Global inputs of biological nitrogen fixation in agricultural systems
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Plant Soil (2008) 311:1–18
DOI 10.1007/s11104-008-9668-3
MARSCHNER REVIEW
Global inputs of biological nitrogen fixation
in agricultural systems
David F. Herridge & Mark B. Peoples &
Robert M. Boddey
Received: 7 March 2008 / Accepted: 22 May 2008 / Published online: 11 July 2008
# Springer Science + Business Media B.V. 2008
Abstract Biological dinitrogen (N2) fixation is a green manure legumes, other literature was accessed
natural process of significant importance in world to obtain approximate estimates in these cases.
agriculture. The demand for accurate determinations Below-ground plant N was factored into the estima-
of global inputs of biologically-fixed nitrogen (N) is tions. The most important N2-fixing agents in agri-
strong and will continue to be fuelled by the need to cultural systems are the symbiotic associations
understand and effectively manage the global N cycle. between crop and forage/fodder legumes and rhizo-
In this paper we review and update long-standing and bia. Annual inputs of fixed N are calculated to be
more recent estimates of biological N2 fixation for the 2.95 Tg for the pulses and 18.5 Tg for the oilseed
different agricultural systems, including the extensive, legumes. Soybean (Glycine max) is the dominant crop
uncultivated tropical savannas used for grazing. Our legume, representing 50% of the global crop legume
methodology was to combine data on the areas and area and 68% of global production. We calculate
yields of legumes and cereals from the Food and soybean to fix 16.4 Tg N annually, representing 77%
Agriculture Organization (FAO) database on world of the N fixed by the crop legumes. Annual N2
agricultural production (FAOSTAT) with published fixation by soybean in the U.S., Brazil and Argentina
and unpublished data on N2 fixation. As the FAO is calculated at 5.7, 4.6 and 3.4 Tg, respectively.
lists grain legumes only, and not forage, fodder and Accurately estimating global N2 fixation for the
symbioses of the forage and fodder legumes is
Responsible Editor: Yongguan Zhu. challenging because statistics on the areas and
productivity of these legumes are almost impossible
D. F. Herridge
to obtain. The uncertainty increases as we move to the
New South Wales Department of Primary Industries,
4 Marsden Park Rd, other agricultural-production systems—rice (Oryza
Calala, NSW 2340, Australia sativa), sugar cane (Saccharum spp.), cereal and
oilseed (non-legume) crop lands and extensive,
M. B. Peoples
grazed savannas. Nonetheless, the estimates of annual
CSIRO Plant Industry,
P.O. Box 1600, Canberra, ACT 2601, Australia N2 fixation inputs are 12–25 Tg (pasture and fodder
legumes), 5 Tg (rice), 0.5 Tg (sugar cane),2 Plant Soil (2008) 311:1–18
would be reduced with the publication of more respectively. The latter estimate was revised down-
accurate statistics on areas and productivity of forage wards at an international conference in Sweden soon
and fodder legumes and the publication of many more afterwards to 122 Tg N, principally by downgrading
estimates of N2 fixation, particularly in the cereal, inputs of fixed N in forests and natural grasslands.
oilseed and non-legume crop lands and extensive Burris (1980) accepted this amended figure of
tropical savannas used for grazing. 122 Tg N fixed annually and noted that it seemed
to be compatible with the published values for the
Keywords Associative . Cyanobacteria . global carbon (C) cycle. The global N2 fixation
Dinitrogen (N2) fixation . Endophytic . Free-living . estimates of Delwiche (1970), Burns and Hardy
Global . Legumes . Nitrogen (N) . Oilseed legumes . (1975) and Burris (1980) have been widely quoted
Pulses . Rhizobia . Soybean ever since. Note that these estimates cover both
agricultural and natural systems, including marine,
and were largely derived using acetylene (C2H2)
Introduction reduction, N difference and N balance methodologies.
The different N2-fixing organisms and symbioses
Just over 25 years ago, Bob Burris wrote a paper entitled found in agricultural and terrestrial natural ecosystems
“The global nitrogen budget—science or séance?” in are shown in Fig. 1.
which he discussed the challenges of scaling up plot New figures for global N2 fixation have been
measurements of dinitrogen (N2) fixation and other published more recently (e.g. Galloway et al. 1995;
nitrogen (N) flows to calculate global N budgets Smil 1999) and are also widely quoted (Vitousek et
(Burris 1980). With tongue in cheek, he suggested al. 1997; Boyer et al. 2004; Galloway et al. 2004;
that potential authors could use a variety of methods to Mosier et al. 2004). Galloway et al. (1995) and Smil
fill in the values in the N cycle, from gazing at crystal (1999) estimated global N2 fixation for cultivated
balls, consulting sages to cranking out computer- agricultural systems, i.e. excluding the extensive
generated random numbers. He did acknowledge, tropical savannas, at 43 Tg (range 32–53 Tg) and
however, that the common method was to consult the 33 Tg (range 25–41 Tg) annually. Cleveland et al.
literature, choose the data that seem to make sense, (1999) estimated terrestrial global N2 fixation by
then construct the budget accordingly. considering 23 biome types covering the whole
Delwiche (1970) and Burns and Hardy (1975) had planet, but did not consider the extent of agricultural
previously estimated annual, global biological N2 activity in these biomes, or the presence of cultivated
fixation at 100 and 175 million tonnes (Tg) N, legumes capable of large per ha inputs of N2 fixation.
