3.4.46
DEVELOPMENT OF SEROLOGICAL METHODS FOR THE DETECTION OF MACROPHOMINA PHASEOLINA, INCITANT OF ASHY STEM IN COWPEA (VIGNA UNGUICULATA)

L AFOUDA1, K WYDRA2and G WOLF1

1Institute of Plant Pathology and Plant Protection, University Gottingen, Germany; 2International Institute of Tropical Agriculture, PHMD, Cotonou, Benin

Background and objectives
Macrophomina phaseolina is a worldwide distributed seed- and soilborne pathogen with a wide host range. Infected plants show various symptoms such as damping-off of young seedlings, stem and root fissures, ashy stem, leaf blight and wilt, and sudden death of mature plants. Apparently healthy plants can carry the fungal infection in a latent form [1]. Yield loss can be high, particularly in regions with high temperatures and pronounced water deficit. The production of clean seed is a prerequisite for a healthy plant stand and can reduce damping off in early stages of plant growth as well as during the latent development of the mycelium in growing plants. To develop integrated methods to control the disease, an accurate knowledge of the biology of the fungus in infected plants and seeds is necessary. Therefore, a sensitive method was developed to detect the fungus even during latent stages of infection.

Materials and methods
Antigens were prepared from mycelium and culture fluids of Macrophomina phaseolina and injected into rabbits. Four polyclonal antibody fractions were purified and tested in a direct DAS-ELISA for their ability and sensitivity to detect Macrophomina phaseolina in infected plants. Fractions with high sensitivity in the DAS-ELISA were selected to study the biology of Macrophomina phaseolina in plants.

Results and conclusions
Three from the four tested antibodies allowed the detection and quantification of Macrophomina phaseolina in different parts of plants with symptoms as well as in symptomless plants. In infected plants, Macrophomina phaseolina was found to be mainly located in the roots and in the hypocotyl and to a small extent in the epicotyl. There was no evidence of fungal presence in the leaves. The latent occurrence of Macrophomina phaseolina was confirmed by detecting the fungus in inoculated but healthy looking plants.

References
1. Toudou A, 1986. Thesis de Docteur-Ingenieur en Sciences Agronomiques, Universite de Rennes.