2.2.122
MOLECULAR CHARACTERISATION OF THE BLAST PATHOGEN MAGNAPORTHE GRISEA FROM SOME WEST AFRICAN RICE SCREENING SITES
MOLECULAR CHARACTERISATION OF THE BLAST PATHOGEN MAGNAPORTHE GRISEA FROM SOME WEST AFRICAN RICE SCREENING SITES
S SREENIVASAPRASAD and J CHIPILI
Horticulture Research International, Wellesbourne, Warwickshire CV35 9EF, UK
Background and objectives Blast caused by Magnaporthe grisea is the most widespread and damaging disease of rice. Despite the tremendous work on disease management, epidemics of blast often devastate the crop across Africa, Asia and Latin America. Absence of durable resistance in the field is a major problem, which has been attributed to the high levels of virulence polymorphism in pathogen populations. Use of molecular techniques, particularly the M. grisea repeat elements, has enabled an understanding of the genetic diversity of the pathogen populations in a geographic location [1], [2]. We are in the process of characterising the M. grisea populations from various rice screening sites in west Africa, as part of a collaborative project with West Africa Rice Development Association (WARDA), using different molecular markers, and by pathotyping (with the International Mycological Institute, UK). The objectives are to assess the genotype and pathotype diversity of M. grisea populations, analyse the relationship between genotype lineages and blast pathotypes and identify sites suitable for resistance screening.
Materials and methods
Field isolates of M. grisea were obtained from surface sterilised individual lesions. Monoconidial isolates were obtained by picking single germinating spores on 4% water agar. Preparation of mycelial mats, DNA extraction and Southern hybridisation with pCB586 [1] were based on standard procedures followed for M. grisea. MGR586 fingerprints were visually inspected and similarity values generated based on the presence or absence of a particular band from pairwise comparisons. Isolates showing 80% or more similarity were assigned to a single lineage. RAPD analysis and automated sequencing of the ITS region were undertaken following standard protocols.
Results and conclusions
More than 200 rice blast samples were collected from various screening sites in west Africa, following preliminary surveys co-ordinated by WARDA and associated National Agriculture Research Service scientists. Up to 50 isolates have been isolated so far and are being characterised using different molecular markers. MGR586 fingerprint profiles were generated for ten monoconidial isolates each, obtained from a limited number of field isolates (IDR1, ONR1, FKR8 and IKR1012). No variation in fingerprint pattern was observed among the monoconidial isolates belonging to each of the field isolates. Albeit based on a limited number, it appears that field isolates of M. grisea are genetically homogenous and stable. A similarity matrix generated from pairwise comparison of MGR586 fingerprint profiles from more than 30 isolates divided them into at least 4 distinct lineages, with the overall similarity ranging from 13 - 100%. Two isolates (IKR1 012 and NB12), obtained from rice blast samples from Nigeria, showed atypical patterns with only 7 - 9 MGR586-hybridising bands, but their epidemiological significance is unknown. RAPD-PCR analysis of the atypical rice isolates along with four each of typical rice and non-rice isolates did not reveal any close relationship between them. ITS sequence of the atypical isolates was determined and comparison with ITS sequences of M. grisea from a number of different hosts, obtained from MH Lebrun, University of Paris, France, revealed a close relationship between the two atypical isolates from this study to M. grisea isolates from Pennisetum, Ginger and Digitaria. An objective assessment of the genetic diversity of west African M. grisea isolates examined so far and the distribution pattern of various lineages among the screening sites surveyed will be presented.
References
1. Levy M, Romac J, Marchetti MA, Hamer JE, 1991. The Plant Cell 3, 95-02.
2. Chen D, Zeigier RS, Leung H, Nelson R, 1995. Phytopathology 85, 1011-20.