Fig. 1 Biological N2-fixing
agents in agricultural and Biological Nitrogen Fixation
terrestrial natural systems
Agricultural systems Natural systems
Crop Pastures & Fodder
Plant-associated Plant-associated Plant-associated
legume-rhizobia (symbiotic) legume-rhizobia (symbiotic) legume-rhizobia (symbiotic)
Azolla-cyanobacteria (symbiotic) cereal-associative bacteria nonlegume-Frankia (symbiotic)
cereal-associative bacteria cereal-endophytic bacteria Azolla-cyanobacteria (symbiotic)
cereal-endophytic bacteria cycad-cyanobacteria (symbiotic)
cereal-associative bacteria
Free-living
Free-living cereal-endophytic bacteria
cyanobacteria
cyanobacteria heterophic bacteria
heterophic bacteria
autotrophic bacteria
Free-living
autotrophic bacteria cyanobacteria
heterophic bacteria
autotrophic bacteriaPlant Soil (2008) 311:1–18 3
Galloway et al. (2004) covered all aspects of the N Measurement of N2 fixation
cycle and incorporated estimates of N2 fixation in
cultivated agricultural systems (32 Tg N/year) using Notwithstanding the difficulties and errors, the demand
data from earlier papers (Galloway et al. 1995; Smil for accurate determinations of global inputs of biolog-
1999). ically-fixed N is strong and will continue to be fuelled
In this review we reconsidered N2 fixation inputs by the need to understand and effectively manage the
into agricultural systems. As in past reviews (e.g. global N cycle. There are five basic methodologies
Smil 1999), we included cultivated land used for available to quantify biological N2 fixation:
agriculture, but also included uncultivated agricul-
1. The enzyme nitrogenase, universally responsible for
tural lands, such as the tropical savannas used for
biological N2 fixation, is also capable of reducing
grazing. Our strategy was to combine data on the
acetylene (C2H2) to ethylene (C2H4). Both gases
areas and yields of legumes and cereals from the
can be readily detected and quantified using gas
Food and Agriculture Organization (FAO) database
chromatography (Schollhorn and Burris 1967;
on world agricultural production (FAOSTAT) with
Hardy et al. 1968). Thus, the C2H2 reduction assay
published and unpublished data on N2 fixation. As
is a sensitive measure of nitrogenase activity at a
the FAO lists grain legumes only, and not forage,
point in time and can be very useful for detecting
fodder and green manure legumes, other literature
N2 fixation activity of, for example, bacterial
was accessed to obtain approximate estimates in
cultures or plant residues that may be harbouring
these cases.
N2-fixing bacteria. However, in enclosing the
The difficulties and potential errors in calculating
particular agent in a gas-tight vessel to evaluate
N2 fixation at global scales are magnified substantial-
ethylene (C2H4) evolution, physical disturbance of
ly when moving from agricultural systems to the
the N2-fixing species is almost inevitable and this
natural systems. The agents of N2 fixation are
results in a decline in activity (Minchin et al. 1986;
essentially the same as in agricultural systems,
Boddey 1987). Even the partial substitution of N2
although the species may be different. The main
by C2H2 is sufficient to reduce N2-fixing activity
problems are the uncertainty in estimating N2 fixation
(Minchin et al. 1983). Scaling up point-source
intensity per unit area, the likely bias of those
C2H2 reduction values to account for spatial and
estimates, and the difficulty in scaling up because of
temporal variations and converting them to
uncertainties in spatial coverage of the putative N2-
amounts of N fixed is difficult, if not impossible,
fixing species. Galloway et al. (2004) stated: “In a
and is not recommended.
recent compilation of rates of natural biological
Hydrogen is an obligate product of N2 reduction
nitrogen fixation (BNF) by Cleveland et al. (1999),
and its measurement can also be used to assay
symbiotic BNF rates for several biome types are based
nitrogenase activity (Hunt and Layzell 1993;
on one-to-few published rates of symbiotic BNF at the
Dong et al. 2001). However, the method has
plot scale within each particular biome. For example,
never been applied as a routine field assay owing
based on a few estimates of symbiotic BNF available
to practical difficulties.
for tropical rain forests, estimated BNF in these
systems represents ∼24% of total natural terrestrial 2. The total N-balance method is based on the
BNF globally on an annual basis (Cleveland et al. principal that the plant/soil system will accumu-
1999). While the relative richness of potential N2- late N over time if there is an input of N2 fixation.
fixing legumes in tropical forests suggests that symbi- However, measures of N2 fixation may be under-
otic BNF in these systems is relatively high (Crews estimated because of N losses from the system
1999), the paucity of actual BNF rate estimates in these during the period of study through ammonia
systems suggest caution when attempting to extrapo- volatilisation, denitrification, leaching etc, or
late plot scale estimates of BNF and highlights the confounded by other external inputs of N unre-
difficulties to attempting to estimate natural BNF at the lated to N2 fixation (e.g. N dissolved in rainfall, N
global scale.” Because of the uncertainties, we have in dust, gaseous N etc). Hence N balance requires
not attempted in this review to quantify global N2 measurements of as many potential N inputs and
fixation in natural systems. outputs as possible. The time-frame is generally4 Plant Soil (2008) 311:1–18
several years because of the need to measure incorporation—Warembourg et al. 1982) fol-
incremental changes in the N content of the soil lowed by measurement of incorporation of 15N
against large background amounts (Peoples and by the plants, and (ii) growing the plants in 15N-
Herridge 1990; Giller and Merckx 2003). Clearly enriched soil or other growth medium (15N
the methodology is technically challenging, requir- isotope dilution—McAuliffe et al. 1958; Chalk
ing substantial inputs of labour for long periods. 1985) and calculating the extent of dilution of 15N
Additionally, errors in quantifying the N fluxes, in the plants by atmospheric (fixed) 14N. A later
and inaccuracies in sampling and analysing soil for variation of 15N isotope dilution utilised the
changes in total N and bulk density, can introduce natural 15N enrichment of soils, thereby avoiding
substantial uncertainties into the final estimates of the need to add 15N-enriched materials (natural
15
N2 fixation (Chalk 1998). The N balance method N abundance—Shearer and Kohl 1986).
