title: Staphylococcus epidermidis biofilms with higher proportions of dormant bacteria induce a lower activation of murine macrophages.
abstract: Staphylococcus epidermidis is an opportunistic pathogen and, due to its ability to establish biofilms, is a leading causative agent of indwelling medical device-associated infection. The presence of high amounts of dormant bacteria is a hallmark of biofilms, making them more tolerant to antimicrobials and to the host immune response. We observed that S. epidermidis biofilms grown in excess glucose accumulated high amounts of viable but non-culturable (VBNC) bacteria, as assessed by their low ratio of culturable bacteria over the number of viable bacteria. This effect, which was a consequence of the accumulation of acidic compounds due to glucose metabolism, was counteracted by high extracellular levels of calcium and magnesium added to the culture medium allowing modulation of the proportions of VBNC bacteria within S. epidermidis biofilms. Using bacterial inocula obtained from biofilms with high and low proportions of VBNC bacteria, their stimulatory effect on murine macrophages was evaluated in vitro and in vivo. The inoculum enriched in VBNC bacteria induced in vitro a lower production of tumour necrosis factor alpha, interleukin-1 and interleukin-6 by bone-marrow-derived murine macrophages and, in vivo, a lower stimulatory effect on peritoneal macrophages, assessed by increased surface expression of Gr1 and major histocompatibility complex class II molecules. Overall, these results show that environmental conditions, such as pH and extracellular levels of calcium and magnesium, can induce dormancy in S. epidermidis biofilms. Moreover, they show that bacterial suspensions enriched in dormant cells are less inflammatory, suggesting that dormancy can contribute to the immune evasion of biofilms.
authors.affiliation: IBMC - Instituto de Biologia Molecular e Celular, Rua do Campo Alegre 83, Porto, Portugal.|ICBAS - Instituto de Ciências Biomédicas de Abel Salazar, Largo do Professor Abel Salazar 2, 4099-003 Porto, Portugal.
authors.forename: Filipe
authors.initials: F
authors.lastname: Cerca
authors.affiliation: ICBAS - Instituto de Ciências Biomédicas de Abel Salazar, Largo do Professor Abel Salazar 2, 4099-003 Porto, Portugal.
authors.forename: Filipa
authors.initials: F
authors.lastname: Andrade
authors.affiliation: CEB-IBB, Campus de Gualtar, Universidade do Minho, 4710-057 Braga, Portugal.|ICBAS - Instituto de Ciências Biomédicas de Abel Salazar, Largo do Professor Abel Salazar 2, 4099-003 Porto, Portugal.
authors.forename: Ângela
authors.initials: Â
authors.lastname: França
authors.affiliation: IBMC - Instituto de Biologia Molecular e Celular, Rua do Campo Alegre 83, Porto, Portugal.|ICBAS - Instituto de Ciências Biomédicas de Abel Salazar, Largo do Professor Abel Salazar 2, 4099-003 Porto, Portugal.
authors.forename: Elva Bonifácio
authors.initials: EB
authors.lastname: Andrade
authors.affiliation: ICBAS - Instituto de Ciências Biomédicas de Abel Salazar, Largo do Professor Abel Salazar 2, 4099-003 Porto, Portugal.
authors.forename: Adília
authors.initials: A
authors.lastname: Ribeiro
authors.affiliation: REQUIMTE, Departamento de Química-Física, Faculdade de Farmácia, Universidade do Porto, Rua Aníbal Cunha 164, 4099-030 Porto, Portugal.
authors.forename: Agostinho A
authors.initials: AA
authors.lastname: Almeida
authors.affiliation: CEB-IBB, Campus de Gualtar, Universidade do Minho, 4710-057 Braga, Portugal.
authors.forename: Nuno
authors.initials: N
authors.lastname: Cerca
authors.affiliation: Channing Laboratory, Department of Medicine, Brigham and Women's Hospital, Boston, MA 02115, USA.
authors.forename: Gerald
authors.initials: G
authors.lastname: Pier
authors.affiliation: CEB-IBB, Campus de Gualtar, Universidade do Minho, 4710-057 Braga, Portugal.
authors.forename: Joana
authors.initials: J
authors.lastname: Azeredo
authors.affiliation: IBMC - Instituto de Biologia Molecular e Celular, Rua do Campo Alegre 83, Porto, Portugal.|ICBAS - Instituto de Ciências Biomédicas de Abel Salazar, Largo do Professor Abel Salazar 2, 4099-003 Porto, Portugal.
authors.forename: Manuel
authors.initials: M
authors.lastname: Vilanova
mesh_terms: D000818:Animals; D001422:Bacterial Adhesion; D018441:Biofilms / Q000254:growth & development*; D001854:Bone Marrow Cells / Q000378:metabolism; D002118:Calcium / Q000378:metabolism / Q000494:pharmacology; D018797:Cell Cycle Proteins / Q000096:biosynthesis; D002478:Cells, Cultured; D005947:Glucose / Q000378:metabolism; D000949:Histocompatibility Antigens Class II / Q000096:biosynthesis; D006863:Hydrogen-Ion Concentration; D007375:Interleukin-1 / Q000096:biosynthesis; D015850:Interleukin-6 / Q000096:biosynthesis; D008262:Macrophage Activation; D008264:Macrophages / Q000378:metabolism* / Q000382:microbiology; D008274:Magnesium / Q000378:metabolism / Q000494:pharmacology; D051379:Mice; D008807:Mice, Inbred BALB C; D013212:Staphylococcus epidermidis / Q000254:growth & development / Q000472:pathogenicity / Q000502:physiology*; D014409:Tumor Necrosis Factor-alpha / Q000096:biosynthesis
chemical_list: D018797:Cell Cycle Proteins; D000949:Histocompatibility Antigens Class II; D007375:Interleukin-1; D015850:Interleukin-6; D014409:Tumor Necrosis Factor-alpha; D008274:Magnesium; D005947:Glucose; D002118:Calcium