Effect of Fluopsin C from Pseudomonas aeruginosa LV strain on established biofilms of extensively drug-resistant Acinetobacter baumannii

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Acinetobacter baumannii is one of the critical pathogens for the development of new antibiotics according to World Health Organization, especially due to the crescent rates of antimicrobial resistance. Biofilm formation allows the attachment of A. baumannii to multiple types of surfaces, enhancing its survival and dissemination, while protects the sessile cells from inhibitory agents (disinfectants and antimicrobials). Development of new antimicrobials is urgent, and compounds that affect both planktonic and sessile cells are highly desirable. In this research, we aimed to determine the effect of Fluopsin C (FlpC), derived from Pseudomonas aeruginosa LV strain cultures, on the biomass of established biofilms of extensively drug-resistant A. baumannii (XDRAB). XDRAB CI 230 and CI 226 were recovered from tracheal secretion of inpatients of a public tertiary hospital in Londrina (Paraná, Brazil; 2019), and characterized as XDRAB according to their susceptibility profiles, determined by automatized systems. A. baumannii ATCC 19606 was used as the reference strain. FlpC was obtained from P. aeruginosa LV cultures in cupric nutrient broth (g/L: peptone 2; beef extract 1.2; CuCl2∙2H2O 0.03; pH = 6.8) kept at 28 °C/170 rpm for 8 days. The cultures were acidified (pH 4), centrifugated (9000 rpm at 4 °C/15 min), dried, and submitted to extraction with dichloromethane (2:1 v/v) (3x). FlpC was purified by two flash chromatography processes. Biofilms were formed in polystyrene 96-well plates using 200 μL of 106 CFU/mL cellular suspensions, incubated for 24 h/37 °C on tryptic soy broth plus 1% (w/v) glucose (TSBG). Wells were washed three times with NaCl 0.85% (w/v), aspirated, and then added with TSBG plus FlpC at five concentrations, ranging from 56 to 3.5 μg/mL. Treated cultures were incubated for 24 h/37 °C. Wells were washed and biofilms were fixed with methanol. Biofilm biomass was determined by reading the intensity of crystal violet staining (0.1% w/v; 15 min) in methanol at λ = 570 nm. 24 h-biofilms were extensively affected by FlpC exposure, in all concentrations (>70%). The best treatment was 3.5 μg/mL, achieving 95.10% (CI 230) and 85.88% (CI 226). In ATCC 19606 from 28 to 3.5 μg/mL, 100% of biofilm biomass was reduced, while 56 μg/mL achieved 88.59%. FlpC is a promissory candidate for drug development and here we describe its antibiofilm effect, also indicating its use as a disinfectant agent.

  • 1 Departamento de Microbiologia / Centro de Ciências Biológicas / Universidade Estadual de Londrina
  • 2 Departamento de Patologia, Análises Clínicas e Toxicológicas / Centro de Ciências da Saúde / Universidade Estadual de Londrina
  • Inovação e Biotecnologia
natural product
antibiofilm effect
established biofilms