Abstract
As beneficial microorganisms in the soil, arbuscular mycorrhizal fungi (AMF) and rhizobia (Rh) form a mutualistic symbiosis with plant roots, while it is unclear whether AMF and Rh synergistically promote root colonization, plant growth, and nitrogen (N) acquisition during the colonization. The aim of this study was carried out to analyze the effect of inoculation with an arbuscular mycorrhizal fungus (Paraglomus occultum) and a rhizobium (Rhizobium trifolii), singly and in combination, on root mycorrhizal colonization and nodules as well as changes in plant growth, hydrogen peroxide (H2O2), and N acquisition of white clover over a period of 90 days. Double inoculation of AMF and Rh favored root AMF colonization and nodules compared to single inoculation. AMF colonized roots earlier than the formation of root nodules, with AMF colonized the root nodules after 30 days. H2O2 (a signaling molecule for plant defense against microbial infestation) was increased in roots from day 5 to day 10 following dual inoculation and single AMF inoculation, and from day 10 to day 15 following single Rh inoculation, indicating a pre-response of roots to AMF colonization during the two microbial colonization. At 20 days after inoculation, single or double inoculation of AMF and Rh showed positive effects on leaf and root biomass production and plant height, while improved effects on taproot length began at 15 days of inoculation, along with the improved magnitude most prominently with double inoculation. The earliest effect of significant improvement in leaf N appeared on day 10 of double inoculation, while the improvement in root N started 15 days later, following the prominent effect with double inoculation among three treatments. It is concluded that AMF and Rh could synergistically promote each other's colonization in roots, thus further enhancing plant growth and N acquisition.