was more commonly used some time ago (e.g. The 15N2 incorporation method is limited in
Vallis 1972; Wetselaar et al. 1973), but in recent application to short experimental periods in a
years has been largely replaced by 15N and ureide laboratory or growth chamber. 15N isotope dilution
methods, described below. with artificial enrichment of soil was, until a few
3. A simple variation of N balance for quantifying N2 years ago, used widely to quantify N2 fixation in
fixation is N difference. With this method, total N agricultural systems (Chalk and Ladha 1999),
accumulated by N2-fixing plants is compared with although rarely on-farm in unreplicated, non-
that of neighbouring non N2-fixing plants, with the experimental studies. In recent years, natural 15N
difference between the two assumed to be due to N2 abundance has gained prominence for work in
fixation. The main assumption is that the N2-fixing both experimental plots and in farmers’ fields,
plants assimilate the same amount of soil mineral owing to the greater accessibility of scientists to
N as the neighbouring non N2-fixing plants. In high-precision, automated isotope-ratio mass spec-
soils of limited N supply, this method can be used trometers. Although natural 15N abundance has
with considerable success, especially if the N2- been widely utilised in agricultural settings, there
fixing plants derive large amounts of N from N2 are a number of potential limitations that restrict its
fixation. It may be less useful in moderate-to-high application in natural ecosystems (Boddey et al.
N soils because differences between N2-fixing and 2000). In those systems, estimates of the percent-
non N2-fixing plants in root morphology and age of plant N derived from N2 fixation (%Ndfa)
rooting depth can result in different capacities to may not be possible owing to the large spatial
exploit soil N (Herridge et al. 1995; Chalk 1998). variability, diversity and complexity of available-N
It is also of limited value for on-farm surveys pools in the soil with different 15N signatures (e.g.
where appropriate non N2-fixing plants may not be Pate et al. 1993; Gehring and Vlek 2004).
present. Good examples of the application of this 5. The ureide method (McClure et al. 1980; Herridge
technique were published in the 1960–1970s and Peoples 1990) exploits the fact that many of
(Weber 1966; Bell and Nutman 1971). As with N the agronomically-important legumes of tropical
balance, this method has been largely replaced by origin (e.g. soybean [Glycine max], common bean
15
N and ureide methods. [Phaseolus vulgaris], Desmodium spp.) export
4. The heavy isotope of nitrogen, 15N, was first used allantoin and allantoic acid (collectively known
to evaluate N2 fixation by bacteria in the 1940s as ureides) as the products of N2 fixation from
(Burris et al. 1942), but the availability of their nodules to the shoots. In these legumes, the
materials enriched with 15N and mass spectrom- ratio of ureide N to total N in xylem sap or stem
eters to analyse the samples severely restricted its segments is highly correlated with %Ndfa. Al-
general application for many years. That situation though not applicable to all legumes, and to no
started to change in the 1970s, facilitating more other N2-fixing associations, the technique has
widespread use of 15N-based methodologies been widely used with both experimental and non-
during the 1980s and beyond. Experimental experimental (farmer) crops. The analytical proce-
protocols involved: (i) labelling N2 in the atmo- dures are simple with minimal requirements for
sphere surrounding the N2-fixing plants (15N2 sophisticated or expensive equipment.Plant Soil (2008) 311:1–18 5
The principles behind these methods and how to estimates of N2 fixation of crop legumes in agricul-
use them effectively have been described in varying tural systems are likely to be sound because they draw
degrees of detail in a substantial number of publica- on many hundreds of individual values of %Ndfa and
tions for nodulated legumes (e.g. Chalk 1985; Shearer the annual area and production statistics of the FAO,
and Kohl 1986; Witty and Minchin 1988; Witty et al. published as FAOSTAT (Table 1). FAOSTAT is the
1988; Peoples and Herridge 1990; Hardarson and web-based statistical database of the FAO (http://
Danso 1993; Danso et al. 1993; Vessey 1994; faostat.fao.org) covering many aspects of world
Unkovich and Pate 2000; Giller 2001; Peoples et al. agriculture, including crops in the section Produc-
2002; Unkovich et al. 2008), and associative and free- tion/Crops. Estimates of N2 fixation of forage and
living N2-fixing agents (Boddey 1987; Chalk 1991; fodder legumes will be less reliable because global
Boddey et al. 2001; Giller 2001; Giller and Merckx areas of land with forage and fodder legumes are
2003; Unkovich et al. 2008). The N balance and N difficult to assemble as are estimates of %Ndfa of
difference methods provide estimates of N2 fixation legumes in those lands.
on an area basis, i.e. kg N/ha. The 15N and ureide The most reliable information on the other N2-
methods, on the other hand, provide estimates of % fixing agents in agricultural systems—the azolla/
Ndfa, i.e. the percentage of total N of the organism cyanobacteria association, free-living cyanobacteria
(bacteria, plant) that is derived from N2 fixation. An and other autotrophic bacteria, and the numerous
amount of N2 fixed per unit area or unit of production genera of heterotrophic bacteria that utilise either
can only be calculated when %Ndfa is combined with C-rich exudates of living plants or degrading crop
an estimate of organism biomass and total N content. residues as energy sources—are the areas in which
Although all methods have their unique limitations they potentially exist. For example, the FAOSTAT
and sources of error, the N balance, N difference, 15N database can provide figures for the global area and
(isotope dilution and natural abundance) and ureide production of rice (Oryza sativa) that can be
methods arguably represent the best of what is combined with published estimates of N2 fixation of
currently available. free-living cyanobacteria and the azolla–cyanobacteria
association to calculate potential N2 fixation in this
system (Smil 1999). Similarly, FAOSTAT can also
Reliability of current estimates of N2 fixation provide accurate data on areas and production of
in the different agricultural systems sugarcane (Saccharum sp.) for calculating potential
N 2 fixation of the endophytic and associative
The key ingredients for accurately estimating N2 bacteria in this particular system. To calculate actual,
fixation at any scale—unit area (m2 or ha), individual rather than potential, N2 fixation is far more difficult
field, catchment, region, country, globe—are reliable because of the uncertainty in determining the
values for %Ndfa and total N accumulation of the N2- occurrence and activity of the N2-fixing agents
fixing agent for a specific period of time. Thus, global across the global reach of these systems (Table 1).
