Management, host pathogenicity, and rapid identification of Magnaporthe poae, causal agent of summer patch on annual bluegrass and Kentucky bluegrass turf.

Principal investigators:
Katerina S. Jordan

Project summary:

With increasing demands placed on golf course putting green turf combined with increased temperatures over the last few years, the incidence of diseases such as summer patch have increased throughout Ontario. Summer patch is a root disease caused by the fungus Magnaporthe poae and it is most pathogenic on annual bluegrass (Poa annua), Kentucky bluegrass (Poa pratensis), and fine fescues (Festuca spp.). The pathogen grows best under conditions of warm air and soil temperatures and high soil moisture. Symptom development can occur at any time when the turfgrass is stressed, although the above conditions are usually when symptoms develop on turf as well.

The disease is managed primarily through preventative chemical applications in conjunction with cultural practices. However, the appropriate method of pesticide application (e.g. application volume, additional irrigation) and the effects of various cultural practices on pathogen survival and disease development are not well known for the disease on annual bluegrass putting greens. In addition, pathogenicity and natural resistance in annual bluegrass ecotypes are not known in Ontario. Finally, proper diagnosis of this disease relies on visual observation of symptoms and signs on the roots, both of which can sometimes be misleading.

The objectives of this study have been to develop best management practices for summer patch, determine host specificity and pathogenicity to gain a better understanding of disease development, and to develop a rapid and simple diagnostic tool for presence of Magnaporthe poae in the hopes of decreasing fungicide use by increasing the efficacy of preventive applications.

CTRF Update – February, 2011

Molecular work for identification of Magnaporthe poae

Small sections of cup cutter sized-samples collected from various golf courses in Summer 2009 that showed signs of colonization with fungi believed to be Magnaporthe poae were planted, grown and maintained in a growth chamber (set at 28ºC). Once plants began to show foliar symptoms, roots were examined microscopically. Roots containing dark runner hyphae (ectotrophic mycelia) believed to be colonized by M. poae were isolated, sterilized and plated on antibiotic-amended potato dextrose agar (APDA). Plates were examined for visible fungal growth, reminiscent of M. poae. Cultures have been maintained on fresh media through periodic re-culturing.

To definitively identify/diagnosis M. poae in samples collected, molecular/genetic work has begun. DNA was isolated from a few of the cultures and amplifed, using primers specific for the ectotrophic root-infecting fungi, through polymerase chain reaction (PCR). To ensure PCR was successful, a small quantity of each PCR product was run on an agarose gel. As shown in Figure 1, five DNA bands (~ 800 base pairs (bp) in size) were evident and appropriate for M. poae according to previously published literature.

Once PCR products were confirmed, PCR purification was performed. Purified samples were sent to the University of Guelph Genomics Facility for sequencing. Unfortunately the samples did not sequence successfully (no sequencing signal), likely due to the loss of the PCR products during purification. Molecular/genetic work is continuing to date and samples will be re-submitted to the U of G Genomics Facility but will not be purified prior to submission.

Field study 2010

A preliminary field study was conducted during the 2010 season at the Guelph Turfgrass Institute (GTI) on the Poa annua green established for this project. The randomized trial consisted of seven treatments including a variety of fungicide application regimes and an untreated control (Figure 2). Four replicates each of preventative application of azoxystrobin (watered-in and not watered-in) and curative application of azoxystrobin (watered-in and not watered-in) were performed. Various control treatments (no inoculum + no azoxystrobin controls, no inoculum + azoxystrobin, and inoculum + no azoxystrobin) were used.

A preliminary infiltration study was performed prior to the start of the trial, in which 20 L of water per 2m2 plot was applied for the watered-in treatments. Inoculum of M. poae was prepared using Kentucky bluegrass seed and various fresh cultures collected from courses throughout southwestern Ontario. The inoculum was grown for several weeks, dried and then weighed into separate envelopes for use in the field study. Plots were watered after the inoculum was applied (to the appropriate plots) and twice daily throughout the entire trial duration. Photographs, disease severity ratings, Normalized Difference Vegetation Index (NDVI) and visual observation of the soil using core samples were taken during the trial to assess treatments. As well, cup cutter samples were taken from each plot at the end of the trial. Further research and data analysis using the disease severity ratings and NDVI readings is currently underway. This trial will be repeated in summer 2011, along with newly developed trials.

Determination of pathogenicity and potential resistance:

During summer 2010, a number of turf samples from golf courses were submitted through the turfgrass diagnostic laboratory. Samples with signs and symptoms suggestive of Magnaporthe poae & summer patch, respectively, were collected. Roots containing dark runner hyphae were isolated, sterilized and prepared on culture media. Cultures were grown and maintained (fresh & periodic re-culturing) as previously discussed.

Regular correspondence with volunteer golf course superintendants is also maintained for sample collection. In addition to collecting numerous isolates of M. poae, various ecotypes of Poa annua were collected and are being maintained for future pathogenicity testing. Pathogenicity tests will commence in winter 2011 using M. poae cultures received from the American Type Culture Collection (ATCC) as well as with the isolates collected from Ontario golf courses. All cultures are grown and maintained on PDA media.

Once we have determined both morphologically and genetically (through DNA analysis) that M. poae is indeed present in the samples, work will begin to develop a diagnostic assay. The plans for the summer of 2011 are to inoculate P. annua in the field and conduct further fungicide and cultural management studies and to continue to collect isolates of M. poae and Poa annua ecotypes.