The legume- rhizobia relationship in the Mediterranean basin Howieson J.G., O' Hara G.W., Loi A. in

Zaragoza : CIHEAM

Cahiers Options Méditerranéennes; n. 45 2000

pages 305-314

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--- Howieson J.G., O'Hara G.W., Loi A. Th e legu me-rh izobia relation sh ip in th e Mediterran ean basin . In : Sulas L. (ed.). Legumes for Mediterranean forage crops, pastures and alternative uses . Zaragoza : CIHEAM, 2000. p. 305-314 (Cahiers Options Méditerranéennes; n. 45)

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The legume -

Basin

J.G. G.W. A.

Studies, of Science &

6150,

WA, South 6151, and

The of Nedlands, 6907,

Summary

-

the is not always optimal. options to N-fixation depending on

the of the limitation to legume words: legume, N-fixation,

- renforcer la valeur de la fixation synlbiotique de l'azote, il est nécessaire de comprendre le produit G2 x 02 G correspond génotyyes de la légumineuse et de son associé, et l'environnement édaphique dans lequel la symbiose doit avoir Dans le bassin méditerranéen, est évident que malgré une Co-évolution des légumineuses et de leur rhizobium, l 'association n'est pas toujours optimale. alternatives de recherche sontproposées afin d'anzéliorer la fixation de l'azote en fonction du type de contrainte lirnitant lespe$ornlances de la légumineuse ou du rhizobium.

légumineuse, fixation d'azote,

The adaptation of plants to climate is often discussed in of G ^X E -the

Genotype X When dealing with the legume - the

becomes one of the second i.e. G' ^X E, G to the genotypes of both the legume and its To effectively the potential of this symbiosis, we must fully these multiple

The capacity to fix N2 in symbiosis with plants is found in of

the (Allovhizobiz.m,

and Sinovhizobizlm,), actinomycetes (Frankia) and (Anabaena,

(Young, 1996). legumes the of the N%-fixed in

in a that is not leached.

is also supplied in equivalent amounts, ton of N

consumes 1.3 ton of oil equivalents and Cocking, 1997). Thus, the inputs of N into systems by N-fixation in nodulated legumes fundamental to sustainable and economic

At least some of the legumes of the basin have been studied

in to (e.g. and Cocks, 1990; et al. 1993).

the most astonishing aspect of the of the basin is how little we

know about ecology! Given to in that as well as

in that a climate, it is that so few

studies of the ecology of in the basin have been Of the many

studies of legume ecology that have been basin, few have focussed upon the symbiosis, whilst even have investigated symbiotic with the The publication by Loi et al., (1999) is a notable exception. The aim of this

is to outline to and manipulating the

in

N optimally?

has been that genotypes adapted to soil

conditions and may be advantaged both and competitively conditions that suit

them, of the symbiotic with host 1999). we follow this

statement to its logical conclusion it that symbiotic effectiveness a selection upon a population than does adaptation to soil and climate.

This is intuitively logical and was, in a sense, implied in a study by and

(1987) that soil a in nodule

occupancy. all -what advantage does a being fully effective at N-fixation than '75% effective? As long as the host has sufficient N to and to compete with its the photosynthate supply to the nodules is and

the well We cannot assume that in any given

the legume symbiosis is optimised simply because both and

have evolved in that evidence this

1 shows that was no yield advantage in Biserrula pelecinus L.

inoculation with a collected as its host. The genotype of B.

yelecim~s yielded well with and vice vemz. fact, the data

indicate that the effective at fixing N per se than those

This possibility needs to be investigated.

This to be the d e than the exception. et al., (1999) that monospecific genus, Hyrnenocarpus circinnatus L. (Savi) the

of high effectiveness gave no indication of specificity. Eight

isolated H. circimatus collected in the Cyclades of nodulated nine

host ecotypes only one actually fixed

Sulas et al., (1998) the yield of coronarium L. on a soil in inoculation with selected high N-fixation, indicating the soil in that a effective suite of

examples populations of N-fixation in the

basin et al. 1995; et al. 1995; and 1991) and

et al. 1999).

