Monoclonal antibodies to isolate-, species-, and genus-specific components on the surface of zoospores and cysts of the fungus Phytophthora cinnamomi

Abstract

Monoclonal antibodies have been raised to components on the surface of glutaraldehyde-fixed zoospores and cysts of an isolate of the pathogenic fungus Phytophthora cinnamomi. Hybridoma supernatants were screened using an immunofluorescence assay, and of 35 cell lines producing antibodies that reacted with the P. cinnamomi cells, 10 have been selected and their specificities examined in detail. The monoclonal antibodies were found to possess a valuable spectrum of taxonomic specificities, and have revealed, for the first time, the presence of isolate-specific antigens on the surface of P. cinnamomi cells. The monoclonal antibodies were tested against six isolates of P. cinnamomi, six species of Phytophthora, and three species of Pythium. In addition to the isolate-specific monoclonal antibodies, species-specific and genus-specific markers which are unambiguous in tests conducted so far have been obtained. The monoclonal antibodies have also revealed the presence of spatially restricted antigens on the surface of the zoospores. Some of these segregated antigens are species-specific and others are more general, occurring in all Phytophthora and Pythium species examined. All of the monoclonal antibodies promise to be of great assistance in investigations of the biology and taxonomy of P. cinnamomi. The methods described should be readily applicable to studies of other fungal pathogens.

New Approaches to the Detection of Microbial Plant Pathogens

“…campestris and obtained monoclonal antibody lines that were specific at the genus, pathovar and strain levels. Similarly, Hardham, Suzaki and Perkin (1986) used zoospores and cysts of Phytophthora cinnarnomi as immunogens and obtained monoclonal antibody lines that were isolate, species or genus specific. Interestingly, this work showed no obvious relationship between the epitopes recognized by the antibody and the specificity of the antibody (Figure 1).…”

Evaluation of a polyclonal antibody immunoassay for detection and quantification of Macrophomina phaseolina

“…Antibodies against fungal pathogens have been raised from soluble proteins, crude cell components, fungal homogenate, culture fluids, and ribosomal proteins (Savage & Sall, 1981;Hardham et al, 1986; Ribosomal proteins were used to raise the antisera. Gleason et al, 1987;Reddick & Collins, 1988;Lange et al, 1989).…”

“…Polyclonal antisera were raised from whole cells, wall components, soluble proteins, and ribosomes of different fungal species (Savage & Sall, 1981;Johnson et al, 1982;Hardham et al, 1986;Gleason et al, 1987;Takenaka, 1992). Though there are reports that polyclonal antisera to fungi generally lack specificity to ELISA (enzyme-linked immunosorbent assay) and cross-react with both related and unrelated fungal species (Dewey et al, 1991), some workers demonstrated that species-specific polyclonal antibodies are present in antisera raised against fungi and can be useful for detection and quantification purposes (Adams & Butler, 1979;Jamaux & Spire, 1994).…”

Phytophthora nicotianae transformants lacking dynein light chain 1 produce non-flagellate zoospores

“…Immune sera were collected after third and subsequent inoculations and tested in enzyme-linked immunosorbent assays. Mouse monoclonal antibodies directed towards PnLpv, PnCpa and PnVsv proteins in zoospore peripheral vesicles (Gautam et al, 1999;Hardham et al, 1986;Hardham and Gubler, 1990) were also used. Immunofluorescence assays were conducted using Phytophthora zoospores and mycelia fixed in 4% formaldehyde in 50 mM Pipes buffer as described previously .…”