Table 1 Assessments of the reliability of estimating %Ndfa and total N of the different N2-fixing agents in agricultural systems (the
more +++ the better)
N2-fixing agent Agricultural system Reliability in Reliability in estimating
estimating %Ndfa total N of the N2-fixing
agent globally
Legume–rhizobia Legume cropping +++++ +++++
Legume–rhizobia Pasture/fodder +++++ +++
Azolla–cyanobacteria, cyanobacteria Rice ++++ +++
Endophytic, associative and free-living bacteria Sugar cane ++ ++
Endophytic, associative and free-living bacteria Other cropping lands + +
Endophytic, associative and free-living bacteria Extensive tropical savannas + +
used for grazing6 Plant Soil (2008) 311:1–18
Below-ground N—the underestimated component pulse and oilseed legumes, soybean, faba bean (Vicia
of N2-fixing plants faba), chickpea (Cicer arietinum), mungbean (Vigna
radiata), narrow-leafed lupin (Lupinus angustifolius),
The majority of published values for legume N2 fixation pea (Pisum sativum) and pigeonpea (Cajanus cajan),
were based on shoots only. Fixed N contained in and 34–68% for the pasture/fodder legumes, subter-
attached and detached roots and nodules, and rhizode- ranean clover (Trifolium subterraneum), serradella
position was essentially ignored (e.g. Evans and (Ornithopus compressus), white clover (Trifolium
Herridge 1987; Danso et al. 1993; Unkovich et al. repens) and alfalfa (Medicago sativa) (Zebarth et al.
1997; Smil 1999; Carlsson and Huss-Danell 2003; 1991; Russell and Fillery 1996b; McNeill et al. 1997;
Russelle and Birr 2004). In other reports, a factor was Jørgensen and Ledgard 1997; Rochester et al. 1998;
used to account for below-ground N (BGN), usually Khan et al. 2002, 2003; Yasmin et al. 2006; Mahieu et
based on a published or experimentally-determined al. 2007; McNeill and Fillery 2008).
value derived from the physical recovery of roots (e.g. Clearly, there is no single value for BGN, with the
Herridge et al. 1995; Evans et al. 2001). We are now variation in published estimates reflecting effects of
starting to see a change, however, with acknowledge- species, soil and climate on the partitioning of N within
ment that published values for legume N2 fixation are the plant. To account for BGN when calculating N2
low because they do not account for the large fixation, we used a multiplication factor of 2.0 for the
proportion of below-ground N contained in non- pasture/fodder legumes and chickpea (assumes 50% of
recovered roots, detached nodules, and products of root plant N is below-ground), 1.5 for soybean (assumes
and nodule necrosis (Carlsson and Huss-Danell 2003; 33% BGN) and 1.4 for the remainder of the pulse and
Crews and Peoples 2005; McNeill and Fillery 2008). oilseed legumes (assumes 30% BGN). Although these
For example, Walley et al. (2007) assumed root N was factors are approximations, we would argue that the
14% of total plant N and rhizodeposited N an additional errors associated with their use are far less than the
10% when calculating N2 fixation of the pulse legumes errors associated with ignoring BGN or using values
in the Northern Great Plains of North America. This for physically-recovered roots. It is also worth noting
change in thinking has been brought about by advances that reported BGN values for non-legumes, such as
in methodologies for estimating BGN. wheat and barley, are similar to those of the legumes.
In the past, the most simple and commonly-used For example, Khan et al. (2003) estimated BGN of
method for determining BGN was to physically remove field-grown barley (Hordeum vulgare) at 30%.
roots from the soil. Values for BGN as a percentage of
total plant N were usuallyPlant Soil (2008) 311:1–18 7
Table 2 Average values for %Ndfa for the major crop legumes in experiments and farmers’ fields
Legume Experimentsa Farmers’ fieldsb
%Ndfa range %Ndfa average %Ndfa average
Common bean 0–73 40 36
Chickpea, lentil, pea, cowpea, mungbean, pigeonpea etc 8–97 63 65
Soybean, groundnut 0–95 68 58
Fababean, lupin 29–97 75 68
a
Collated from Peoples et al. (2008) in which data from a number of reviews and experimental papers were summarised with
additional information on N2 fixation of common bean from Rennie and Kemp (1982a, b) and Hardarson et al. (1993)
b
Sourced from Peoples et al. (2008), comprising >800 determinations
papers were summarised (Peoples and Craswell 1992; for the farmers’ fields are in reasonable agreement with
Herridge and Danso 1995; Peoples et al. 1995; Wani et the experimental data and support three of the four
al. 1995; Jensen 1997; Unkovich et al. 1997; Schulz et groupings of the crop legumes. The %Ndfa values for
al. 1999; Unkovich and Pate 2000; Giller 2001; soybean in farmers’ fields are lower than those in the
Rochester et al. 2001; Turpin et al. 2002; Aslam et experiments, principally reflecting the regions in which
al. 2003; Shah et al. 2003). Additional information on these particular crops were grown. Only 21 of the 133
N2 fixation of common bean was sourced from Rennie estimates were from Brazil and none were from
and Kemp (1982a, b) and Hardarson et al. (1993). We Argentina. The two countries together grow >40% of
grouped the legumes according to their ability to fix N the world’s soybean with relatively high %Ndfa values
in experiments. Common bean has the lowest capacity (Alves et al. 2003; Hungria et al. 2005) (see also
for N2 fixation and is in a group by itself, with an Table 3).
average Ndfa of 40%. The next group includes most of To differentiate %Ndfa for the different legumes at
the winter and summer pulses, with an average Ndfa of smaller scales, i.e. field, catchment, region, according
63%. The third group includes soybean and groundnut to local soil and plant-growth conditions and then
(Arachis hypogaea), with Ndfa of 68% and the final aggregate those estimates to generate country and
group includes faba bean and lupin (Lupinus spp.) with global values would be extremely difficult and may
Ndfa of 75%. The ranges of values within the four not improve accuracy. Having said that, %Ndfa of
groups are large and reflect variations in legume soybean needs to be differentiated for the principal
growth, set by genetic, agronomic, environmental and soybean-producing countries as this crop is respon-
experimental factors, the availability of soil mineral N sible for most of the N fixed by legumes, and there
and numbers and effectiveness of rhizobia in the are considerable differences in soil type, climate and
vicinity of the growing root system. The groupings plant-cultural practices amongst those countries
are reasonably consistent with those described by (Table 3).
