Using Stigma Imprints to Determine Fire Blight Risk and Improve Control of Blossom Blight

1. Using Stigma Imprints to Determine Fire Blight Risk and Improve Control of Blossom Blight Scott C. Ockey Utah State Horticulture Association annual meeting Provo, Utah January 26-28, 2004

2. Stigma Imprint Background: • Fire blight bacteria multiply on the stigmatic surfaces of blossoms. • Stigma imprints indicate presence of bacteria on the stigma surfaces and percentage of blossoms within an orchard with epiphytic bacterial populations. • Stigma imprint results can be used to guide bactericide applications. • Current stigma imprinting protocols provide results in 36-48 hrs.

3. 2003 Objectives: • Improve timeliness of stigma imprinting results by bringing stigma imprinting closer to home. • Improve timeliness of stigma imprinting by implementing current molecular detection procedures. • Continue to work on stigma imprinting technique and evaluate stigma imprint data.

4. Improve timeliness of stigma imprinting results. • We continued having Lester perform stigma sampling. • Previously poured Petri plates were on hand at Mountainland. • Lester followed an established sampling protocol in each selected orchard.

5. We brought the lab to Mountainland. • Incubator—reduced results to 24-30 hours. • Microscope—Verified ease of identification.

6. Molecular Detection Procedure rtPCR- real time Polymerase Chain Reaction • Used a 944bp segment of DNA common to all virulent Fire Blight Bacteria. • Modified PCR protocol published by Al Jones et al. to rtPCR. • Used Idaho Technologies R.A.P.I.D. rtPCR unit. • Set up molecular grade lab at Mountainland packing shed.

10. What Happened????? • Strong inhibitors present on stigmatic and blossom surfaces. • Length of replicated DNA strand to long for rtPCR procedure. • Complex of both factors.

11. What Next??? • Evaluate shorter DNA sequences. • 447bp sequence found PEANT. • New protocol tested. • Work with published protocols for removing inhibitors. • Use of Llop extraction procedure developed specifically for bacterial plant pathogens.

12. Testing done on crabapple stigma and blossom using PEANT primers and/or Llop extraction protocol.

13. Continue to work on stigma imprinting technique and evaluate stigma imprint data. • Stigma imprinting with Streptomycin amended media provides resistance information. No Strep Strep

14. Fire blight Bacteria present before Maryblyt, Cougarblight or the temerature mean models predicted. • Stigma imprints indicated that 62% of the colonized flowers had streptomycin resistant strains. • Suggested spraying blossoms with Mycoshield.

15. Fire blight Bacteria present before Maryblyt, Cougarblight or the temerature mean models predicted. • Stigma imprints indicated that most colonized flowers had streptomycin resistant strains. • No sprays applied, 8strikes/tree some trees near cankers had 95% blossoms infected. Infection occurred at low temps.

16. Blossom blight infections can and do occur when climatic conditions and models suggest otherwise. • Numerous over wintering cankers increases blossom blight risk even with non-conducive climatic conditions. • Two row planting systems inhibit spray movement, resulting in spray coverage problems. • Streptomycin and Mycoshield applications do not reduce bacterial populations on the stigma. However, they do coat the floral cup to reduce infection potential.

17. Stigma imprints may be able to help identify “hot spots” within an orchard. • Set a sampling route to be followed each sampling day. • Mark plates to correspond to points on the sampling route. • Check for bacterial growth on plates and determine if there is an occurrence pattern. • Go back to the orchard and check for overlooked cankers or other causes (beehives).

18. We would like to thank: USHA Mountainland Grower Cooperators Idaho Technologies Doug Rowley Lester Ryan Black