Preliminary Application Of Bacteriophages Against Antibiotics-resistant Stenotrophomonas Maltophilia And Pseudomonas Aeruginosa

Purpose:Stenotrophomonas maltophilia is a common opportunistic pathogen which can colonise respiratory-tract epithelial cells and surfaces of medical devices, and this characteristic makes it a ready coloniser of hospitalized patients, and most of the infections are pneumonia. Pseudomonas aeruginosa is also a common conditioned pathogen. The intrinsic resistance to many antibiotics classes makes them great threats to human health, especially to the immunocompromised hosts.To tackle the serious issues of antibiotics resistance of Stenotrophomonas maltophilia and Pseudomonas aeruginosa, we successfully isolated two bacteriophage strains IME13and IME15using the antibiotics-resistant Stenotrophomonas maltophilia SMA2and bacteriophage Pa27P1using Pa44. We also analyzed their biological characteristics and established SMA2immune inhibition Balb/c mice lung infection and treatment model, Pseudomonas aeruginosa Pa44abdominal and lung infection model and its therapeutic models.Method:Using antibiotics-resistant Stenotrophomonas maltophilia SMA2and Pseudomonas aeruginosa Pa44as indicator bacteria, the bacteriophages were isolated using plaque method; the phage’s titer, optimal multiplicity of infection and one-step growth curve were measured by the double-layer plate culture method; phage genome was extracted using proteinase K/SDS method and the whole phage genome sequence were determined by the454sequencer.Infecting the Balb/c mice lung with minimal lethal dose of Stenotrophomonas maltophilia SMA2through nasal inhalation, then the infected mice were treated with Stenotrophomonas maltophilia bacteriophages IME13and IME15at different time points, and the mice mortality and bacteriophage treatment effect were evaluated. We used intraperitoneal injection and nasal drops to build mouse systemic infection model and pulmonary infection model of Pa44. And then we applied phage Pa27P1to treat the mice infected by Pa44, and observed the therapeutic effect.Result:Bacteriophages IME13and IME15were successfully isolated. IME13possessed unique plaque morphology and a surprisingly big burst size exceeding3000. IME15is the first T7-like Stenotrophomonas maltophilia bacteriophage. We also isolated approximately90Pseudomonas aeruginosa phages, and sreemed out five relatively broad spectrum phages Pa3Pl, Pa4P5, Pa12P1, Pa20P5, and Pa28P2, which when mixed together, could split all18Pseudomonas aeruginosa in vitro.The animal experiment indicated that bacteriophages IME13and IME15had good therapeutic effects on the pneumonia caused by Stenotrophomonas maltophilia infection, and phage Pa27P1also showed good results on Pa44systemically infected mice model and pulmonary infection mice model. The timely treatment after infection is meaningful to reduce the mice mortality.Conclusion:As the problem of antibiotics resistance of Stenotrophomonas maltophilia and Pseudomonas aeruginosa increases seriously, they do great harm to immunosuppressed patients, which makes it urgent to explore new treatment other than antibiotics. Our research showed that it was feasible to treat the bacterial infections by the phages. Thus the phage therapy could be expected to supplement the traditional antibiotics therapy.11figures,4tables,43references.