Abstract: Studies on the effects of the interspecific interactions of maize||soybean intercropping on the rhizosphere microbial community structure and their relationship with crop yield are of theoretical and practical significance for elucidating the yield effects of interspecific crops in intercropping systems. The aim of this study was to explore the changes in microbial community structure in the rhizospheres of soybean and maize planted under an intercropping system (soybean||maize) with a 2∶3 line ratio and a randomized design pattern with three types of partitions between two crop roots. The intercropping partitions were a mesh barrier (MB, with exchange of root extudates without roots interaction) or a polythene film barrier (PB, without exchange of root extudates and roots interaction) to separate the maize roots from soybean roots or no barriers (NB) between the roots. An independent monoculture (M) was set up as a control. BIOLOG and terminal restriction fragment length polymorphism (T-RFLP) assays were used to investigate the microbial community diversity in the maize||soybean rhizospheres. The results showed that the land equivalent ratios (LERs) under NB, MB, and PB conditions were 1.39, 1.13, and 0.98, respectively, at a plant row ratio of maize||soybean of 2∶3. These findings suggest that the LER increases with increased interspecific root interactions from PB to NB under the same intercropping pattern. Further analysis revealed that the microbial diversity and evenness indexes in the rhizosphere of both intercropped maize and soybean similarly increased with the increase in interspecific root interactions from PB to NB. Average well color development (AWCD) analysis showed that the rhizospheric microbial communities under NB and MB conditions had the strongest overall ability to utilize carbon sources as substrates, whereas those under PB and M conditions had a lower ability in this regard. The enhancement of interspecific root interactions increased the ability of rhizospheric microbes of intercropped soybean to utilize amines, polymers, amino acids, and carbohydrates (four types of carbon-source substrates) by 181.01%, 32.6%, 37.84%, and 78.28%, respectively. However, the capability of microbes in the intercropped soybean rhizosphere for utilizing two other carbon sources (phenols and carboxylic acids) decreased. Moreover, the ability of the microorganisms in the intercropped maize rhizosphere to utilize carboxylic acids, carbohydrates, and amines increased by 46.26%, 6.54%, and 15.84%, respectively, whereas their ability to utilize phenols, polymers, and amino acids decreased. T-RFLP analysis revealed a significant increase in the abundance of dominant bacteria, such as Rhodococcus (Actinomycetes) and Halobacillus (Firmicutes), in the rhizosphere of intercropped soybean under NB compared with that under PB; whereas the abundance of beneficial dominant bacteria, such as Rhodococcus (Actinomycetes) and Bacillus (Spirochetes), markedly increased in the rhizosphere of intercropped maize under NB conditions compared with that under PB conditions. As a result, the crop yield and LER increased under intercropping conditions.