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      Complete Killing of Agar Lawn Biofilms by Systematic Spacing of Antibiotic-Loaded Calcium Sulfate Beads

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          Abstract

          Background: Pseudomonas aeruginosa (PA) and Staphylococcus aureus (SA) are the major causative agents of acute and chronic infections. Antibiotic-loaded calcium sulfate beads (ALCSB) are used in the management of musculoskeletal infections such as periprosthetic joint infections (PJI). Methods: To determine whether the number and spatial distribution of ALCSB are important factors to totally eradicate biofilms, ALCSBs containing vancomycin and tobramycin were placed on 24 h agar lawn biofilms as a single bead in the center, or as 16 beads placed as four clusters of four, a ring around the edge and as a group in the center or 19 beads evenly across the plate. Bioluminescence was used to assess spatial metabolic activity in real time. Replica plating was used to assess viability. Results: For both strains antibiotics released from the beads completely killed biofilm bacteria in a zone immediately adjacent to each bead. However, for PA extended incubation revealed the emergence of resistant colony phenotypes between the zone of eradication and the background lawn. The rate of biofilm clearing was greater when the beads were distributed evenly over the plate. Conclusions: Both number and distribution pattern of ALCSB are important to ensure adequate coverage of antibiotics required to eradicate biofilms.

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          Most cited references 36

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          Microbial biofilms.

          Direct observations have clearly shown that biofilm bacteria predominate, numerically and metabolically, in virtually all nutrient-sufficient ecosystems. Therefore, these sessile organisms predominate in most of the environmental, industrial, and medical problems and processes of interest to microbiologists. If biofilm bacteria were simply planktonic cells that had adhered to a surface, this revelation would be unimportant, but they are demonstrably and profoundly different. We first noted that biofilm cells are at least 500 times more resistant to antibacterial agents. Now we have discovered that adhesion triggers the expression of a sigma factor that derepresses a large number of genes so that biofilm cells are clearly phenotypically distinct from their planktonic counterparts. Each biofilm bacterium lives in a customized microniche in a complex microbial community that has primitive homeostasis, a primitive circulatory system, and metabolic cooperativity, and each of these sessile cells reacts to its special environment so that it differs fundamentally from a planktonic cell of the same species.
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            Multidrug-resistance efflux pumps - not just for resistance.

             Laura Piddock (2006)
            It is well established that multidrug-resistance efflux pumps encoded by bacteria can confer clinically relevant resistance to antibiotics. It is now understood that these efflux pumps also have a physiological role(s). They can confer resistance to natural substances produced by the host, including bile, hormones and host-defence molecules. In addition, some efflux pumps of the resistance nodulation division (RND) family have been shown to have a role in the colonization and the persistence of bacteria in the host. Here, I present the accumulating evidence that multidrug-resistance efflux pumps have roles in bacterial pathogenicity and propose that these pumps therefore have greater clinical relevance than is usually attributed to them.
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              Persister cells and tolerance to antimicrobials.

              Bacterial populations produce persister cells that neither grow nor die in the presence of microbicidal antibiotics. Persisters are largely responsible for high levels of biofilm tolerance to antimicrobials, but virtually nothing was known about their biology. Tolerance of Escherichia coli to ampicillin and ofloxacin was tested at different growth stages to gain insight into the nature of persisters. The number of persisters did not change in lag or early exponential phase, and increased dramatically in mid-exponential phase. Similar dynamics were observed with Pseudomonas aeruginosa (ofloxacin) and Staphylococcus aureus (ciprofloxacin and penicillin). This shows that production of persisters depends on growth stage. Maintaining a culture of E. coli at early exponential phase by reinoculation eliminated persisters. This suggests that persisters are not at a particular stage in the cell cycle, neither are they defective cells nor cells created in response to antibiotics. Our data indicate that persisters are specialized survivor cells.
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                Author and article information

                Journal
                Materials (Basel)
                Materials (Basel)
                materials
                Materials
                MDPI
                1996-1944
                05 December 2019
                December 2019
                : 12
                : 24
                Affiliations
                [1 ]Department of Microbial Infection and Immunity, The Ohio State University, Wexner Medical Center, Columbus, OH 43210, USA; devendra.dusane@ 123456osumc.edu (D.H.D.); brooks.922@ 123456buckeyemail.osu.edu (J.R.B.); sindeldecker.3@ 123456osu.edu (D.S.); peters.690@ 123456osu.edu (C.W.P.); li.5960@ 123456osu.edu (A.L.); Nicholas.Farrar@ 123456osumc.edu (N.R.F.); Scott.Diamond@ 123456beaumont.org (S.M.D.); knechtc2@ 123456ccf.org (C.S.K.)
                [2 ]Division of Bacterial, Parasitic, and Allergenic Products, Center for Biologics Evaluation and Research, Food and Drug Administration, Silver Spring, MD 20993, USA; Roger.Plaut@ 123456fda.hhs.gov
                [3 ]Biocomposites Ltd., Keele Science Park, Keele, Staffordshire ST5 5NL, UK; cpd@ 123456biocomposites.com (C.D.); sa@ 123456biocomposites.com (S.S.A.); pl@ 123456biocomposites.com (P.A.L.)
                [4 ]Department of Orthopaedics, The Ohio State University, Wexner Medical Center, Columbus, OH 43210, USA; anne.sullivan@ 123456osumc.edu (A.S.); jgranger230@ 123456gmail.com (J.F.G.)
                [5 ]National Centre for Advanced Tribology at Southampton (nCATS) and National Biofilm Innovation Centre (NBIC), Department of Mechanical Engineering, University of Southampton, Southampton SO17 1BJ, UK
                Author notes
                [* ]Correspondence: paul.stoodley@ 123456osumc.edu ; Tel.: +1-614-292-7871
                [†]

                Department of Microbial Infection and Immunity, The Ohio State University, 760 Biomedical Research Tower, 460 West, 12th Avenue, Columbus, OH 43210, USA.

                Article
                materials-12-04052
                10.3390/ma12244052
                6947297
                31817373
                © 2019 by the authors.

                Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license ( http://creativecommons.org/licenses/by/4.0/).

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