Antifungal Compound Identification

Identify antifungal compounds with activity against Armillaria spp. that are naturally produced in Prunus (phytoanticipins).

More recently, an ornamental cherry species, Prunus maackii, was found to have ARR resistance (Warnstrom et al. 2011; Hammerschmidt, unpublished observations). Using a twig and root segment colonization assay, P. maackii displayed reduced colonization of Armillaria ostoyae compared to many other Prunus species. Bark extracts from P. maackii were subsequently found to inhibit growth of four species of Armillaria (Kanisewski and Hammerschmidt 2015; Hammerschmidt unpublished research). In particular, two anti-fungal compounds were isolated and characterized as 6-methoxyflavonoids (Kaniszewski and Hammerschmidt 2015). These flavonoids were characterized as phytoanticipins, pre-formed antimicrobial compounds produced by plants that confer host plant resistance. Thus, the resistance exhibited by P. maackii is due to the presence of these unique phenolic compounds that are present in periderm tissue providing a chemical barrier to Armillaria species.

Twig and root colonization assay.

Twig and root segments colonization assay (above). From left to right, a stem/root segment is placed on an actively growing colony of Armillaria (a, b); colonization of the tissue after peeling back the periderm (white coloration is the Armillaria mycelial fan) (c); infected tissue measurement (d).

Tissue culture.The third objective addresses the need to identify anti-Armillaria compounds in two known ARR resistant Prunus rootstock species, P. cerasifera and P. munsoniana, to contribute toward the breeding of ARR resistant rootstocks. Labs at Michigan State University are evaluating plant material suspected of being resistant using a twig and root colonization assay.

To seek new ARR resistance sources, the Gasic lab (Clemson University) adopted an in vitro screening protocol (Baumgartner et al. 2018) to evaluate 81 Prunus accessions, representing 28 species, from the USDA-ARS National Clonal Germplasm Repository in Davis, CA.

Microscopy image of roots.Microscopy is used to confirm the fungal presence and penetration from accessions that appear to be tolerant. Pictured to the right is a 40x magnification of the root (red) and fungus (green). A susceptible accession (left) displays fungal root penetratiion while the resistant genotype (right) shows no fungal penetration.

References:

Warnstrom, E. L., Outwater, C. A., Jacobs, J. L., & Hammerschmidt, R. (2011). Development of an in vitro bioassay to screen Prunus spp. for resistance to Armillaria ostoyae. Phytopathology, 1(6).

Kaniszewski, L. J., Hammerschmidt, R. (2015). Novel phytoanticipins from Prunus maackii: Possible factors in disease resistance. Michigan State University.

Baumgartner, K., Fujiyoshi, P., Ledbetter, C., Duncan, R., & Kluepfel, D. A. (2018). Screening Almond Rootstocks for Sources of Resistance to Armillaria Root Disease. HortScience53(1), 4-8.