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Pseudomonas aeruginosa biofilm killing beyond the spacer by antibiotic-loaded calcium sulfate beads: an in vitro study.

ABSTRACT: Introduction: Bacterial biofilms are an important virulence factor in chronic periprosthetic joint infection (PJI) and other orthopedic infection since they are highly tolerant to antibiotics and host immunity. Antibiotics are mixed into carriers such as bone cement and calcium sulfate bone void fillers to achieve sustained high concentrations of antibiotics required to more effectively manage biofilm infections through local release. The effect of antibiotic diffusion from antibiotic-loaded calcium sulfate beads (ALCS-B) in combination with PMMA bone cement spacers on the spread and killing of Pseudomonas aeruginosa Xen41 (PA-Xen41) biofilm was investigated using a "large agar plate" model scaled for clinical relevance. Methods: Bioluminescent PA-Xen41 biofilms grown on discs of various orthopedic materials were placed within a large agar plate containing a PMMA full-size mock "spacer" unloaded or loaded with vancomycin and tobramycin, with or without ALCS-B. The amount of biofilm spread and log reduction on discs at varying distances from the spacer was assessed by bioluminescent imaging and viable cell counts. Results: For the unloaded spacer control, PA-Xen41 spread from the biofilm to cover the entire plate. The loaded spacer generated a 3 cm zone of inhibition and significantly reduced biofilm bacteria on the discs immediately adjacent to the spacer but low or zero reductions on those further away. The combination of ALCS-B and a loaded PMMA spacer greatly reduced bacterial spread and resulted in significantly greater biofilm reductions on discs at all distances from the spacer. Discussion: The addition of ALCS-B to an antibiotic-loaded spacer mimic increased the area of antibiotic coverage and efficacy against biofilm, suggesting that a combination of these depots may provide greater physical antibiotic coverage and more effective dead space management, particularly in zones where the spread of antibiotic is limited by diffusion (zones with little or no fluid motion).

SUBMITTER: Brooks JR

PROVIDER: S-EPMC8137860 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:BackgroundPseudomonas aeruginosa is a ubiquitous environmental microorganism and also a common cause of infection. Its ability to survive in many different environments and persistently colonize humans is linked to its presence in biofilms formed on indwelling device surfaces. Biofilm promotes adhesion to, and survival on surfaces, protects from desiccation and the actions of antibiotics and disinfectants.ResultsWe examined the genetic basis for biofilm production on polystyrene at room (22 °C) and body temperature (37 °C) within 280 P. aeruginosa. 193 isolates (69 %) produced more biofilm at 22 °C than at 37 °C. Using GWAS and pan-GWAS, we found a number of accessory genes significantly associated with greater biofilm production at 22 °C. Many of these are present on a 165 kb region containing genes for heavy metal resistance (arsenic, copper, mercury and cadmium), transcriptional regulators and methytransferases. We also discovered multiple core genome SNPs in the A-type flagellin gene and Type II secretion system gene xpsD. Analysis of biofilm production of isolates of the MDR ST111 and ST235 lineages on stainless-steel revealed several accessory genes associated with enhanced biofilm production. These include a putative translocase with homology to a Helicobacter pylori type IV secretion system protein, a TA system II toxin gene and the alginate biosynthesis gene algA, several transcriptional regulators and methytransferases as well as core SNPs in genes involved in quorum sensing and protein translocation.ConclusionsUsing genetic association approaches we discovered a number of accessory genes and core-genome SNPs that were associated with enhanced early biofilm formation at 22 °C compared to 37 °C. These included a 165 kb genomic island containing multiple heavy metal resistance genes, transcriptional regulators and methyltransferases. We hypothesize that this genomic island may be associated with overall genotypes that are environmentally adapted to survive at lower temperatures. Further work to examine their importance in, for example gene-knockout studies, are required to confirm their relevance. GWAS and pan-GWAS approaches have great potential as a first step in examining the genetic basis of novel bacterial phenotypes.

Project description:Resistance to conventional antibiotics is a growing public health concern that is quickly outpacing the development of new antibiotics. This has led the Infectious Diseases Society of America (IDSA) to designate Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, and Enterobacter species as "ESKAPE pathogens" on the basis of the rapidly decreasing availability of useful antibiotics. This emphasizes the urgent need for alternative therapeutic strategies to combat infections caused by these and other bacterial pathogens. In this study, we used Staphylococcus aureus (S. aureus) as a proof-of-principle ESKAPE pathogen to demonstrate that an appropriate antibiotic (daptomycin) can be incorporated into polydopamine-coated gold nanocages (AuNC@PDA) and that daptomycin-loaded AuNC@PDA can be conjugated to antibodies targeting a species-specific surface protein (staphylococcal protein A; Spa) as a means of achieving selective delivery of the nanoconstructs directly to the bacterial cell surface. Targeting specificity was confirmed by demonstrating a lack of binding to mammalian cells, reduced photothermal and antibiotic killing of the Spa-negative species Staphylococcus epidermidis, and reduced killing of S. aureus in the presence of unconjugated anti-Spa antibodies. We demonstrate that laser irradiation at levels within the current safety standard for use in humans can be used to achieve both a lethal photothermal effect and controlled release of the antibiotic, thus resulting in a degree of therapeutic synergy capable of eradicating viable S. aureus cells. The system was validated using planktonic bacterial cultures of both methicillin-sensitive and methicillin-resistant S. aureus strains and subsequently shown to be effective in the context of an established biofilm, thus indicating that this approach could be used to facilitate the effective treatment of intrinsically resistant biofilm infections.

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Pseudomonas aeruginosa biofilm killing beyond the spacer by antibiotic-loaded calcium sulfate beads: an in vitro study.

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