiments either during or after biofilm formation directly on a microdialysis probe. Continuous sampling of C4-AHL, C6-AHL, C8-AHL, C6-OXO-AHL, and C12-OXO-AHL was achieved over a period of up to 4 days. The AHLs and AOHLs in dialysates were concentrated with solid-phase extraction and quantified using LC-MS. Dialysate concentrations obtained for the AOHLs and AHLs ranged between 1 and 100 ppb (ng/mL) and varied between sampling days. This work demonstrates the initial use of microdialysis sampling to collect quorum sensing signaling chemicals during biofilm formation by a Gram-negative bacterial species.

Iloprost Affects Macrophage Activation and CCL2 Concentrations in a Microdialysis Model in Rats.

PURPOSE: The hypothesis that locally-released iloprost, a synthetic prostacyclin analog, affects macrophage phenotype at a microdialysis implant in the subcutaneous space of rats was tested. Macrophage activation towards alternatively-activated phenotypes using pharmaceutical release is of interest to improve integration of implants and direct the foreign body reaction toward a successful outcome. METHODS: Macrophage cell culture was used to test iloprost macrophage activation in vitro. Microdialysis sampling probes were implanted into the subcutaneous space of Sprague-Dawley rats to locally deliver iloprost in awake- and freely-moving rats. Monocyte chemoattractant protein -1 (CCL2) was quantified from collected dialysates using ELISA. Immunohistochemical staining was used to determine the presence of CD163+ macrophages in explanted tissues. RESULTS: Iloprost reduced CCL2 concentrations in NR8383 macrophages stimulated with lipopolysaccharide. CCL2 concentrations in collected dialysates were similarly reduced in the presence of iloprost. Iloprost caused an increase in CD163+ cells in explanted tissue surrounding implanted microdialysis probes at two days post probe implantation. CONCLUSIONS: Localized delivery of iloprost caused macrophage activation at the tissue interface of a microdialysis subcutaneous implant in rat. This model system may be useful for testing other potential macrophage modulators in vivo.