1.10.19
ANALYSIS OF THE EXTRACELLULAR MATRIX SURROUNDING GERM TUBES AND APPRESSORIA OF THE ANTHRACNOSE FUNGUS COLLETOTRICHUM LINDEMUTHIANUM

S RAWLINGS1, K HUTCHISON1, JR GREEN1 and RJ O'CONNELL2

1School of Biological Sciences, University of Birmingham, Birmingham B15 2TT, UK; 2 IACR-Long Ashton Research Station, Department of Agricultural Sciences, University of Bristol, Long Ashton, Bristol BS 18 9AF, UK

Background and objectives
The germ tubes and appressoria formed by the bean anthracnose fungus Colletotrichum lindemuthianum are coated with an extracellular matrix (ECM) which probably mediates attachment to the plant surface. We have produced two monoclonal antibodies (MAbs) that bind to this ECM [1, 2]. One of these MAbs, UB26, binds primarily to the sheath around germ tubes, whereas the other, UB31, labels the appressoria strongly. Both MAbs recognise high-molecular-weight glycoproteins which adhere to the substratum on which fungal cells are grown. The objectives of the work are: (i) to clone and sequence the genes encoding these glycoproteins; (ii) to study the interaction of these glycoproteins with other components of the ECM; and (iii) to determine their role in fungal adhesion.

Results and conclusions
Using immunofluorescence of infection structures grown on glass slides, we have shown that UB26 recognises a small number of Colletotrichum species (C. lindemuthianum, C. malvarum, C. orbiculare and C. trifolii), whereas UB31 labels all Colletotrichum species tested. In Western blots, UB31 labels glycoproteins extracted from a melanin-deficient mutant of C. lagenarium but shows no labelling of extracts from the wild type. This suggests that the glycoproteins recognised by UB31 are linked to melanin at the surface of appressoria. Both antibodies label C. lindemuthianum hyphae grown in liquid culture, and a cDNA library has been prepared from these hyphae for immunoscreening with the MAb UB26.

References
1. O'Connell RJ, Pain NA, Hutchison KA et al., 1996. Journal of Microscopy 181, 204-212.
2. Pain NA, Green JR, Jones GL, O'Connell RJ, 1996. Protoplasma 190, 119-130.