Mode of reproduction of Brazilian species of Adesmia (Leguminosae)

Mode of reproduction was studied in 15 species of Adesmia DC. (Leguminosae). In six species, three treatments were used: mutual pollination, mechanical stimulation and control. Fifty-four plants of these six species were grown in a greenhouse, indi vidually isolated in nylon screen boxes. Flowers were labelled and submitted to the different treatments. In addition, the frequency of spontane us self-pollination in the absence of pollinators was studied in 200 plants of nine other species. These 200 plants were kept in a greenhouse, which avoided contact with any possible pollinator. Adesmia bicolor, A. muricata, A. punctata and A. riograndensis produced seed both by crossand self-pollination. Adesmia punctata nd A. riograndensis need mechanical stimulation for self-pollination. Adesmia incana reproduced by self-pollination; however, the possibility of cross-pollination cannot be totally ruled out. A esmia tristis reproduced mainly by cross-pollination and a mechanism of self-incompatibility is suggested. Among the nine species that were not ex pos d t pollinators, A. securigerifolia produced a large amount of seed, indicating that it is a self-pollinating species. Adesmia arillata, A. araujoi, A. ciliata, A. psoraleoides , A. rocinhensis, A. reitziana, A. sulina and A. vallsii did not produce any seed under the experimental conditions, suggesting that they are cross-pollinated or that they need mechanical stimulation to reproduce. Departamento de Plantas Forrageiras e Agrometeorologia, Faculdade de Agronomia, Universidade Federal do Rio Grande do Sul, Av. Bento Gonçalves, 7712, Caixa Postal 776, 91501-970 Porto Alegre, RS, Brasil. Send correspondence to S.B.T. Rua Conde de Porto Alegre, 1368, ap. 301, 97015-110 Santa Maria, RS, Brasil. E-mail: tedesco@infoway.com.br Embrapa Recursos Genéticos e Biotecnologia, 70770-900 Brasília, DF, Brasil.


INTRODUCTION
The South American genus Adesmia DC. belongs to tribe Adesmieae (Benth.)Hutch.and comprises about 200 species.Seventeen species and one variety, annual or perennial, herbaceous or shrubby, are found in Brazil.All of them are endemic to the southern region, which includes the States of Rio Grande do Sul, Santa Catarina and Paraná (Miotto and Leitão-Filho, 1993).
Many of the Brazilian species are promising as forage crops and have attracted the interest of researchers.Several species of Adesmia from southern Brazil present good winter growth, when there is normally lack of cattle forage.They are well adapted to the regional environment, are widespread and have a high nutritional value.Crude protein percentages range from 6.9 to 17.5 in A. tristis, 18.6 in A. latifolia, 17.9 to 21.5 in A. ciliata, 19.7 in A. psoraleoides and 23.4 in A. punctata (Dall´Agnol and Gomes, 1994).High crude protein values, as well as good in vitro organic matter digestibility (IVOMD), were also found for A. latifolia, A. tristis and A. punctata, by Scheffer-Basso (1999).
Germplasm accessions of the Brazilian species of Adesmia have been stored at Embrapa Recursos Genéticos e Biotecnologia/CENARGEN, Brasília, DF, since 1984, when intensive efforts to collect and conserve this germplasm were initiated.Germplasm characterization, including knowledge about the mode of reproduction, is essential for any further research.The reproductive success of a given species depends on the reproductive strategy used to ensure perpetuation.
The determination of the mode of reproduction should be one of the first steps in germplasm characterization, since the process of collecting, multiplication, and further plant breeding procedure depends on how each species reproduces (Valls, 1988).Depending on the mode of reproduction, different plant breeding methods should be used (Poehlman, 1965;Borém, 1997).Native legumes are important components of many natural pastures in several countries and regions, such as in southern Brazil; nevertheless the reproductive processes are unknown for many species (Izaguirre et al., 1994).Very little is known about the mode of reproduction of Adesmia (Valls, 1984).Adesmia latifolia is considered a versatile species, allowing for self and cross fertilization (Tedesco et al., 1998), but there is no published information about the other Brazilian species.Visual observation has shown that the flowers are visited and pollinated by insects.
We determined the mode of reproduction of 15 Brazilian species of Adesmia.This is expected to be useful for plant breeding.vidually isolated in nylon screen boxes of 0.5 x 0.5 or 0.5 x 1.0 m, depending on plant height.The effects of three treatments were tested: T1: pairs of plants, which were mutually pollinated by reciprocally transferring pollen from one to the other.Flowers were labelled and manually pollinated with a small cuneiform piece of cardboard, suitable to pollinate without damaging the flowers; T2: flowers were labelled and mechanically stimulated by manually pressing the keel with a piece of cardboard as in T1, one per flower, thus liberating the anthers and releasing pollen.This artificial tripping simulates the action of the pollinating insect; T3: flowers were labelled and left undisturbed.The number of flowers in each treatment varied among species and among treatments, depending on the availability of exposed flowers.T1 and T2 were carried out from 10:00 a.m. to 1:00 p.m. for several days.The number of fruits produced was counted for each plant and treatment, and compared by a χ 2 test.

RESULTS AND DISCUSSION
Manual stimulation was the most efficient treatment for A. riograndensis (Table III), suggesting that this species is autogamous, but needs mechanical stimulation.Per-centages of cross-pollination are only estimates, since there was no emasculation, but just mutual pollen transfer, so that some of the fruits may have been formed by self-pollination.As A. riograndensis is annual, a higher percentage of autogamy would be expected.A. bicolor needed mechanical stimulation to set fruit, suggesting that it is a preferentially allogamous species (Table III), even considering that some of these fruits could have been formed by self-pollination.Similar results were found for A. punctata and A. tristis.
A. muricata is probably mainly cross-pollinating (Table III).In A. incana, no fruits were formed by T1 and T3 but 6.77% of fruits were formed by T2 (mechanical stimulation).The number of flowers treated was small (Table III), but the hypothesis that this species is an inbreeder that needs mechanical stimulation cannot be ruled out.
Apparently A. tristis, A. bicolor, A. incana and A. punctata need mechanical stimulation for self-pollination, which in nature is done by pollinating insects.There may be a self-incompatibility mechanism in A. tristis, as the percentage of fruits formed was small (Table III).
Self-incompatibility may hinder genetic breeding, avoiding selfing, but at the same time assures maintenance of high heterozygosis.It is genetically controlled (Nettancourt, 1997) and common in several economically important legumes of the genera Trifolium, Lotus, Melilotus and Values followed by the same letters do not differ by the χ 2 test (P > 0.05).T1-Mutual pollination; T2-mechanical stimulation; T3-control.
Observations with the other nine species suggest that A. securigerifolia is an inbreeder, since it produced a large amount of seed by self-pollination (Table IV).The other species did not produce any seed at all in the absence of pollinators, suggesting that they are obligate outbreeders or need mechanical stimulation for selfing.
Data on mode of reproduction should be taken into account when sampling new populations, to assure a representative portion of the existing genetic variability, as well as when plant breeding experiments are designed.Studies with molecular markers may help to elucidate the mode of reproduction of these Adesmia species; they are being planned by our research group.

Table I -
List of Adesmia accessions utilized for the determination of the mode of reproduction (Experiment 1).

Table II -
List of Adesmia accessions utilized for the observation of the mode of reproduction (Experiment 2 ).

Table III -
Percentage of fruits formed under treatments T1, T2 and T3 to determine the mode of reproduction of Brazilian species of Adesmia.