Pseudomonas tolaasii 박테리오파지에 특이적인 다클론항체 형성 및 이를 이용한 파지 교차 반응성 = Pseudomonas tolaasii bacteriophage-specific polyclonal antibody formation and its cross reactivity to various phages

국문 초록 (Abstract)

Pseudomonas tolaasii는 느타리버섯에 갈반병을 일으키는 병원균주로, 다양한 변이균주들을 분리하여 P1α와 P1β, P1γ 세 가지 소그룹으로 분류하였다. 각 그룹별 균주들에 특이적인 박테리오파지를 이용한 파지테라피는 갈반병 방제에 매우 성공적이었다. 본 연구에서는, 박테리오파지들의 특성을 구명하기 위하여 소그룹별 대표균주을 이용하여 파지를 분리하였고, 이들의다클론항체를 제작하여 파지들 사이에 유연관계를 조사하였다.
파지 준비물은 1010 pfu/mL 이상으로 토끼의 다리 근육에 주사하였고, 3회의 반복주사에 의해 다클론항체가 얻어졌다. 파지φ6264에 대한 항체의 역가는 2×107 Ab/mL 이상, 파지 φHK2 에 대해서는 1×106 Ab/mL, 파지 φHK19와 φHK23에 대해서는 1×107 Ab/mL 이상이었다. 항체와 이에 특이적인 파지 사이에는 매우 높은 반응특이성이 있었고, 소그룹이 다른 파지의 항체와 파지 사이에도 일부 교차반응성을 확인하였다. 파지 φ6264 에서 생성된 Abφ6264는 모든 P1α 소그룹의 파지들과 반응성을 보였으나, 파지 φHK16을 제외한 다른 소그룹의 파지들과는반응하지 않았다. P1γ 소그룹에서 생성된 AbφHK23은 P1β 소그룹의 모든 파지들을 불활성화시켜 가장 넓은 항체범위를 보였다. 항체와 파지를 이용한 숙주균과의 관계를 분석하였을 때, 16S rRNA 유전자 분석에 의한 숙주균의 근연관계와 항체를 이용한 숙주균의 파지들 사이의 구조적 근연관계는 상당히 차이가있음을 확인하였다. 결론적으로, 박테리오파지의 숙주균 특이성과 항체를 이용해 측정한 파지의 껍질단백질 구조유사성 사이에는 약한 상관성을 보였다.

다국어 초록 (Multilingual Abstract)

Pseudomonas tolaasii causes brown blotch disease on the oyster mushroom (Pleurotus ostreatus). Various pathogenic strains of P. tolaasii were isolated and divided into three subtypes, P1α, P1β, and P1γ. For phage therapy, bacteriophages against to these subtype strains were applied to mushroom cultivation and very successful to prevent from the disease. In this study, bacteriophages were isolated against the representative strains of subtype pathogens and their polyclonal antibodies were synthesized to investigate structural relationship among capsid proteins of phages. Phage preparations over 1010 pfu/mL were injected to rabbit thigh muscle and polyclonal antibodies were obtained after three times of boost injection. Titers of the antibodies obtained were over 2×107 Ab/mL for the phage φ6264, 1×106 Ab/mL for the phage φHK2, and 1×107 Ab/mL for the phage φHK19 and phage φHK23. High specific activities were observed between antibodies and the corresponding bacteriophages. Some crossreactivities between the antibodies and non-corresponding bacteriophages were also measured. Antibody Abφ6264 inactivated all phages of P1α subtype and only phage φHK16 among P1β subtype phages. Antibody AbφHK23 of P1γ subtype neutralized all phages of P1β subtype as well as the phage φHK23, showing the widest phage-inactivation range. When the structural-similarity studies of phages were investigated by using phage antibodies, closeness obtained by phylogenetic analysis of 16S rRNA genes of pathogenic strains were quite different from that of polyclonal antibody-specific structural similarity of phage capsid proteins. In conclusion, there is weak correlation between the host strain specificity of bacteriophage and its capsid structural similarity measured by phage antibodies.

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