The cultivation of transgenic () has received worldwide attention since Bt crops were first released. Its ecological risks on arbuscular mycorrhizal fungi (AMF) have been widely studied. In this study, after cultivation for five seasons, the AMF diversity and community composition of two Bt maize varieties, 5422Bt1 (event Bt11) and 5422CBCL (event MO10), which both express Cry1Ab protein, and their isoline non-Bt maize 5422, as well as Bt straw after cultivation had been returned to subsequent conventional maize variety, were analyzed using Illumina MiSeq sequencing. A total of 263 OTUs (operational taxonomic units) from 511,847 sequenced affiliated with the AMF which belonged to Mucoromycota phylum Glomeromycotina subphylum were obtained. No significant difference was detected in the AMF diversity and richness (Shannon, Simpson, ACE, and Chao 1 indices) and community composition in rhizosphere soils and roots between Bt and non-Bt treatment revealed by NMDS (non-metric multidimensional scaling) and NPMANOVA (non-parametric multivariate analysis). Moreover, was the most dominant genus in all samples. Although there was no significant difference in the AMF community in roots and rhizosphere soils between the Bt and non-Bt maize treatments, total phosphorus (TP), total nitrogen (TN), organic carbon (OC), and pH were driving factors affecting the AMF community, and their composition varied between rhizosphere soils and roots during the maturity period of the fifth season. Compared to our previous study, the results were identical. In conclusion, no significant difference was observed between the Bt and non-Bt treatments, and the Illumina MiSeq method had higher throughput and higher quality read cover, which gave us comprehensive insight into AMF communities in agro-ecosystems.