Diazotrophy is the ability to biologically fix atmospheric nitrogen gas (N2) into ammonia (NH3). This magical process allows otherwise inert, biologically unavailable nitrogen gas to fuel the primary production which brings forth all higher life on the planet. It cannot be overstated how vital this novel process is to the function of both wild ecosystems as well as humanity’s global food supply. Without diazotrophy, there would be no fields, no forests, no seaweed beds, no bogs, no jungles or prairies, no coral reefs nor meadows, mangroves or magnolia gardens. With diazotrophy, oases bloom from deserts; cactuses can grow in sand, tillandsia grow without soil and fields of sargassum seaweed emerge from the desolate open sea.
Diazotrophy is still not very well-understood, but its mysteries have inspired new generations of scientists to dissect and wield the particularities of this almost divine power. What is certain is that diazotrophy is key to understanding the basic function of our world’s ecology and is paramount to sustaining humanity’s place within it. For millennia, humans have been indirectly utilizing diazotrophs to enhance agriculture. Traditional agriculture relies on ‘aged’ soil to produce greater and greater crop yields over time on the same plot.
Not to be confused with crop fertilization itself, the process of aging a soil begins with the establishment of both photosynthetic and diazotrophic microorganisms. The purple non-sulfur bacteria (PNSB) of the genera Rhodopseudomonas, Rhodospirillum, Rhodobacter, etc. fit both of these functional roles. Because of their ‘swiss army knife’ metabolism, they can conduct a great many biochemical feats--and when exposed to nitrogen-devoid soil, they seize N2 gas from the air and transform it into ammonia (NH3). This NH3 is the perfect fertilizer for photosynthesizers who in turn propagate and produce food for the greater biological community. By this process, aged soil ‘breathes’ in N2 and ‘exhales’ an enormous functional diversity of critters. It is this rich soil diversity which allows for enhanced plant productivity, quality and resiliency.
A return to Nature
“Interestingly, the diazotrophic communities in dryland soil are more robust and contain more generalists compared to irrigated soils.” (Alleman et al 2021)
Traditional agriculture was, and is, dependent on the natural function of the soil; this however proved insufficient to support larger-scale monoculture farming. Innovations in farming practices, such as tilling and fertilization, increased crop yields in the short-term but depleted the natural performance of the soil in the long-term. Constant shuffling of the various soil layers prevents the establishment of stable and diverse microbial communities. Fertilizing soils can improve their productivity in the short term; still, just as an aquarium can be overfed, flooding soils with "raw" nitrate and phosphate ultimately poisons a soil’s natural ecology. The more such techniques are employed, the more natural soil ecosystems are erased, rendering farmland less productive and more expensive to manage. The very same consequences arise from overfertilization of hydroponic/aquaponic water assets.
Farmers, scientists and state agencies of the past Industrial Age possessed little to no appreciation for the natural ecology which allowed their soil to breathe in N2 and exhale life. As a result, they systematically dismantled and destroyed what had nurtured their forefathers and instead paid fortunes to mine, ship and distribute bulk sources of ammonia (guano, saltpeter, and so on). Such practices found the 20th Century in constant food deficit as the human population rose and nations became willing to murder each other over bat feces--amongst other natural resources.
Though ravenous in its need for fossil fuel, the Haber-Bosch Process allowed Imperial Germany to both farm and produce munitions throughout WWI despite being cut off from South America’s guano-rich islands. After the war, this technology spread to other nations and fundamentally changed agriculture forever. Artificial fertilizer was produced and distributed on an unprecedented scale, markedly reduced in cost and subsequently applied with feverishly increasing intensity and frequency! This is our world today, a world where artificial fertilizers have allowed for true industrial farming--food is produced in incalculable surplus (albeit of an overall compromised quality). The Haber-Bosch Process allowed humanity to divorce itself from natural soils and for a brief time succeed---we have survived, grown and fattened but it has also come at a substantial cost.