1 4 0

¹ T

¿# 60

9

F

20 O

Y

- -

Figure l. Yield of two genotypes of Biserrula pelecinus (95GCN68 and

when inoculated with collected as

is yield of tops as a of the inoculant in

(Loi, Yates and unpublished data).

is scope N-fixation in legumes within the

basin, as is legumes have been

then can this optimisation be best achieved? is in G2 X E that will

be made and we have developed a set of this we must

the status quo with to the symbiosis as it applies to legume of

2 outlines common leading to options that when

investigating legume nodulation and N-fixation:

1

a high population of OOO/g soil)

Select a host legume genotype to nodulate effectively with soil

I

genotype that has no

Figure 2. A schematic of to N-fixation in legumes

selection of the host genotype.

1 the soil contains a high population of effective that cause 2 the soil contains a low population of effective that 3 the soil does not contain capable of nodulating with the host W-fixation

nodulation and N-fixation legume of and hence inoculation is

each a of options legume nodulation, with

the most likely option to succeed denoted by line density in 2. Some examples the options A-F have been successful in given in Table 1.

Of exist. example, is. a high population of effective

and no to inoculation (the case with many pulse legumes in Asia)

the legume of is with N and inoculation is

ineffectual (e.g. vulgaris in inoculation is

and having decided upon the a of techniques available to select adapted

Table 1. examples applying the pathways A-F in Fig. 2 have been successful in developing symbiotic N-fixation in

A

Bi, Bii

C

F

Trifolium nzichelianum Lotus ornithopodiodes, Biserrula pelecinus

Ornithopus spp Trifolium spp

Vicia, Lens polymorpha

Cicer, Hymenocarpus, Scorpiurus, Biserrula, Hedvsarum. etc.

2000, this issue

Loi unpubl., et al.

1995

(unpubl.) et al. 1999 Watlun 1999

et al. 1999 and Ewing 1986

and Ewing 1989

CV. well adapted to the medic population in alkaline soils

legumes which avoid with a

effective medic

population on soils 6-9.

Good nodule occupancy in the of sowing on acid soils.

A host

selected new

and to

WU95.

Selection of adapted inoculants acid soils An acid inoculant was selected to colonise soils

combination with E, a symbiotically competent legume was selected the soils of 4-5.

The with

many legumes in

4.5-6

Techniques for selecting and evaluating rhizobia to match strains with both legume hosts and soil conditions

The genetic of populations can now be investigated with

methodologies (Vinuesa et al. 1998). This, time, ecologists to follow population dynamics, population and investigate genetic

doing so, ecologists can that

inoculant quality, of

1. high NZ-fixation with the intended host species without

2. adaptation to the edaphic host;

The needed in inoculant-quality species;

3. genetic stability in and soil;

4. and in inoculant and

5. competitiveness with indigenous soil

l . Selecting effective rhìzobìal strains with broad host-range characteristics To genetic compatibility NZ-fixation between host and

uses a lit, glasshouse than

pouches. We emphasise fundamental aspects:

0 the must be limiting only in plant available N

0 we expect within species NZ-fixation

0 we acknowledge the necessity to select that will not

existing legumes in the

We symbioses nodulation and NZ-fixation in sand et al., 1995)

than because it has to optimise

conditions each “new” legume studied. The method consists of steamed, sand held pots, with a system in the base and alltathene beads on the

The beads minimise losses and contamination

Autoclaved plant solution added as a capped tube. This

system can be utilised legumes of all seed sizes. is with seeded legumes

(>5 mg) to select size and of attention

must be paid to hygiene in the glasshouse to avoid contamination by Effectiveness of NZ-fixation is by yields and %N of inoculated plants with

and uninoculated

2. Screening edaphic adaptation

many symbioses, the challenge is to develop a consistent nodulation

the legume in the selection focusses on

effective and then between them on the basis of

ability to in, and to colonise, soils. The methodology has been

the technique as by and Ewing (1986).

