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      Optimal environmental and culture conditions allow the in vitro coexistence of Pseudomonas aeruginosa and Staphylococcus aureus in stable biofilms

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          Abstract

          The coexistence between species that occurs in some infections remains hard to achieve in vitro since bacterial fitness differences eventually lead to a single organism dominating the mixed culture. Pseudomonas aeruginosa and Staphylococcus aureus are major pathogens found growing together in biofilms in disease-affected lungs or wounds. Herein, we tested and analyzed different culture media, additives and environmental conditions to support P. aeruginosa and S. aureus coexistence in vitro. We have unraveled the potential of DMEM to support the growth of these two organisms in mature cocultured biofilms (three days old) in an environment that dampens the pH rise. Our conditions use equal initial inoculation ratios of both strains and allow the stable formation of separate S. aureus microcolonies that grow embedded in a P. aeruginosa biofilm, as well as S. aureus biofilm overgrowth when bovine serum albumin is added to the system. Remarkably, we also found that S. aureus survival is strictly dependent on a well-characterized phenomenon of oxygen stratification present in the coculture biofilm. An analysis of differential tolerance to gentamicin and ciprofloxacin treatment, depending on whether P. aeruginosa and S. aureus were growing in mono- or coculture biofilms, was used to validate our in vitro coculture conditions.

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          Physiological heterogeneity in biofilms.

          Biofilms contain bacterial cells that are in a wide range of physiological states. Within a biofilm population, cells with diverse genotypes and phenotypes that express distinct metabolic pathways, stress responses and other specific biological activities are juxtaposed. The mechanisms that contribute to this genetic and physiological heterogeneity include microscale chemical gradients, adaptation to local environmental conditions, stochastic gene expression and the genotypic variation that occurs through mutation and selection. Here, we discuss the processes that generate chemical gradients in biofilms, the genetic and physiological responses of the bacteria as they adapt to these gradients and the techniques that can be used to visualize and measure the microscale physiological heterogeneities of bacteria in biofilms.
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            Molecular aspects of bacterial pH sensing and homeostasis.

            Diverse mechanisms for pH sensing and cytoplasmic pH homeostasis enable most bacteria to tolerate or grow at external pH values that are outside the cytoplasmic pH range they must maintain for growth. The most extreme cases are exemplified by the extremophiles that inhabit environments with a pH of below 3 or above 11. Here, we describe how recent insights into the structure and function of key molecules and their regulators reveal novel strategies of bacterial pH homeostasis. These insights may help us to target certain pathogens more accurately and to harness the capacities of environmental bacteria more efficiently.
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              Hydrogen Ion Buffers for Biological Research*

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                Author and article information

                Contributors
                mcendra@ibecbarcelona.eu
                etorrents@ibecbarcelona.eu
                Journal
                Sci Rep
                Sci Rep
                Scientific Reports
                Nature Publishing Group UK (London )
                2045-2322
                8 November 2019
                8 November 2019
                2019
                : 9
                : 16284
                Affiliations
                ISNI 0000 0004 0536 2369, GRID grid.424736.0, Bacterial Infections and Antimicrobial Therapies Group, , Institute for Bioengineering of Catalonia (IBEC), The Barcelona Institute of Science and Technology, ; Baldiri Reixac 15-21, 08028 Barcelona, Spain
                Author information
                http://orcid.org/0000-0002-3010-1609
                Article
                52726
                10.1038/s41598-019-52726-0
                6841682
                31705015
                c2a9bf93-a0bb-43ea-848a-7a51e8a6f588
                © The Author(s) 2019

                Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

                History
                : 17 June 2019
                : 22 October 2019
                Funding
                Funded by: FundRef https://doi.org/10.13039/501100010198, Ministerio de Economía, Industria y Competitividad, Gobierno de España (Ministerio de Economía, Industria y Competitividad);
                Award ID: RT12018-098573-B-100
                Award ID: RT12018-098573-B-100
                Award ID: RT12018-098573-B-100
                Award ID: RT12018-098573-B-100
                Award Recipient :
                Funded by: FundRef https://doi.org/10.13039/100010434, "la Caixa" Foundation (Caixa Foundation);
                Award ID: L16ALFQCAIXA
                Award Recipient :
                Funded by: FundRef https://doi.org/10.13039/501100002809, Generalitat de Catalunya (Government of Catalonia);
                Award ID: 2017 SGR01079
                Award Recipient :
                Funded by: FundRef https://doi.org/10.13039/501100003329, Ministerio de Economía y Competitividad (Ministry of Economy and Competitiveness);
                Award ID: BIO2015-63557-R
                Award Recipient :
                Categories
                Article
                Custom metadata
                © The Author(s) 2019

                Uncategorized
                applied microbiology,biofilms
                Uncategorized
                applied microbiology, biofilms

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