棘胸蛙脑膜炎病原双重PCR快速诊断技术及疫苗免疫保护效果研究
Meningitis Pathogenic Bacteria Identification, Doublex PCR Rapid Diagnosis and Immune Protective Effect of the Vaccine in the Frog (Quasipaa spinosa)

本研究从患脑膜炎的棘胸蛙体内分离到QSM01、QSM02和QSM03三株菌，采用细菌形态学、生理生化特征及分子生物学技术对其进行鉴定，并开展了药物筛选、快速诊断PCR诊断技术和免疫保护试验。结果显示：QSM01为朱斯特金黄杆菌(Chryseobacterium joostei)，QSM02为脑膜炎败血伊丽莎白菌(Elizabethkingia meningoseptica)，QSM03为比目鱼金黄杆菌(Chryseobacterium scophthalmum)；药敏实验表明QSM01对利福平等5种抗生素敏感，对氯霉素5种抗生素中度敏感；QSM02菌株对新生霉素、红霉素、利福平敏感，对氯霉素、链霉素、万古霉素中度敏感；QSM03菌株对万古霉素等抗生素6种敏感，对四环素等5种抗生素中度敏感。建立了双重PCR快速诊断方法，可特异性检测出致病性最高的QSM02菌株。采用注射、浸泡和喷雾方式给蛙接种QSM02甲醛灭活疫苗，经攻毒后免疫保护率分别达到100%、87.5%和75%。因此，采用浸泡和喷雾法接种疫苗方式在该蛙防治脑膜炎上是可行的。
This experiment made the identification of the morphological characteristics, physical and chemical characteristics and the molecular biology to QSM01, QSM02, QSM03 which are isolated from a disease Quasipaa spinosa in meningitis and at the same time, made the drug screening, rapid PCR diagnostic techniques and immune prevention tests. The result showed QSM01 is Chryseobacterium joostei; QSM02 is Elizabethkingia meningoseptica; QSM03 is Chryseo-bacterium scophthalmum. Drug sensitivity test showed that QSM01 was sensitive to 5 antibiotics such as rifampicin and moderately sensitive to 5 chloramphenicol such as Chloramphenicol, QSM02 strain was sensitive to erythromycin, vancomycin and rifampicin, moderately sensitive to chloram-phenicol, streptomycin and vancomycin, QSM03 strain was sensitive to 6 antibiotics such as vanco-mycin, moderately sensitive to 5 antibiotics such as tetracycline. The duplex PCR can detect the heaviest pathogenic bacterial strain QSM02 specifically. The immune protection rate for Quasipaa spinosa after injection, soak and spray of QSM02 inactivated vaccine can reach 100%, 87.5%, 75%. Therefore, it’s feasible to control the disease by soaking and spraying the vaccine.