Hardarson and Atkins (2003) for food legumes In the U.S., soils used for soybean production tend to
involved in FAO/International Atomic Energy Agency be fertile, with moderate-high concentrations of clay,
co-ordinated research programs across a number of organic matter and plant-available N (e.g. Russelle and
countries and with those of Walley et al. (2007) for the Birr 2004). As a result, reported Ndfa values mostly
pulse legumes in the Northern Great Plains of North range between 40% and 80% (van Kessel and Hartley
America. 2000; Peoples et al. 2008; Salvagiotti et al. 2008), with
Average %Ndfa values for legumes growing in >800 an overall average value of 60%.
farmers’ fields in Europe, Africa, Asia, South America The average Ndfa value for soybean in Brazil is
and Australia are shown in the final column, Table 2. calculated at 80%, reflecting the widespread use of
Values were taken from Peoples et al. (2008) using rhizobial inoculants, the high N demand of the crops
data sourced from Rupela et al. (1997), Rochester et al. (about 300 kg N/ha) coupled with low inputs of fertiliser
(1998), Schwenke et al. (1998), Maskey et al. (2001), N, and the high proportion of the crops that are no-tilled
Peoples et al. (2001), Hiep et al. (2002), Hoa et al. (Hungria and Vargas 2000; Hungria et al. 2005, 2006;
(2002) and Herridge et al. (2005). The %Ndfa values Alves et al. 2003; FAOSTAT). Alves et al. (2003) and8 Plant Soil (2008) 311:1–18
Table 3 Estimates of amounts of N fixed annually by soybean in the major soybean-producing countries, using FAO statistical data
for 2005 (FAOSTAT), estimates of country-specific %Ndfa, and estimates of harvest index, %N shoots and below-ground N as % of
total crop N
Country Area (Mha) Grain yield (Tg) Shoot DM (Tg)a Shoot N (Tg)b Crop N (Tg)c %Ndfa Crop N fixed (Tg)
U.S. 30.0 85.0 212.6 6.38 9.56 60 5.74
Brazil 22.9 51.2 128.0 3.84 5.76 80 4.61
Argentina 14.0 38.3 95.8 2.87 4.31 80 3.44
China 9.6 16.8 42.0 1.26 1.88 50 0.95
Soybean 93.4 214.8 537.1 16.12 24.17 68 16.44
a
Using harvest index (grain dry matter as a proportion of total above-ground dry matter) value of 0.4 (Jefing et al. 1992; Herridge and
Holland 1992; Guafa et al. 1993; Herridge and Peoples 2002; Shutsrirung et al. 2002; Gan et al. 2002, 2003; Salvagiotti et al. 2008)
b
Using %N shoots of 3.0% (Herridge et al. 1990; Herridge and Holland 1992; Herridge and Peoples 2002; Shutsrirung et al. 2002;
Gan et al. 2002, 2003; Salvagiotti et al. 2008)
c
Multiplying shoot N by 1.5 (Rochester et al. 1998)
others (see review by van Kessel and Hartley 2000) 0.54 Tg fertiliser N was applied to 10.5 Mha soybean
reported consistent increases in nodulation and N2 and groundnut in 1994. The fertiliser N inputs plus
fixation of no-tilled soybean compared with crops grown residual mineral N in the soil from previous crops
under cultivation. The increases under no till were would depress N2 fixation activity substantially. Thus,
thought to be due principally to reduced levels of nitrate we estimate the average Ndfa value for China at 50%
coupled with improved moisture conditions in the soil. (Ruiz Sainz et al. 2005).
Thus, Alves et al. (2003) reported that Brazilian soybean The total amount of N2 fixed by soybean for each of
derived 70–85% of crop N from N2 fixation, equivalent the four major soybean-producing countries can now
to 70–250 kg N/ha. In the case of high-yielding crops, be estimated by combining the %Ndfa values with
i.e. >4.0 t/ha, as much as 350–400 kg N/ha may be production statistics from FAOSTAT. First, the total
fixed. Similarly, Hungria et al. (2005) reported Ndfa amount of soybean N is calculated by dividing the
values of 69–94% for inoculated soybean in Brazil. FAOSTAT crop production data (Column 3, Table 3)
There are very few reports quantifying N2 fixation of by an average harvest index value (0.4) to determine
soybean in Argentina. Published Ndfa values are 30– shoot dry matter (DM) (Column 4). Shoot N (Column
70% (Garcia 2004) and 40–50% (Gutiérrez-Boem et al. 5) and crop N (Column 6) are then calculated using
2004; Di Ciocco et al. 2004), but these estimates were 3% for the N concentration of shoots and a multipli-
from experimental sites and not farmer’s fields. cation factor of 1.5 to account for below-ground N
However, Argentinian soybean farmers, like the Brazil- (Rochester et al. 1998). Crop N fixed (final column) is
ian farmers, commonly use inoculants and no-tillage then determined as crop N×%Ndfa. Thus, estimates of
production systems with negligible fertiliser N (Garcia total crop N fixed by soybean range between 0.95 Tg
2004; Hungria et al. 2005; Peloni 2006; FAOSTAT). annually for China to 3.4 Tg for Argentina, 4.6 Tg for
Garcia (2004) also noted that most of the soils used for Brazil and 5.7 Tg for the U.S.
soybean production in Argentina have nutrient defi- We used the same series of calculations to estimate
ciencies, including N. Taken together, these reports global N2 fixation of the major pulse and oilseed
suggest that the high N demand crops would need to fix legumes (Table 4). The final column contains the calcu-
a large proportion of their N requirements. We therefore lated values for annual crop N fixed for each species
assume the same average Ndfa value for soybean in plus total values for the pulse legumes (2.95 Tg), oilseed
Argentina as for soybean in Brazil, i.e. 80%. legumes (18.5 Tg) and all crop legumes (21.5 Tg).
