Integrated Environmental Biotechnology 259will be described here only in outline. The process of root nodule formation is
shown diagrammatically in Figure 10.5. In total, there is a limited number of
organisms able to fix nitrogen, all of which are prokaryotes. All living organisms
are dependent ultimately on such organisms, due to a universal and essential
requirement for nitrogen, normally in the form of nitrate, ammonia or ammo-
nium ion or as amino acids from which the amino group may be transferred as
required. Nitrogen is fixed by reduction to ammonia either by free living organ-
isms or by plant symbiots. In both cases, it is essential to have an oxygen free
environment as the enzymes involved in the process are irreversibly inactivated
by the presence of oxygen. Of the free living organisms able to fix nitrogen,
some achieve this naturally while others have to create such an environment.
ClostridiumandKlebsiellaachieve this since they are both anaerobes and so
are already adapted to life in an oxygen-free atmosphere, whileCyanobacteria
andAzobacter have developed means of creating one for themselves.Azobac-
terdoes this by having a very high oxygen consumption rate thus effectively
creating an oxygen-free environment. Other nitrogen-fixing bacteria include the
filamentous bacteria such as theCorynebacteriumspecies, and photosynthetic
bacteria referred to elsewhere, such asRhodospirillum.The latter makes use of
photosynthesis to provide the energy for these reactions and so is presented with
the problem of removing the oxygen produced during photosynthesis away from
nitrogen fixation reactions. Although these free living organisms have a vital role
to play in their particular niches, approximately 10 times more nitrogen is fixed
by plant symbiots. Presumably this is because the plant is better able to provide
the necessary levels of ATP to meet the high energy demands of the process
than are free living bacteria. In addition, the plant supplies the endophytes with
dicarboxylic acids, such as malate and succinate and other nutrients, like iron,
sulphur and molybdenum which is a component of the nitrogen-fixing enzymes.
It also provides its nitrogen-fixing symbiots with an oxygen-free environment,
as described later in this discussion. The analogous chemical process has an
enormous energy requirement to achieve the necessary high temperature, in the
region of 500◦C, and a pressure in excess of 200 atmospheres. This is part of
the reason why manufacture of fertilisers for agricultural purposes, which is in
effect the industrial equivalent of the biological process, is a drain on natural
resources. Together with the unwelcome leaching of surplus fertiliser into water-
ways, causing algal blooms among other disturbances, this makes the widespread
application of fertiliser recognised as a potential source of environmental dam-
age. It is understandable to see a drive towards the engineering of plants both
to increase the efficiency of nitrogen fixation, which is estimated at being 80%
efficient, and to extend the range of varieties, and especially crop species, which
have this capability. It is also worth noting that unlike superficially applied fer-
tilisers which may exceed locally the nitrogen requirement and so leach into the
surrounding waterways, nitrogen fixation by bacteria occurs only in response to
local need and so is very unlikely to be a source of pollution.