Abstract: Bt Cry8Ea protein presents insecticidal activity to Holotrichia oblita larvae. The interaction between Bt Cry8Ea protein and the midgut microorganism in H. oblita larvae was investigated by metagenome DNA analysis. The midguts in normal-fed or Cry8Ea protein-fed H. oblita（2nd-instar larva）were collected and then used to extract metagenome DNAs of midgut microorganism for constructing 16S rDNA gene libraries. Polymerase chain reaction restriction fragment polymorphism（PCR-RFLP）was employed to determine the polymorphisms of 16S rDNA enzyme-spliced fragments（Msp I，Rsa I，and Hae III restriction enzyme）from the midgut microorganism of two fed larvae. Different types of clones were sequenced and then aligned with NCBI gene database，and the phylogenetic tree was constructed to study the differences of microbial communities. The results showed that there were 30 bacteria in the normal group of H. oblita，and the predominant bacteria were Desulfosporosinus（29.63%），Clostridium（12.59%），and Bacillus（10.37%）respectively. However，there were only 4 bacteria in the Cry8Ea protein-fed H. oblita group，and the predominant bacteria were Enterococcus（50%）and Streptococcus（46.88%）. These data revealed that the treatment with Cry8Ea protein changed the abundance and composition of midgut microbial flora.

Key words: Holotrichia oblita, midgut microorganism, Bt Cry8Ea protein, PCR-RFLP