Chinese farmers reportedly apply fertiliser N to In a previous publication we calculated global N2
soybean and rely on the naturalised soil rhizobia to fixation by the pulse and oilseed legumes by using
nodulate the crops rather than use inoculants (Gan et estimates of average amounts of N fixed per unit
al. 2002; Ruiz Sainz et al. 2005). P.W. Singleton shoot biomass (Peoples et al. 2008). This approach
(personal communication) estimated that about was based on the observation that amounts of N2Plant Soil (2008) 311:1–18 9
Table 4 Estimates of amounts of N fixed annually by the major pulse and oilseed (crop) legumes, using FAO statistical data for 2005
(FAOSTAT), values for average %Ndfa from Table 2 and estimates of values for harvest index, %N shoots and below-ground N as %
of total crop N
Legume Area (Mha) Grain yield (Tg) Shoot DM (Tg)a Shoot N (Tg)b Crop N (Tg)c %Ndfa Crop N fixed (Tg)
Common bean 25.1 18.1 51.7 1.03 1.45 40 0.58
Cowpea 9.2 4.6 13.3 0.27 0.37 63 0.23
Chickpea 10.4 8.4 23.9 0.48 0.96 63 0.60
Pea 6.6 11.3 32.3 0.65 0.90 63 0.57
Lentil 4.1 4.1 11.8 0.24 0.33 63 0.21
Fababean 2.7 4.3 12.4 0.27 0.38 75 0.29
Other pulses 11.4 9.4 26.8 0.54 0.75 63 0.47
Total pulses 69.7 60.2 171.9 3.48 5.14 57 2.95
Groundnut 23.4 37.6 93.9 2.16 3.03 68 2.06
Soybean 93.4 214.8 537.1 16.11 24.17 68 16.44
Total oilseeds 116.7 252.4 707.8 18.27 27.20 68 18.50
Total crop legumes 186.4 312.6 879.7 21.75 32.34 66 21.45
a
Using harvest index (grain dry matter as a proportion of total above-ground dry matter) values of 0.4 for groundnut and soybean and
0.35 for the remainder (see references in footnote Table 3; also Schwenke et al. 1998; Evans et al. 2001; Hiep et al. 2002; Hoa et al.
2002; MJ Unkovich, personal communication)
b
Using %N shoots of 3.0% for soybean, 2.3% for groundnut, 2.2% for fababean and 2.0% for the remainder (see references in
footnote Table 3; also Schwenke et al. 1998; Evans et al. 2001; Hiep et al. 2002; Hoa et al. 2002)
c
Multiplying shoot N by 2.0 (chickpea), 1.5 (soybean) and 1.4 (remainder) to account for below-ground N.
fixed by legumes in any agroecosystem were primarily Comparisons of the Smil (1999) estimates of legume
regulated by plant growth and DM production. The N2 fixation (area basis, kg N/ha) and estimates using
provisos were that effective rhizobia were present in the the data in Table 3 are shown in Table 5.
soil and concentrations of soil mineral N were not There is generally good agreement between the Smil
excessive. Data collected from both experimental trials (1999) values for crop N2 fixed (kg/ha) and our values
and farmers’ crops indicated that crop legumes generally calculated from Table 4, except for soybean and pea
fix 15–25 kg shoot N for every Mg shoot DM (Table 5). The difference in the case of soybean can be
accumulated, with averages of 20 kg shoot N/Mg shoot explained by the recent expansion of production in
DM (Fig. 2; see also Evans et al. 2001; Maskey et al. Argentina and Brazil where the use of fertiliser N is low,
2001; Peoples et al. 2001). Fixed N associated with the inoculation is widespread and the N demands of the
nodulated roots increased the value to 30 kg total crop predominantly no-tilled crops are large because of rela-
N/Mg shoot DM. Common bean, chickpea and soybean tively high grain yields (2.73 Mg/ha for Argentina and
were identified as the exceptions, with values for 2.23 Mg/ha for Brazil, FAOSTAT for 2005). The long-
common bean of 15 kg total crop N fixed/Mg shoot standing notion that soybean fix, on average, about 50%
DM, and for chickpea and soybean of 40 kg crop N of their N needs would appear to be no longer valid.
fixed/Mg shoot DM. We used these values to calculate Smil (1999) estimated crop legumes to fix a total
global N2 fixation of 4 and 18 Tg N (total 22 Tg N) of 10 Tg N annually, compared with our estimate of
annually by the pulses and oilseed legumes, respective- 21.5 Tg annually. As mentioned above, the discrep-
ly, using FAOSTAT production statistics for 2000–2004. ancy results mainly from the different values of %
Smil (1999) used yet another approach to calculate Ndfa for pea and soybean, our inclusion of estimates
average annual values for global N2 fixation by the of below-ground fixed N associated with, or released
crop legumes. Ranges of values (minimum, mean, from, roots and nodules, and the use of updated
maximum) for crop N fixed for each species were FAOSTAT statistics, i.e. 2005 data used for calcu-
estimated on an area basis (kg N/ha), then applied to lations in Tables 3 and 4 compared with mid 1990s
the global areas of the legumes from FAOSTAT. data used by Smil (1999).10 Plant Soil (2008) 311:1–18
Fig. 2 Examples of the re-
lationship between amounts
of N2 fixed (as kg N/ha in
shoots) and shoot dry matter
(Mg/ha) for crop legumes
growing in different geo-
graphic regions. Data
includes both rainfed and
irrigated cool-season (open
circles) and warm-season
legumes (closed triangles).
