Towards incorporating the biosynthetic transformation required for Striga inhibition from Desmodium into edible legume intercrops

In sub-Saharan savannah agriculture, a primary activity is the cultivation of maize as a subsistence crop. The farmers are resource poor and have a range of problems with which to contend, in particular the loss of yield due to damage by insect pests (stem-borers) and parasitic weeds (striga). The insect stem-borers are the caterpillars of indigenous and introduced moths, and attack maize stems causing extensive damage and even the loss of the plant by collapse. During the course of work on controlling insect pests, by intercropping with repellent plants and trap cropping with plants that attract the adult moths away from the maize, an intercrop was found that provided dramatic control of striga. These parasitic weeds from the Striga genus are in the snapdragon family and are a major threat to the staple food crops of over 100 million people in the region. The striga-controlling intercrops were cattle forage legumes, commonly called desmodium, and when grown on-farm as a one-to-one intercrop between maize and other subsistence cereals such as sorghum, the parasitic striga was controlled to the extent that a yield of well under 1 tonne/hectare of maize could be raised to over 5 tonnes/hectare. To show this protection was based on chemicals generated from the root system of desmodium, it was demonstrated that the effect could be provided by passing water over the roots of desmodium and then into soil containing the striga and maize seeds. Furthermore, when desmodium was grown in water with nutrients but without soil (hydroponically), the water captured the activity, which could then be transferred to the striga and maize seeds in soil, and again conferred control of the parasite. The chemical composition from the water effecting this control has been studied and fractions of the content separated from each other. After the striga seed germinates, a radicle (root) is produced that will develop into the attachment organ for parasitism of maize. However, one particular fraction causes interference with the development of the radicle and from this we have identified a potent radicle inhibitor. The structure of this compound is a di-C-glycosylflavone called isoschaftoside. Many plants possess the ability to produce flavones, however the glycosylation steps required to produce isoschaftoside are unusual. We propose to identify the enzyme in desmodium that performs this glycosylation by searching for C-glycosyltransferase activity on substrates in the biosynthetic pathway in protein extracts of root tissue. We will purify the protein by following this activity using standard protein purification techniques and characterise its structure by determining its amino acid sequence. This sequence will allow us to search the genomes of edible crop legumes in an effort to find this protein and so breed edible crop legumes that, when intercropped, also prevent striga from parastising maize. In addition, the desmodium gene itself, which generates the C-glycosyltransferase protein in the plant, provides a target for genetic modification of an edible crop legume that will prevent striga parasitizing maize. This work will provide the first elucidation of how a valuable weed suppression mechanism can be utilised and could provide the basis for attempts to control weeds in agriculture in other developing countries and potentially also in developed countries, such as the UK.

Desmodium uncinatum, when intercropped with maize and other cereals in soil with a high level of seeds of the parasite Striga hermonthica (Scrophulariaceae), gives yield increases of 5-fold and when repeatedly used, helps to reduce the Striga seed bank. Activity has been demonstrated to be in both root exudate and extracts of D. uncinatum and acts without the need for soil or rhizobial inoculation. Such suppressive activity is not present in the root exudates of other commonly used intercrops such as cowpea, beans, soybean. Bioassay-guided fractionation of D. uncinatum root exudate, from hydroponic culture, or extract has identified, by NMR spectroscopy, lipophilic components conferring high germination of the Striga seed and hydrophilic components that cause inhibition of the rate of elongation of the Striga radicle. The main inhibitory compound identified is an unusual di-C-linked glycoside of a flavonoid. D. uncinatum is a legume that can be used as cattle forage but edible crop legumes, such as cowpea, do not show this protection mechanism despite possessing the biosynthetic pathway to flavones. The ability to biosynthesise the active component requires this glycosylation process. We propose to isolate the C-glycosyltransferase (CGT) responsible for biosynthesising the active component from D. uncinatum and using amino acid sequence data, search for genes in sequenced legumes, in particular cowpea, that confer the same biosynthetic transformation. This provides a basis for breeding the protection pathway into an edible crop legume. In addition, the C-glycosyltransferase gene will be a target for genetic modification of cowpea to produce an edible legume intercrop that possesses the Striga protection mechanism.