title: Role of daptomycin in the induction and persistence of the viable but non-culturable state of Staphylococcus aureus biofilms.
abstract: We have recently demonstrated that antibiotic pressure can induce the viable but non-culturable (VBNC) state in Staphylococcus aureus biofilms. Since dormant bacterial cells can undermine anti-infective therapy, a greater understanding of the role of antibiotics of last resort, including daptomycin, is crucial. Methicillin-resistant S. aureus 10850 biofilms were maintained on non-nutrient (NN) agar in the presence or absence of the MIC of daptomycin until loss of culturability. Viable cells were monitored by epifluorescence microscopy and flow cytometry for 150 days. All biofilms reached non-culturability at 40 days and showed a similar amount of viable cells; however, in biofilms exposed to daptomycin, their number remained unchanged throughout the experiment, whereas in those maintained on NN agar alone, no viable cells were detected after 150 days. Gene expression assays showed that after achievement of non-culturability, 16S rDNA and mecA were expressed by all biofilms, whereas glt expression was found only in daptomycin-exposed biofilms. Our findings suggest that low daptomycin concentrations, such as those that are likely to obtain within biofilms, can influence the viability and gene expression of non-culturable S. aureus cells. Resuscitation experiments are needed to establish the VBNC state of daptomycin-exposed biofilms.
authors.affiliation: Department of Life and Environmental Sciences, Polytechnic University of Marche, Ancona 60131, Italy. s.pasquaroli@univpm.it.
authors.forename: Sonia
authors.initials: S
authors.lastname: Pasquaroli
authors.affiliation: Department of Biomolecular Sciences, Sect. Toxicological, Hygiene, and Environmental Sciences, University of Urbino Carlo Bo, Urbino 61029, Italy. barbara.citterio@uniurb.it.
authors.forename: Barbara
authors.initials: B
authors.lastname: Citterio
authors.affiliation: Department of Life and Environmental Sciences, Polytechnic University of Marche, Ancona 60131, Italy. andrix.di.cesare@alice.it.
authors.forename: Andrea Di
authors.initials: AD
authors.lastname: Cesare
authors.affiliation: Department of Life and Environmental Sciences, Polytechnic University of Marche, Ancona 60131, Italy. amiri.m1983@gmail.com.
authors.forename: Mehdi
authors.initials: M
authors.lastname: Amiri
authors.affiliation: Department of Earth, Life and Environmental Sciences, University of Urbino Carlo Bo, Urbino 61029, Italy. anita.manti@uniurb.it.
authors.forename: Anita
authors.initials: A
authors.lastname: Manti
authors.affiliation: Microbial Biofilm Laboratory, IRCCS Fondazione Santa Lucia, Rome 00179, Italy. c.vuotto@hsantalucia.it.
authors.forename: Claudia
authors.initials: C
authors.lastname: Vuotto
authors.affiliation: Department of Life and Environmental Sciences, Polytechnic University of Marche, Ancona 60131, Italy. f.biavasco@univpm.it.
authors.forename: Francesca
authors.initials: F
authors.lastname: Biavasco