N2 fixation by free-living and associative organotrophic diazotrophs in marine and terrestrial habitats*

David Patriquin, Dalhousie University

I met Dr. Roger Knowles in 1970 while in the final stages of my PhD on nutrient sources for tropical seagrasses. I had concluded that there had to be high levels of nitrogen fixation in the sediments in which they
Roger Knowles
Ruth & Roger Knowles, 2004
are rooted. Roger encouraged me to work on the problem in his lab and later I took up a postdoc there. A vintage mass spec enabled 15N studies of N2 fixation, while we used the latest GCs to analyze atmospheric gases and apply the novel acetylene reduction technique to probe sites and processes of N2 fixation. On a memorable visit to DuPont labs, Ralph Hardy talked optimistically about the potential for N2 fixation and the newly discovered C4 photosynthetic pathway to reduce fertilizer needs and increase crop production. From Brazil came reports of high N2-fixing activity associated with roots of tropical grasses including maize. Amidst the excitement of these discoveries, Roger instilled in us the value of stepwise, hypotheticodeductive approaches for investigating questions in microbial ecology and the importance of evaluating the related methodologies. Research I began in Roger's lab and carried on through the 70s and 80s with students at Dalhousie University and collaborators at other institutions added new genera to the list of marine diazotrophs, (Vibrio and Campylobacter - C. nitrofigilis later became the type species for the genus Arcobacter), documented or suggested novel sites of diazotroph activity (rhizosphere and sediments of seagrasses, internal tissues of graminoid roots and stems, sea urchin guts, saltmarsh detritus, sugarcane litter) and revealed a fungal-diazotroph association (Helicomyces roseus/Azospirillum brasilense) in sugarcane litter that fixes nitrogen under fully aerobic conditions. Following on the pioneering work by Johanna Dobereiner, research by many investigators has uncovered a high diversity of organotrophic diazotrophs, including many new species, associated with terrestrial plants. It has also provided detail at molecular and higher levels about colonization of roots and stems by diazotrophs. However, recent evaluations of the contribution of N to major crops from associative N2 fixation suggest it is small, with the possible exceptions of rice, sugarcane and some perennial forages under particular conditions. Efforts to manipulate graminoid-diazotroph associations to enhance N2 fixation have so far not succeeded. Recent studies have increased the estimates of N2 fixation by free-living and associative organotrohic diazotrophs in marine systems, much of it attributed to sulfate reducers. In association with seagrasses, sulfate reducers fix N2 in excess of their own needs, there is rapid transfer of newly fixed N to above-ground biomass and as much as a third of N assimilated by the plant can come from associative nitrogen fixation.

*Presentation in the Roger Knowles Tribute Symposium, Canadian Society of Microbiologists Annual Conference, Memorial University June 20-23, 2011. Slides (PDF)