Tools to evaluate Herbaspirillum seropedicae abundance and nifH and rpoC expression in inoculated maize seedlings grown in vitro and in soil

The plant growth promoting bacteria Herbaspirillum seropedicae is an important model to study biological nitrogen fixation and it is proposed as crop inoculants for grasses. In this study we developed new tools to evaluate the abundance of H. seropedicae SmR1 and its expression in planta and investigated the association of the plant growth promoting bacteria H. seropedicae with maize grown in sterile and nonsterile conditions. Maize seedlings (P30F53) were inoculated with H. seropedicae SmR1 and grown in vitro and in soil. The plants were sampled at 4, 7 and 10 (in vitro) or 14, 21 and 28 days after inoculation (soil). Using qPCR we quantified H. seropedicae DNA and measured nifH, rpoC and hrcN levels of bacterial transcripts. In vitro assay inoculated plants presented highest amount of DNA and transcript contents, not detected in control plants. nifH and rpoC gene expression were detected on roots of inoculated maize cultivated in both growth conditions. However, it was not possible to detect hrcN gene expression in maize roots cultivated in soil. TaqMan assay is species-specific for H. seropedicae DNA and nifH and rpoC transcript levels could be used to monitor H. seropedicae gene expression in planta in sterile and nonsterile growth conditions. We developed specific, reliable and efficient tools to monitor H. seropedicae DNA abundance and H. seropedicae nifH and rpoC expression in planta.