Phage Therapy against Experimental Pseudomonas aeruginosa Infections in Mice and Drosophila melanogaster
- 주제(키워드) Pseudomonas aeruginosa , bacteriophage therapy
- 발행기관 서강대학교 대학원
- 지도교수 조유희
- 발행년도 2009
- 학위수여년월 2009. 2
- 학위명 석사
- 실제URI http://www.dcollection.net/handler/sogang/000000044859
- 본문언어 영어
초록/요약
최근 항생제 내성을 지닌 세균의 증가로 인해 Pseudomonas aeruginosa (녹농균) 의 감염에 대한 박테리오파아지 요법이 다시 각광받고 있다. 본 논문에서는 녹농균 PAO1 균주에 감염하여 크고 뚜렷한 plaque을 형성하는 두 가지의 새로운 용해성 파아지 (MPK1과 MPK6) 를 오물 환경에서 분리하였고, 이들이 PAO1 균주의 성장과 생물막 형성을 저해하는 것을 확인하였다. 형태적으로 MPK1은 Myoviridae에 속하고 MPK6는 Podoviridae에 속한다. 두 가지 파아지의 주된 수용체는 lipopolysaccharide의 B-polysaccharide로서 이는 두 가지 파아지의 숙주특이성이 녹농균 균주들의 혈청형과 연관될 것으로 생각된다. 두 파아지의 생체 밖에서의 살상 능력과 PAO1 균주에 의한 생쥐 복막염 모델을 이용한 파아지 요법의 효율성을 측정해 보았더니, 두 파아지의 근육 또는 복막 내 투여는 감염된 쥐의 생존율을 효과적으로 높이며 그 쥐들의 폐, 비장, 간장 내의 세균 수도 감소시키는 것을 알 수 있었다. Drosophila melanogaster (초파리) 전신 감염 모델에서는 파아지를 먹임으로써 파아지의 감염억제 효율을 측정해 보았다. 이러한 결과들을 종합해 볼 때, MPK1과 MPK6는 모두 녹농균에 의한 치명적인 감염을 효과적으로 제어함을 알 수 있었다.
more초록/요약
Bacteriophage therapy against Pseudomonas aeruginosa infections has received renewed attention owing to the increasing prevalence of multiple antibiotic-resistances in this bacterium. Here, I isolated and characterized two new lytic bacteriophages (MPK1 and MPK6) from sewage samples, which produced large, clear plaques on P. aeruginosa strain PAO1 and inhibited the planktonic growth and biofilm formation of PAO1. Based on their morphology, MPK1 belongs to the Myoviridae, while MPK6 belongs to the Podoviridae. The primary receptor for both phages is the B-polysaccharide of lipopolysaccharide, suggesting their hostspectra are associated with the serotypes of the P. aeruginosa strains. I evaluated the lytic activity of these phages in vitro and the efficacy of phage therapy using a murine model of peritoneal infections caused by PAO1. Both intraperitoneal and intramuscular administrations of MPK1, and to the less extent, MPK6 significantly protected infected mice from peritonitis-derived mortality. Mice treated with phages also had the lower bacterial loads in their lung, spleen and liver. The efficacy of both MPK1 and MPK6 was also evaluated by using Drosophila melanogaster systemic infection model, for which the phages were administered by feeding. Taken together, both MPK1 and MPK6 are effective against fatal infections caused by P. aeruginosa.
more목차
Introduction = 3
Materials and Methods = 8
Bacterial strains and media and culture conditions = 8
Preparation of phage lysates and phage particles = 8
Electron microscopy = 10
Biofilm experiments = 10
Phage infection = 11
Mouse experiments = 11
Determination of numbers of viable bacteria and phages in blood and organs = 12
Drosophila melanogaster experiments = 12
Results = 14
Identification of the new Caudovirales phage strains for P. aeruginosa = 14
Host spectra of the 6 lytic phages (MPK1 to MPK6) = 18
Receptor of phages MPK1 and MPK6 = 18
Lytic activity of phages MPK1 and MPK6 in vitro = 20
Pharmacokinetics of phage MPK1 and MPK6 in mouse = 23
Evaluation of therapeutic efficacy of MPK1 and MPK6 against mouse peritoneal infection = 26
Evaluation of therapeutic efficacy of MPK1 and MPK6 against D. melanogaster systemic infection = 29
Discussion = 34
References = 38
List of Tables
Table 1. Bacterial strains and plasmids used in this study = 9
Table 2. Killing spectra of MPK phages = 16
List of Figures
Fig. 1. Electron micrograph of phages MPK1 and MPK6 = 15
Fig. 2. Plaque formation by MPK1 and MPK6 on various lipopolysaccharide (LPS) mutants = 19
Fig. 3. In vitro growth of MPK1 and MPK6 = 21
Fig. 4. Effect of phage treatment on biofilm formation = 22
Fig. 5. Pharmacokinetics of MPK1 and MPK6 in noninfected mice = 24
Fig. 6. Protective effect of MPK1 and MPK6 on the survival of mice with peritonitis-sepsis caused by P. aeruginosa = 27
Fig. 7. Bacterial burdens in organs of mice with or without phages = 28
Fig. 8. Pharmacokinetics of MPK1 and MPK6 in D. melanogaster = 30
Fig. 9. Bacteriophage therapy against D. melanogaster systemic infection = 32
