Phylogenetic study on CTO primer-amplified ammonia-oxidizing bacteria and β-Proteobacteria associated with rice roots grown in a flooded paddy soil

Ammonia-oxidizing bacteria associated with rice roots that developed from different nodes at different growth stages were studied by using polymerase chain reaction (PCR)-denaturing gradient gel electrophoresis (DGGE) and sequencing analysis. Root samples used in this study were collected three times on July 2 (tillering stage), on July 21 (maximum tillering stage), and on September 12 (ripening stage). The fact that many DGGE bands of bacterial 16S rDNA amplified by the CTO primer set that was originally designed for ammonia-oxidizing beta-Proteobacteria, were observed in every nodal root on July 2 and July 21 and in nodal roots with small number on September 12 indicated the presence of ammonia-oxidizing bacteria under reduced soil conditions. All the ammonia-oxidizing bacteria associated with rice roots belonged to Nitrosospira spp. In addition, many 16S rDNA of beta-Proteobacteria were also amplified by the CTO primer set, and they belonged to the Rhodocyclus and Methylophilus groups. The growth stage of rice plant was the primary factor affecting CTO-amplifled bacterial communities. Root age was also estimated to be an important factor influencing the communities throughout the growth stages of rice plants.