The dashed lines indicate 15
and 25 kg N fixed per Mg
dry matter. Relationship
modified from Peoples et al.
(2008) who used published
and unpublished data col-
lated from studies undertak-
en in the Middle East and
Asia (Syria, Nepal, Paki-
stan, Thailand), Oceania
(Australia), South America
(Brazil), North America
(Canada and USA), and
Europe (Austria, Denmark
and France)
Forage/fodder legumes–rhizobia to obtain. Smil (1999) reported 100–120 Mha of land
in fodder and forage legumes and green manure
Accurately estimating global N2 fixation for the crops. He assumed average annual N2 fixation rates of
symbioses of the forage and fodder legumes is 200 kg N/ha for alfalfa, 150 kg N/ha for the clovers
challenging because statistics on the areas and (Trifolium spp.), 100 kg N/ha for other forages and
productivity of these legumes are almost impossible 50 kg N/ha for legume–grass pastures. Thus, total N2
Table 5 Comparing estimates of N2 fixation/unit area (kg/ha) by Smil (1999) with estimates calculated from legume global areas
(Table 4, column 2) and crop N fixed (Table 4, column 8)
Legume Smil (1999) ranges of values (kg N/ha/year) Calculated from
Table 4 (kg N/ha/year)
Minimum Mean Maximum
Common bean 30 40 50 23
Chickpea 40 50 60 58
Pea 30 40 50 86
Lentil 30 40 50 51
Fababean 80 100 120 107
Other pulses 40 60 80 41
Groundnut 60 80 100 88
Soybean 60 80 100 176Plant Soil (2008) 311:1–18 11
fixation for the forage and fodder legumes was forage and fodder legumes can be calculated by
calculated at 12 Tg annually (average of about combing the overall annual production of 500 Tg
110 kg N/ha/year) (Table 6). with the rate of N2 fixation per unit of forage (50 kg
A substantial body of work in Australia and N fixed/Mg shoot biomass). Thus, a value of 25 Tg
northern Europe shows that forage/fodder legumes N/annually is obtained, a value about double that of
have an average Ndfa value of about 70% and 25 kg Smil (1999).
N is fixed in the shoots for every Mg shoot biomass The same value of 25 Tg N can be calculated if the
produced (Peoples and Baldock 2001; Carlsson and following figures and assumptions are used: globally
Huss-Danell 2003). It should be noted that Peoples 110 Mha legumes with an average Ndfa of 70%,
and Baldock (2001) reported wide variations for this average shoot DM production of 4.5 Mg/ha, shoot N
value, ranging 8–53 kg shoot N fixed/Mg shoot concentration of 3.6% and below-ground N of 50%.
biomass. Such variation would have been caused by Thus, average annual N2 fixation is calculated at
differences in soil nitrate levels and pasture vigour, as 227 kg/ha and global N2 fixation at 25 Tg.
well as species differences in foliage-N content, So, what is a realistic figure for N2 fixation by the
experimental treatment and error. Assuming 50% of forage and fodder legumes in agricultural systems?
forage legume nitrogen is below-ground (McNeill et The Smil (1999) figure of 12 Tg annually may be low
al. 1997; Peoples and Baldock 2001), the overall because it does not reasonably account for below-
average for N2 fixation by forage legumes becomes ground N, but without reliable data on global forage
50 kg N fixed/Mg shoot biomass. and fodder legume areas and production statistics for
Smil (1999) estimated global shoot productivity of those areas, it is impossible to provide an alternative.
the forages at 500 Tg from the 100–120 Mha, The real figure may lie somewhere between 12 and
equivalent to 4.2–5.0 Mg/ha. Global N fixed by the 25 Tg annually (Table 6).
Table 6 Summary of estimates of N fixed annually in agricultural systems by rhizobia in symbiosis with crop, pasture and fodder
legumes, numerous genera of bacteria associated with non-leguminous species and free-living bacteria
Agent Agricultural system Areaa Rate of N2 Crop N Comments on validity of
(Mha) fixation fixed global N2 fixation estimates
(kg N/ha/year) (Tg/year)
Legume–rhizobia Crop (pulse and 186 115 21 May be a robust estimate and substantially
oilseed) legumes higher than the Smil (1999) estimate
of 10 Tg fixed
Legume–rhizobia Pasture and 110 110–227 12–25 Difficult to accurately assess because of
fodder legumes uncertainty in legume areas and
production
Azolla– Rice 150 33 5 Smil (1999) estimate of 5 Tg
cyanobacteria, N/year reasonable, although primarily
cyanobacteria based on C2H2 reduction technique
Endophytic, associative Sugar cane 20 25 0.5 Large variations in apparent N2 fixation,
& free-living bacteria using natural 15N abundance, make
estimations difficult and speculative
Endophytic, associative Crop lands other 80012 Plant Soil (2008) 311:1–18
Azolla–cyanobacteria and free-living (1997) study of 50 Brazilian sugar cane crops, the
cyanobacteria in rice paddies overall average δ15N value for the cane was +5.32‰
(range +2.0‰ to +11.0‰), compared with +6.13‰
Smil (1999) estimated N2 fixation by free-living (range −0.4‰ to +12.9‰) for the reference samples.
cyanobacteria and cyanobacteria in symbiosis with An aggregated estimate of Ndfa, using those average
the water fern Azolla at 4–6 Tg annually. Estimates values, is just 13%.
were based on rates of N2 fixation of 20–30 kg N/ha Boddey et al. (2001) reported a second study to
by cyanobacteria during the growing season and 50– quantify N2 fixation in 11 commercial crops of sugar
90 kg N/ha by the cyanobacteria–Azolla symbiosis. cane in Brazil, also using 15N natural abundance.
