Rice paddy soils are an important biogenic source of methane, a relevant greenhouse gas. Rice rotation with upland crops and pastures affects hydrolytic and fermentative bacteria and methanogenic archaea by sequential exposition to oxic and anoxic conditions. There are several ways to reduce methane emissions from rice paddy soils. Here we examine the effect of intensification in rice rotations with upland crops upon the microbial community structure and methanogenesis when dicarboxylic acids exudated by rice root were amended in microcosms assays. Soils under rotations rice-pasture, rice-soybean and continuous rice, with the last two more intensive systems recently installed in the field experiment, were sampled. Amplicon sequencing of 16S rRNA gene analysis showed that the rotation system was a main driver of the microbial community structure, explaining 33%, 38% and 55% of the differences in the structure of Bacteria, Archaea and methanogens, respectively. The density of mcrA gene copies was significantly higher in continuous rice soil than in other soils, and the relative abundance of methanogenic archaea was 33% for rice monoculture, whereas represented 15% and 14% for the rice-pasture and rice-soybean soils, respectively. The incubation of soils with tartrate and succinate confirmed functional differences among soils since rice monoculture showed similar or higher methane production and significantly less acetate and propionate accumulation than other soils. The T-RFLP analysis of mcrA gene in soils and incubated soils revealed that Rice Cluster I increased after incubation in all rotational soils, whereas Methanosarcinaceae and Methanobacteriaceae remained relevant after incubation. Furthermore, a more stable community of methanogenic archaea was established in soil under continuous rice, since T-RF profiles of soils and incubated soils with dicarboxylic acids grouped together with 62% similarity, while communities from other rotational soils experimented a great shift after incubation with these substrates. Altogether, these results showed that the conversion of rice-pasture rotational fields to a more intensive rice crop production strongly impacts the methanogenic community structure and their potential for methane emission.