Identification of phytate mineralizing bacteria colonized on the extraradical hyphal surface of arbuscular mycorrhizal fungi in a maize field

【Objectives】 The objective of this study was to investigate whether phosphate mineralizing bacteria (PMB) could colonize on the hyphal surface of arbuscular mycorrhizal (AM) fungi in maize field and to identify these bacteria and test their abilities in mineralizing organic phosphate. This can help to go to deep into understanding the functions of hyphosphere bacteria in soil phosphorus turnover and biogeochemical cycle.【Methods】 We collected the extraradical hyphae of AM fungi associated with maize roots from the field at China Agricultural University long-term experimental station in Quzhou, Hebei province. The PMB attached to AM fungal hyphae were selected using the phytate-selective medium contained phytin as the sole phosphorus source. The DNA of selected bacteria was extracted with TIANamp Bacteria DNA Kit according to the manufacturer’s instructions. Then the genera of PMB were identified through bacterial 16S rDNA sequencing. The abilities of PMB to utilize organic phosphate were determined in solid phytate-selective medium contained phytin by the diameter of bacterial colony (d) and phosphorus solubilizing halo (D), and by inorganic phosphorus concentration and pH in liquid phytate-selective medium. Each PMB isolated from AM fungal hyphae was analyzed in triplication. PMB strains belonged to different genera were inoculated into the hyphal compartment of a two-compartment microcosm conducted in the greenhouse to test their abilities to re-colonize on the hyphal surface. 【Results】 Twenty-nine strains of PMB were isolated from the surface of AM fungal hyphae and affiliated to Bacillus, Serratia, Pseudomonas, Staphylococcus and Enterobacter, respectively. The abilities of PMB to utilize phytin in the liquid medium showed that the mineralization rates ranged from 1.9% to 21.9%. Among them, the genus of Pseudomonas had the highest mineralization rate (more than 14%), meanwhile the pH of liquid medium was reduced by 2 to 4 units. All the identified PMB strains were re-inoculated into the hyphal compartment of a two-compartment microcosm in the greenhouse. After 30 days, we found that four genera of the PMB were isolated and identified successfully from the hyphal surface with the exception of the genus of Pseudomonas, interestingly, another PSM strain, Cupriavidus was detected.【Conclusions】 The results demonstrated that PMB could attach to the surface of extraradical hyphae of AM fungi associated with maize in the field and 29 bacterial strains belonged to 5 genera were identified. They had different ability to mineralize phytin and the genus of Pseudomonas had the highest mineralization rate.