In the 21st Century, fossil fuels possess a precious value which almost rivals that of food and water. The mass production of artificial fertilizers has become less a luxury and more an addiction, as modern industrial farms cannot fertilize themselves and are thus incredibly dependent on fossil fuel. As artificial fertilizers are inefficiently absorbed by crops, they contribute to run-off that destabilizes the ecology of rivers, streams, ponds, lakes, estuaries, wetlands and other water bodies. This is where the value of diazotrophy has become obvious, as microbial keystone species, such as Rhodopseudomonas palustris, offer humanity a means of reestablishing a bountiful soil ecology that resembles the ones we have destroyed. Perhaps even more remarkably, they impart a certain degree of natural ecological function to completely artificial grow systems such as hydroponics and aquaponics.
(Haber-Bosch)
(Biological Diazotrophy)
When comparing the Haber-Bosch reaction to that of a generalized diazotroph, one is immediately struck by how comparatively inefficient the artificial process is. Exponentially more energy is required at higher temperatures to produce the same amount of ammonia by burning fossil fuels than allowing a biological cell to naturally fix the N2. Purposefully deploying diazotrophs into agricultural soil/water will be key to establishing an organically grown food supply of ever-increasing quality and abundance.
The triple bond between the two N atoms in nitrogen gas is extremely strong and stable. This is why N2 comprises over 80% of the atmosphere and naturally reacts with very few substances. Artificially, it requires strong reagents such as molten potassium or red-hot magnesium to get it to react. Diazotrophs are especially habile because they have manifested a way to break this N2 triple bond at 77°F. The key to this power is the enzyme nitrogenase.
Though nitrogenase is produced by a diverse variety of diazotrophs, there are only a select few genes which code for it. This has led scientists to believe that several diazotrophs obtained their abilities from methanogenic archaea through horizontal gene transfer. It was previously believed that all nitrogenase enzymes were based around a core of molybdenum (Mb), but studies over the decades have revealed at least two other general variations of the enzyme based around vanadium (V) and iron (Fe). Likely, there is an even greater diversity of nitrogenases than previously appreciated. In all its forms discovered, described and otherwise, nitrogenase allows diazotrophs to break the triple bond and attach hydrogens to the severed nitrogen atoms. This process is energy and resource intensive as 12-15 ATP molecules must be combined with magnesium and iron ions to convert a single N2 molecule into two NH3.
Figure Credit:(Mus et al 2018)
“Using wild type and Nase mutant strains of the metabolically versatile photoheterotroph Rhodopseudomonas palustris, we tested the influence of alternative nitrogen fixation on cellular redox homeostasis, biomass composition, and carbon acquisition. Despite assumptions that alternative nitrogen fixation is less efficient than canonical nitrogen fixation, we find that, depending on the carbon source, use of the alternative V-Nase does not necessarily slow photoheterotrophic growth or result in substantially greater H2 production. The data indicate that the metabolic costs of alternative nitrogen fixation could be less significant than previously assumed, possibly explaining why alternative Nase genes persist in diverse diazotroph lineages and are broadly distributed in the environment.” (Luxem et al 2018)
The plant holobiont
Nitrogen fixation will not occur under aerobic conditions, creating an inherent paradox considering that diazotrophs access N2 from the air. As a result, various diazotrophs have developed specialized organelles/architecture to shield nitrogenase from oxygen, whilst allowing for the import of N2 and export of NH3. Legumes (e.g. beans) are the most classical example of a plant which forms a symbiosis with diazotrophic bacteria. The root nodules of legumes house colonies of diazotrophic Rhizobia spp. bacteria. The pink protein leghemoglobin is secreted by the plant’s cells which then absorb and store all the O2 in a localized area. This allows the diazotrophs to have access to fresh supplies of N2 without aerobic inhibition. A similar association occurs between the brown seaweed Sargassum, which forms a unique floating structure to host its associated diazotrophs. Mutualistic interactions such as these allow plants and other organisms to “fuse” with diazotrophs, achieving higher peaks of robust growth.