to the soil as inocula at a site of chemical and physical

and of the species of of the site should

be in the host-legume, if this is likely to be a upon

Soils with a sandy (5-10% clay) expedite of examination of the nodules and also place upon inoculant

the bio-assay the plots sown as 2m lines of inoculated legume seed

by l m and with all and except N.

allowed to a full season which top weight and N-fixation can be assessed. the soils low in available N, the biomass of the tops is an

excellent of symbiotic This can valuable given that

in stage 1 was based upon N-fixation conditions. the soil contains N, then the abundance method (Unkovich and 1998) can

indicate symbiotic is possible that in

abilities to on the seed in the legume in difficult soils, hence data on situ essential the selection of elite

Following a (usually in the soil, the individual

and movement away of to the soil using a nodulation bio-

assay this assay, uninoculated, seeds sown the line at two

points.

l 2

Figure 3. A schematic of the technique. lines seed sown inoculated 1) uninoculated 2).

plants excavated 10-12 weeks sowing and the nodulation

design can be as blocks, adjusted to take advantage of spatial analysis techniques (Cullis and Gleeson 1991).

3. Genetic stability in culture, storage and soil

sometimes on media that can genetic

changes that affect symbiotic Whilst it is that must

use such media to to such media should be minimised

because the of change is example, in leguminosarunz bv.

viceae non-nodulating in 30% of colonies only 10

comm.). is to at -8OCO in as vacuum-

(Vincent 1970).

Genetic stability in soils is a much difficult to evaluate. lotì

to have a to genetic of a “symbiosis island”

to (Sullivan and 1998). this is amongst

species is a the of substantial genetic in

populations of Sinorhizobium nzeliloti, 1. trijiolii and Bradyrlzizobium sp (Lupinus) anecdotal evidence that genetic exchange these species. Given

of this phenomenon, it may soon be possible to identify the genetic

disposing to gene and to to this

4. Satisfactoly growth and sumival in irlocularzt rnanllfncturing procedures

is evidence that the can affect the adaptation of

inoculants to the soil example, in the if the

is exposed to a it may initiate a of physiological and

biochemical that it to handle an acid soil This is

the adaptive acid in by

and Glenn (1994). Not all possess the capacity to the acid trifolii does, the acid sensitive TA1 does not (Watlin 1999).

5. Cornpetitiveness with indigenous soil rhizobia

Of to the basin is how we can N-fixation

the of competitive of A to study competition is to

label with a such as GUS (Wilson, 1995) and visualise the outcome of competition have been selected on the basis of adaptation to the is a chance they can be competitive with indigenous and less effective genotypes.

the does not have access to technology,

a means to between they

to use a of this (Vinuesa et al.

1998).

Conclusions

is evidence that legumes do not always fix N optimally in the

We should not assume that because of co-evolution, the legume and its

matched to fix N. is not, selection

on optimal N-fixation. view of this, we have discussed

to symbiotic N-fixation developing an of G2 ^X E

and then applying some elite selection.

AS., (1987). of lime and phosphate on nodulation of soil-

Trifolium subterrarzmeum L. by indigenous 53,

Cullis, Gleeson, A.C. (1991). Spatial analysis of field and extension to two dimensions. Biometrics 47, 1449-1460.

J.C. (1999). abundance and symbiotic effectiveness of legunzinosarum bv. trifoZii alkaline

soils in South Aust. (in

T., Cocks, (1990). of annual legumes in

J.G., Ewing, (1986). Acid in the -

2090-209’7.

J. Appl. 27,578-591.

symbiosis. Aust. J. 37, 55-64.

J.G., Ewing, (1989). Annual species of in ability to nodulate on acid soils. Aust. J. 40, 843-50.

J.G., Loi, A., S.J. (1995). Biserrzda pelecinus L. -a legume species with potential acid, duplex soils which is nodulated by unique Aust. J.

Agric. 46, 997-1007.

J.G. (1999). The S.J. Cocks (Eds.)

of and Forage Legumes, Academic

pp. 96-106.

J.G., G.W. and C m , S.J. (1999). Changing legumes in

an Field

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L.A., N., and Altuntas, S. (1995).

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