| Honey bee plays an important role in ecology and economy as the pollinator of a lot of crops and wild plants, and it is an important non-target insect. Due to the rapid decline in distribution of our native honey bee (called Apis cerana cerana) which is in imminent danger, the recent researches focus on Apis mellifera ligustica which is very important in plant pollination and maintaining ecological balance. It is one of the focal points that is concerned a lot that whether transgenic crops will have a negative impact on honey bee. The influence of transgenic crops on the amount and structural diversity of soil microbial is an important indicator to evaluate the safety of transgenic crops, because plants and soil microbial effect each other mutual restraint and mutual promotion via rhizosphere effect. In this paper, to provide theoretical basis for the evaluation system for safety of transgenic plants, independent researched transgenic maize with cry1Ah gene which has high toxicity against Ostrinia furnacalis was used as experimental materials to study its effect on bacterial community in the midgut of Apis mellifera ligustica and the soil microbial community, and the results showed as below.1. The influence of the transgenic maize pollen with gene cry1Ah on the bacterial community in the midgut of Apis mellifera ligustica in the lab.Sugar with the transgenic maize pollen with gene cry1Ah, normal maize pollen, high concentration plasmid, fragment of linear cry1Ah and pure sugar was fed to Italian honeybee separately. Two weeks later, honey bees were dissected to gain the genomic DNA of intestinal bacteria, and Denaturing Gradient Gel Electrophoresis was used to analyse the community composition of bacterial in the midgut of honeybees in the five treatment groups. As the results showed, the Shannon's index of the five treatment groups in DDGE profiles distributed between 2.51-2.79 and the evenness index was between 0.99-1.01. It suggested that the difference on the community composition of intestinal bacteria with different treatment is not significant. The specific bands in DGGE profiles was recycled, cloned and sequenced in succession. The results of sequences analysis suggested that they are mainly belong to Firmicutes, Proteobacteria and Actinobacteria. Firmicutes is mainly Composed of Lactobacillus, while Proteobacteria contains α-,β- andγ-Proteobacteria. Still, there are some kinds of bacteria have not been classified.2. The influence of the transgenic maize pollen with gene cry1Ah on the bacterial community in the midgut of Apis mellifera ligustica in the net room of web house.The transgenic maize and normal maize were planted in the web house in experimental base located in Lang fang apart. Every web house was put into three hives of honeybee which contained more or less 10000 honeybees during the flowering stage of maize. After marking, the larva was sampled after freshing pollen for 0 days, 3 days, 6 days and 9 days separately, while the worker bee was sampled after 0 days, 15 days and 30 days. Every treatment above was sampled 3 times. The community composition of bacterial in the midgut of honeybees in the five treatment groups was analysed with the Denaturing Gradient Gel Electrophoresis after the genomic DNA was gained from intestinal bacteria. As the results showed, the Shannon's index of the larva in DGGE profiles distributed between 2.88-3.11 and the evenness index was between 0.99-1.01. It showed that the difference on the community composition of intestinal bacteria with different days larva is not remarkable. The results of sequences analysis suggested that they are mainly belong to Firmicutes and Proteobacteria. the Shannon's index of the worker bee in DGGE profiles distributed between 2.60-2.78 and the evenness index was between 0.99-1.00. The result showed that there was no notable difference on the community composition of intestinal bacteria with different days work bee. The results of sequences analysis suggested that they are mainly belong to Firmicutes, Proteobacteria and Actinobacteria.The conclusion that transgenic cry1Ah maize would not pose detrimental effect on the larva and work bee of Apis mellifera ligustica was deduced.3. The influence of the transgenic maize with gene cry1Ah on the microorganism community in the rhizosphere soil.The method of conventional culture techniques was employed to analyze the amounts of bacteria, fungi and actinomycetes in the rhizosphere soil planted transgenic maize with cry1Ah gene. Simultaneously, the effect of on microorganism community structure in rhizosphere soil was assessed with culture-independent technique polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE) and BIOLOG assay. The rhizosphere soil sample of the maize was collected after growing for 0 days, 15 days, 30 days, 60 days, 90 days, 180 days, the amount of cultured rhizosphere microorganisms was some variation, but no significant difference between transgenic and regular maize in the same period. Moreover, the results of cluster analysis of DGGE profile of bacteria and fungi indicated that there was no remarkable difference in the same phase. The specific bands in DGGE profiles was recycled, cloned and sequenced. The bacteria results of sequences analysis suggested that they are mainly belong to Firmicutes, Proteobacteria, Actinobacteria and Unknown. The results of fungi sequences that they are mainly belong to Basidiomycotina, Deuteromycotina, Ascomycotina, zygomycotina and Unknown。For the two investigations, the Shannon's index of the bacterial community in the rhizosphere soil was 2.40-2.70 and the evenness index was0.96-0.99; the Shannon's index of the fungal community in the rhizosphere soil was 2.17-2.48 and the evenness index was 0.95-0.99. The soil sample was collected after growing 0 days, 30 days, 90 days, 180 days, BIOLOG assay results showed that there was no remarkable difference of the average well color development, Shannon, Simpson and McIntosh of between transgenic and common maize. It implies that the transgenic maize with the cry1Ah gene did not affected significantly on community structure of microorganism in rhizosphere soil.The work we carried out here will impetus the development of transgenic maize, enhance its international competitiveness. Key words transgenic maize; honey bee; intestinal microbe; denaturing gradient gel electrophoresis; BIOLOG; soil microorganism; community structure... |
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