Figure Credit: (Sigh et al 2019)
Associations between plants/algae/fungi/animals and diazotrophs are far more diverse, complex and intimate then previously appreciated. The rhizosphere refers to host organism architecture (generally on or around roots) specifically devoted to the recruitment and retention of symbiotic diazotrophs. Huang et al 2014 described three major sub-regions of the rhizosphere: The endorhizosphere, the rhizoplane and the ectorhizosphere. These regions are subject to various metabolic secretions from the host. These secretions (amino acids, organic acids, sugars, phenols, polysaccharides, proteins, etc.) mediate the diazotrophic microbial community as well as the multitrophic interactions associated with it.
In plants, the rhizosphere largely exists in the roots but can exist in a variety of structures amongst a variety of organisms. For example, marine sponges and corals (Galaxea, Pavona, Porites, etc.) can host diazotrophs within the deeper crevices of their polyps/mucus/skeleton/pores. Air plants such as Tillandsia and wild maize (Zea mays) secrete mucilage where microanaerobic zones form and foster diazotrophs. Bennet et al 2020 demonstrated how sierra maize obtains 28-82% of its ammonia from atmospheric N2. Additionally, some diazotrophs--certainly including PNS bacteria--confer additional benefits to both aquatic and terrestrial plants by synthesizing phytohormones (e.g. IAA), solubilizing phosphorus and inhibiting pathogenic microbes.
“Traditionally cultivated rice in upland fields and wild rice are likely to harbor unique populations of endophytic bacteria that differ from those in extensively bred modern varieties of rice subjected to the application of various fertilizers and agrochemicals.” (Elbeltagy et al 2000)
A new path forward
Traditional agriculture utilized practices of crop rotation, in which diazotroph-associated crops such as legumes were planted in order to revitalize the soil after nitrogen-intensive crops. While this practice waned in favor of mass monoculture, the best future practice arguably is to maximize diazotroph activity in association with desired/necessary food, biofuel, pharmaceutical and textile crops. This is where PNS bacteria shine brightly! PNSB can survive in a wide variety of aquatic and terrestrial ecosystems, but if starved of NH3, will gradually switch to a diazotrophic metabolism. However, it takes several generations under anaerobic, nutrient starved conditions before PNSB can stabilize into a true ‘diazotrophic community.’
This is where mutualistic relationships between PNSB and crop species become all the more essential to understand and manipulate. Jianbing et al 2018 demonstrated how inoculating Stevia rebaudiana plots with important plant growth-promoting rhizobia (PGPR) Rhodopseudomonas palustris markedly increased soil diversity. Lo et al 2018 identified several stains of PNSB associated with wild Taiwaneese paddy soils; some of these strains enhanced plant uptake of fertilizer while conducting diazotrophy at the same time! Identifying and deploying strains of PNSB best suited to enhance the production of ornamentals, fruits, cereal crops, vegetables, oilseed, legumes, sugars, corn, soy and cannabis is the path towards ever more productive and more sustainable agriculture.
A better understanding of diazotrophs, such as Rhodopseudomonas palustris, is of immeasurable benefit to both humanity and the global ecosystem at large. History has demonstrated how humanity’s past efforts to achieve higher and higher crop yields through disruptive farming practices and excess application of artificial fertilizers has devastated the natural diversity which originally made our lands so fertile. The manufacture and distribution of artificial fertilizers has poisoned the soil, guzzled an ocean of oil and induced harmful algae blooms. Beyond these sins, the Haber-Bosch Process is far too gluttonous in its need for fossil fuels, so much so that there is a pressing economic imperative for it to be replaced. It is the obligation of every nation to pursue food security. This begins with a recognition that farm soil must be treated with the reverence that is customarily bestowed to a rainforest or coral reef. And if the soil’s diversity has been destroyed, then it must be constructed anew.
A bottle of PNSB is poured on scorched earth…
and what was lost in the fire, has been found amongst the ashes.
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