US EPA

2850428 

Journal Article 

A novel intracellular nitrogen-fixing symbiosis made by Ustilago maydis and Bacillus spp 

Ruiz-Herrera, J; León-Ramírez, C; Vera-Nuñez, A; Sánchez-Arreguín, A; Ruiz-Medrano, R; Salgado-Lugo, H; Sánchez-Segura, L; Peña-Cabriales, JJ 

2015 

1 

New Phytologist
ISSN: 0028-646X
EISSN: 1469-8137 

207 

3 

769-777 

English 

25754368 

10.1111/nph.13359 

WOS:000357824400028 

We observed that the maize pathogenic fungus Ustilago maydis grew in nitrogen (N)-free media at a rate similar to that observed in media containing ammonium nitrate, suggesting that it was able to fix atmospheric N2 . Because only prokaryotic organisms have the capacity to reduce N2 , we entertained the possibility that U. maydis was associated with an intracellular bacterium. The presence of nitrogenase in the fungus was analyzed by acetylene reduction, and capacity to fix N2 by use of (15) N2 . Presence of an intracellular N2 -fixing bacterium was analyzed by PCR amplification of bacterial 16S rRNA and nifH genes, and by microscopic observations. Nitrogenase activity and (15) N incorporation into the cells proved that U. maydis fixed N2 . Light and electron microscopy, and fluorescence in situ hybridization (FISH) experiments revealed the presence of intracellular bacteria related to Bacillus pumilus, as evidenced by sequencing of the PCR-amplified fragments. These observations reveal for the first time the existence of an endosymbiotic N2 -fixing association involving a fungus and a bacterium. 

Bacillus pumilus; endosymbiosis; nifH gene; nitrogen fixation; nitrogenase; Ustilago maydis