重组荧光假单胞菌BioP8在环境中消长动态与安全性评价

On dynamics of colonization and biosafety assessment of genetically modified Pseudomonas fluorescens BioP8 in different environments

通过选择分离培养和 PCR鉴定 ,对转苏云金芽孢杆菌(Bacillus thuringiensis,Bt)杀虫蛋白基因荧光假单胞菌 Bio P8及其天然受体菌 P30 3在温室自然土壤和田间大白菜根际、叶面的环境生存竞争能力进行研究。结果表明 ,在温室自然土壤中 ,可培养细菌总量在 1 0 8CFU/g土 (湿重 )左右 ,40 d内 P30 3和 Bio P8群落总量分别能维持在 1 0 5CFU /g土 (湿重 )和 1 0 4 CFU /g土 (湿重 )左右 ,1 0 0 d后检测不到残留 ;在大白菜根际 ,可培养细菌总量在 1 0 1 0 CFU/g根 (湿重 )左右 ,30 d内 P30 3和 Bio P8可维持在 1 0 6 CF U/g根 (湿重 )和 1 0 5CFU /g根 (湿重 ) ,75 d后检测不到残留 ;在大白菜叶面 ,可培养细菌总量在 1 0 3～ 1 0 6 CFU/cm2叶片之间波动 ,处理后 3d的供试菌菌量迅速由 1 0 5CFU/cm2叶片降至最低 (1 0 3CFU/cm2叶片和 1 0 2 CFU/cm2叶片左右 ) ,之后回升并以 (1 0 4和 1 0 3) CF U/cm2叶片维持一段时间 (第 5— 1 5 d) ,30 d检测不到残留。P30 3及其工程菌株在温室自然土壤、田间大白菜根际和叶面的定殖时间至少分别为 6 0 d、5 0 d和1 5 d,它们的消长动态相似 ,在温室自然土壤、田间大白菜根际的定殖能力明显强于叶面 ,Bio P8定殖能力弱于出发菌 P30

The present paper aims to present the authors' comparative study as to the survival and colonization of Pseudomonas fluorescens P303 and its genetically modified derivative BioP8 with Bt cry1Ac, cry1Ab and cry2Aa genes, both in natural soil and in greenhouse environment. The study is done in the rhizosphere and the phyllosphere of cabbage in the fields through selective culture and PCR identified methods. The results show that the total population of cultivable bacteria in soil in the rhizosphere is around 10~8 CFU/g soil (wet weight) and 10^(10) CFU/g roots (wet weight), and the population of P303 and BioP8 retains 10~5CFU/g soil and 10~4 CFU/g soil respectively in natural soil within 40 days, 10~6and10~5 CFU/g roots in rhizosphere of cabbage for 30 days. However, the total population of cultivable bacteria on the phyllosphere of cabbage fluctuates between 10~6and 10~3CFU/cm^2leaf, and that of P303 and BioP8 decline from 10~5to 10~3and 10~2 CFU/cm^2leaf from the first day to the 3rd day sharply, then increases and keeps steadily between 10~3 and 10~4 CFU/cm^2leaf (5th-15th days). However, both P303 and BioP8 can not be detected in soil, in the rhizosphere and the phyllosphere of cabbage after 100, 75 and 30 days respectively. In conclusion, BioP8 and P303 are found to have similar dynamics of colonization and can survive no less than for 60, 50, 15 days in soil, rhizoshpere and phyllosphere respectively, but the ability of colonization of both strains in soil and rhizosphere proves stronger than that in phyllosphere, with P303 being appreciably stronger than BioP8.