Combined Bacteriophage and Antibiotic Treatment Prevents Pseudomonas aeruginosa Infection of Wild Type and cftr- Epithelial Cells

Abstract

With the increase of infections due to multidrug resistant bacterial pathogens and the shortage of antimicrobial molecules with novel targets, interest in bacteriophages as a therapeutic option has regained much attraction. Before the launch of future clinical trials, in vitro studies are required to better evaluate the efficacies and potential pitfalls of such therapies. Here we studied in an ex vivo human airway epithelial cell line model the efficacy of phage and ciprofloxacin alone and in combination to treat infection by Pseudomonas aeruginosa. The Calu-3 cell line and the isogenic CFTR knock down cell line (cftr-) infected apically with P. aeruginosa strain PAO1 showed a progressive reduction in transepithelial resistance during 24 h. Administration at 6 h p.i. of single phage, phage cocktails or ciprofloxacin alone prevented epithelial layer destruction at 24 h p.i. Bacterial regrowth, due to phage resistant mutants harboring mutations in LPS synthesis genes, occurred thereafter both in vitro and ex vivo. However, coadministration of two phages combined with ciprofloxacin efficiently prevented PAO1 regrowth and maintained epithelial cell integrity at 72 p.i. The phage/ciprofloxacin treatment did not induce an inflammatory response in the tested cell lines as determined by nanoString R gene expression analysis. We conclude that combination of phage and ciprofloxacin efficiently protects wild type and cftr-epithelial cells from infection by P. aeruginosa and emergence of phage resistant mutants without inducing an inflammatory response. Hence, phage-antibiotic combination should be a safe and promising anti-Pseudomonas therapy for future clinical trials potentially including cystic fibrosis patients.

Friends or Foes? Rapid Determination of Dissimilar Colistin and Ciprofloxacin Antagonism of Pseudomonas aeruginosa Phages

“…Luscher et al [159] treated 6 h P. aeruginosa PAO1 infections of Calu-3 epithelial cell lines in minimal essential medium with phages in the presence of 16× MIC ciprofloxacin (4.0 µg/mL, which roughly corresponds to a maximum serum concentration of 750 mg dose of ciprofloxacin in the clinic, 4.3 µg/mL, and which corresponds to 14× MIC in our study). After 72 h post infection, they observed substantial bacteria-killing and multiple log increases in PFUs.…”

In vitroanalysis of colistin and ciprofloxacin antagonism ofPseudomonas aeruginosaphage PEV2 infection activities

“…Luscher et al 107 treated 6 h P. aeruginosa PAO1 infections of Calu-3 epithelial cell lines in minimal essential medium with dual phage treatment in the presence of 16× MIC ciprofloxacin (4.0 μg/ml, which roughly corresponds to maximum serum concentration of 750 mg dose of ciprofloxacin in clinic, 4.3 μg/ml, and corresponds with 14× MIC in our study). After 72h post infection they observed substantial bacteria killing and multiple log increases in PFUs.…”

A Method to Determine the Efficacy of a Commercial Phage Preparation against Uropathogens in Urine and Artificial Urine Determined by Isothermal Microcalorimetry

“…Considering the different studies on rapid drug susceptibility testing using isothermal microcalorimetry [ 46 ] and previous studies with phages and viruses [ 29 , 30 , 31 , 32 ], we believe that providing a phagogram or a combined phagogram/antibiogram within 5 to 8 h is possible using patient urine directly, as UTI are often characterized by a high pathogen load with only rare polymicrobial infections. This would be of great interest, as it is expected to prevent the risk of mutant appearance [ 47 , 48 , 49 ]. In order to further speed up the process, isothermal microcalorimetry could be combined with a time-of-flight mass spectrometer MALDI-TOF [ 46 , 50 , 51 ] for preliminary identification of the pathogen and thus narrowing the range of phage product to be tested.…”