title: Comparison of methods to detect the in vitro activity of silver nanoparticles (AgNP) against multidrug resistant bacteria.
abstract: BACKGROUND
  Multidrug resistant microorganisms are a growing challenge and new substances that can be useful to treat infections due to these microorganisms are needed. Silver nanoparticle may be a future option for treatment of these infections, however, the methods described in vitro to evaluate the inhibitory effect are controversial.
  
  
  RESULTS
  This study evaluated the in vitro activity of silver nanoparticles against 36 susceptible and 54 multidrug resistant Gram-positive and Gram-negative bacteria from clinical sources. The multidrug resistant bacteria were oxacilin-resistant Staphylococcus aureus, vancomycin-resistant Enterococcus spp., carbapenem- and polymyxin B-resistant A. baumannii, carbapenem-resistant P. aeruginosa and carbapenem-resistant Enterobacteriaceae. We analyzed silver nanoparticles stabilized with citrate, chitosan and polyvinyl alcohol and commercial silver nanoparticle. Silver sulfadiazine and silver nitrate were used as control. Different methods were used: agar diffusion, minimum inhibitory concentration, minimum bactericidal concentration and time-kill. The activity of AgNPs using diffusion in solid media and the MIC methods showed similar effect against MDR and antimicrobial-susceptible isolates, with a higher effect against Gram-negative isolates. The better results were achieved with citrate and chitosan silver nanoparticle, both with MIC90 of 6.75 μg mL(-1), which can be due the lower stability of these particles and, consequently, release of Ag(+) ions as revealed by X-ray diffraction (XRD). The bactericidal effect was higher against antimicrobial-susceptible bacteria.
  
  
  CONCLUSIONS
  It seems that agar diffusion method can be used as screening test, minimum inhibitory concentration/minimum bactericidal concentration and time kill showed to be useful methods. The activity of commercial silver nanoparticle and silver controls did not exceed the activity of the citrate and chitosan silver nanoparticles. The in vitro inhibitory effect was stronger against Gram-negative than Gram-positive, and similar against multidrug resistant and susceptible bacteria, with best result achieved using citrate and chitosan silver nanoparticles. The bactericidal effect of silver nanoparticle may, in the future, be translated into important therapeutic and clinical options, especially considering the shortage of new antimicrobials against the emerging antimicrobial resistant microorganisms, in particular against Gram-negative bacteria.
authors.affiliation: Department of Infectious Diseases, University of São Paulo, São Paulo, Brazil. emerson_danguy@yahoo.com.br.
authors.forename: Emerson Danguy
authors.initials: ED
authors.lastname: Cavassin
authors.affiliation: Department of Surgery São Paulo, University of São Paulo, São Paulo, Brazil. costasilviaf@ig.com.br.
authors.forename: Luiz Francisco Poli
authors.initials: LF
authors.lastname: de Figueiredo
authors.affiliation: Department of Surgery São Paulo, University of São Paulo, São Paulo, Brazil. pinhata@usp.br.
authors.forename: José Pinhata
authors.initials: JP
authors.lastname: Otoch
authors.affiliation: Faculty of Engenier São Paulo, University of São Paulo, São Paulo, Brazil. marcelo.seckler@usp.br.
authors.forename: Marcelo Martins
authors.initials: MM
authors.lastname: Seckler
authors.affiliation: Faculty of Engenier São Paulo, University of São Paulo, São Paulo, Brazil. robertoangelo@usp.br.
authors.forename: Roberto Angelo
authors.initials: RA
authors.lastname: de Oliveira
authors.affiliation: Nanomedicine and Nanotoxicology Group, University of São Paulo, IFSC, São Paulo, Brazil. fabifantinelli@gmail.com.
authors.forename: Fabiane Fantinelli
authors.initials: FF
authors.lastname: Franco
authors.affiliation: Nanomedicine and Nanotoxicology Group, University of São Paulo, IFSC, São Paulo, Brazil. valeria.s.marangoni@gmail.com.
authors.forename: Valeria Spolon
authors.initials: VS
authors.lastname: Marangoni
authors.affiliation: Nanomedicine and Nanotoxicology Group, University of São Paulo, IFSC, São Paulo, Brazil. zuco@ifsc.usp.br.
authors.forename: Valtencir
authors.initials: V
authors.lastname: Zucolotto
authors.affiliation: Department of Infectious Diseases, University of São Paulo, São Paulo, Brazil. gcih.adm@hc.fm.usp.br.
authors.forename: Anna Sara Shafferman
authors.initials: AS
authors.lastname: Levin
authors.affiliation: Department of Infectious Diseases, University of São Paulo, São Paulo, Brazil. silviacosta@usp.br.|LIM-54 (Laboratório de Bacteriologia), Instituto de Medicina Tropical, Av. Dr. Enéas de Carvalho Aguiar, 470, Prédio II, 1º andar, Sala 112, Cerqueira César, São Paulo, SP, 054503-00, Brazil. silviacosta@usp.br.
authors.forename: Silvia Figueiredo
authors.initials: SF
authors.lastname: Costa
mesh_terms: D000900:Anti-Bacterial Agents / Q000737:chemistry / Q000494:pharmacology*; D001419:Bacteria / Q000166:cytology / Q000187:drug effects* / Q000254:growth & development; D001424:Bacterial Infections / Q000188:drug therapy* / Q000382:microbiology; D024901:Drug Resistance, Multiple, Bacterial; D006801:Humans; D053768:Metal Nanoparticles / Q000737:chemistry*; D008826:Microbial Sensitivity Tests / Q000379:methods*; D012834:Silver / Q000737:chemistry / Q000494:pharmacology*
chemical_list: D000900:Anti-Bacterial Agents; D012834:Silver