Giller (2001) was more conservative, referring to Their data provide a stronger case for consistent and
average rates by free-living cyanobacteria of 12 kg N/ substantial N2 fixation. They reported an overall
ha/cropping season in a study of 190 rice fields in the average δ15N value for the cane of +6.38‰ (range
Philippines and 27 kg N/ha/cropping season in a +3.3‰ to +13.2‰), compared with +9.10‰ (range
review of published estimates. Giller (2001), howev- +5.4‰ to +26.5‰) for the reference samples. An
er, cautioned that the vast majority of the estimates aggregated estimate of Ndfa, using those average
were based on acetylene reduction assays and likely values, is 30%. The authors concluded that N2
to be inaccurate. fixation appeared to supply between zero and 60%
Apparent N2 fixation rates of the cyanobacteria– of the N in the sugar cane crops in the study. They
Azolla symbiosis are impressive, e.g. daily accumu- also acknowledged that the complex interactions
lation rates of Azolla N of 0.4–3.6 kg N/ha with a between plant genotype, the suite of N2-fixing (and
mean of 2 kg N/ha and total growing season other) bacteria associated with the plant and the
accumulation of 25–170 kg N/ha (mean of 40 kg N/ environmental and edaphic conditions need to be
ha) (Giller 2001). It is probable that N2 fixation defined before agronomically-significant inputs of
contributes at least 80% of the Azolla N. fixed N can be guaranteed.
It would be reasonable to assume that most of Given the large variations in apparent N2 fixation
the world’s rice paddies contain free-living cya- of sugar cane in the field studies in Brazil (Yoneyama
nobacteria, but that the cyanobacteria–Azolla sym- et al. 1997; Boddey et al. 2001, 2003), Japan and the
biosis is present in only about 2% (3 Mha) of the Philippines (Yoneyama et al. 1997), Australia (Biggs
paddies (Giller 2001). Thus, the average estimates et al. 2002) and South Africa (Hoefsloot et al. 2005),
of N2 fixation in rice paddies of about 30 kg N/ha/ it is impossible to estimate global N2 fixation with
year and a total of 5 Tg N/year appear reasonable confidence. The proposition of Smil (1999) that the
(Table 6). world’s 20 Mha of sugar cane fix, on average, 100 kg
N/ha/year is not supported by the literature. It is also
unlikely that Brazil’s 7 Mha of sugar cane sustain N2
Endophytic, associative and free-living bacteria fixation at such high rates—a more realistic value for
in sugar cane systems Brazil would be 40 kg N/ha, calculated using average
Ndfa of 20% and total crop N of 200 kg/ha.
Smil (1999) reported that the world’s 20 Mha of sugar Reasonable, but speculative, values for the remaining
cane fix, on average, 100 kg N/ha, based principally 14 Mha might be an average of 20 kg N/ha fixed,
on research in Brazil (e.g. Boddey et al. 1995). The assuming Ndfa of 10% (Table 6).
evidence for substantial inputs of fixed N in Brazilian
sugar cane grown in large pots is strong (Lima et al.
1987; Urquiaga et al. 1992) and is supported by the Endophytic, associative and free-living bacteria
isolation of a large and diverse number of N2-fixing in crop lands not used for legumes and rice
bacteria from inside and outside of the cane roots (see
Boddey et al. 2003). Data on N2 fixation of field- Smil (1999) suggested the plant-associated and free-
grown plants using 15N natural abundance, however, living bacteria in the 800 Mha of cropping lands used
is more equivocal (Yoneyama et al. 1997; Biggs et al. primarily for the cultivation of cereals and oilseeds
2002, Hoefsloot et al. 2005). In the Yoneyama et al. fixed N at an average, annual rate of 5 kg/ha and aPlant Soil (2008) 311:1–18 13 global, annual rate of 4 Tg N (Table 6). These values fixed annually are questionable and are likely to be far are very speculative but, with current knowledge, it is too high (Table 6). impossible to offer alternatives. A number of reviews The savannas do produce substantial quantities of of plant-associated N2 fixation have clearly highlighted C-rich plant residues that are a potential energy source the many methodological problems and inconsistencies for N2-fixing bacteria. As well, a large proportion of in the published studies (Boddey 1987; Chalk 1991; the savannas are now used for grazing and, in Giller 2001; Kennedy and Islam 2001; Giller and countries like Brazil, Venezuela and Colombia, have Merckx 2003). One of the key problems is distinguish- been oversown with improved species of grasses, ing between inputs of N by free-living and associative such as Brachiaria spp., Panicum maximum, and agents and other external sources of N contributing to Andropogon gayanus. There may be about 200 Mha agricultural soils, e.g. N in rainfall and dry deposition. tropical savannas that contain improved grass species Such inputs can represent 3–50 kg N/ha/year (Gould- (RM Boddey, personal communication). Reis et al. ing et al. 1998; Giller and Merckx 2003; McNeill and (2001), using natural 15N abundance, reported Ndfa Unkovich 2007). values of 25–40% for genotypes of P. purpureum and Roper and Ladha (1995) concluded that the free- 2–26% for five species of Brachiaria, and N2 fixation living, heterotrophic bacteria may fix significant values >100 kg N/ha. Although these data suggest a amounts of N in agricultural systems, using crop large potential for N2 fixation by bacteria associated residues as an energy source. However, they did not with some of the tropical grasses, there are still speculate as to what the average rate of N2 fixation questions as to whether the apparent 15N isotope might be. More recently, Gupta et al. (2006) dilution is due to N2 fixation, or to other effects, or to suggested N2 fixation rates of 1–25 kg N/ha/year for a combination of both. Thus, the occurrence and dryland cereal systems in southern Australia. Other intensity of N2 fixation in this system by the reviews present similar ranges, or suggest a maximum cyanobacteria, endophytic and associative bacteria value that is unlikely to be exceeded. For example, and heterotrophic free-living bacteria are essentially Giller (2001) concluded that N2 fixation by free-living unknown. A notional rate of
14 Plant Soil (2008) 311:1–18 production. It is also possible that particular systems together these systems fix
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