Aerial Transport of Rust Fungi and Detection in Rain Les J. Szabo

Transcription

1 Aerial Transport of Rust Fungi and Detection in Rain Les J. Szabo USDAARS Cereal Disease Lab University of Minnesota St. Paul, MN

2 Introduction Rust biology Atmospheric transportation of rust spores

3 Rust Life cycle Spring Sexual Winter Asexual Summer Fall Puccinia graminis

4 Rust Life cycle Asexual 9 14 days Puccinia graminis

5 Spore production Single pustule 10,000 spores/24 h Field with 5% disease severity 50 pustules/tiller 2 trillion spores/hectare/24 h Puccinia graminis Roelfs, 1985

6 Spore cloud

7 Atmospheric transport Free Atmosphere Long distance transport (15 km) Atmospheric boundary layer Mixed layer 13,000 meters Sedimentation Rainout & Washout Source Target Target Adapted from Main et al., 2001

8 Transport into Free Atmosphere Convective updrafts Thunderstorms Fires Solar heating along the equator

9 Atmospheric transport Free Atmosphere Long distance transport (15 km) Atmospheric boundary layer Mixed layer 13,000 meters Sedimentation Rainout & Washout Source Target Target Adapted from Main et al., 2001

10 Atmospheric transport Local inoculum source

11 Atmospheric transport Local inoculum source

12 Atmospheric transport Longdistance inoculum source

13 Atmospheric transport Longdistance inoculum source

14 Aerial Transport Rust Spores Sugarcane rust Wheat Stem rust

15 Sugarcane Rust Puccinia melanocephala Asia and Africa Not found in America prior to 1978

16 PanAmerican epidemic July 3, 1978 Dominican Republic 1.6 ha area July 23, 1978 Dominican Republic 51 ha infected September, 1978 Jamaica October 11, 1978 Puerto Rico March 23, 1979 Canal Point, FL

17 Transoeceanic transport

18 Transoeceanic transport GOES Satellite image June 1997 Dust cloud took 5 days to cross from West Africa to the Caribbean Purdy et al., 1985

19 Transoeceanic transport Forward trajectories for air parcels Purdy et al., 1985

20 Regional transport Back airparcel trajectories Purdy et al., 1985

21 Sugarcane Rust: Summary Rust spores came from Cameroons, West Africa to the Dominica Republic June 1998 with transoeceanic winds Rust quickly spread through the Caribbean Rust spores spread from the Caribbean to Florida, February 1979 by wind in association with a tropical disturbance

22 Wheat Rusts Stem rust Puccinia graminis Stripe rust P. striiformis Leaf rust P. triticina P. recondita f.sp. secalis

23 U.S. Wheat 1998

24 Puccinia Pathway

25 Overwintering 35 30

26 Mean date of first Infection June 29 June 21 June 9 June 5 April 30

27 Spore monitoring

28 Spore monitoring: Airplane Asai, 1957 June 24 Ft. Riley Kansas City spores per m 3 Omaha Sioux City Brookings Fairmont Fairmont Fargo July1 Crookston Fargo Fargo Valley City Asai, 1960 Phytopathology

29 Spore monitoring 1957 First detection of stem rust in wheat fields Kansas June 3 Nebraska June 6 South Dakota June 21 Minnesota June 23 North Dakota July 4

30 Spore monitoring Spores collected were not the first introduction in the region since first field infections were detected before or near the same date as the first detection of spores in air samples Common problem found with other methods of air sampling for wheat stem rust spores These methods often sample rust spores from local sources, Rather than introduction of spores from distant locations

31 Spore monitoring: Rain Collected rain samples Rain collectors used a funnel that filtered the samples at the time of collection. Rust spores were separated from dirt particles by organic phase separation Roelfs et al., 1970 Phytopathology

32 Spore monitoring: Rain Method was inefficient in that each filter was examined by hand using a microscope Difficult to differentiate between different rust spores Stem rust was only one clearly identifiable

33 Realtime PCR Characteristics: Rapid Very sensitive Highly specific Multiplex

34 Realtime PCR: TaqMan PCR primer F Hybridization probe Q F Q

35 Realtime PCR Assay

36 ITS1 Primers and Probes Its1rustF10s 18s ITS1 5.8S Its1GrustR3s P. graminis P. recondita f.sp. secalis P. striiformis P. triticina

37 Realtime PCR Assay: Stem rust P. graminis specific probe Internal standard

38 Realtime PCR Assay: Sensitivity Determine the functional range of the assay lower limits of detection linearity of assay Serial dilutions of genomic DNA 0.5 pg 50 ng

39 Realtime PCR Assay: Sensitivity P. graminis P. recondita f.sp. secalis a b r 2 = r 2 = c r 2 = r 2 = P. striiformis P. triticina Lower limit was 0.5 pg or approximately 3 urediniospore

40 Trial samples Realtime PCR assays Field Spore Observ. Morph. Pg Pr Ps Pt Pc Stripe? Leaf? + Leaf? Leaf? + Stripe? Crown? + Stripe? Stripe? + Stripe? Stripe? + Stem? Stem? + Rust samples were from pastures around Hermiston, OR

41 Realtime PCR:Summary Rusts P. coronata P. graminis P. striiformis P. triticina P. recondita f.sp. secalis Nuclear ribosomal ITS1 region PCR primers are rustspecific TaqMan probe speciesspecific

42 Realtime PCR: Summary Assays are speciesspecific Assays are sensitive 0.5 pg 3 urediniospore linear over 10 5 fold DNA concentration Internal standard Ribosomal 28S PCR primers and TaqMan probe are rustspecific

43 Realtime PCR: Future Applying method to rain samples NADP collection sites Refining methods for removal of spores from filters

44 Rain collection sites National Atmospheric Deposition Program

45 Summary Aerial transport of rust spores Transoeceanic (Sugarcane rust) Transcontinental (Wheat stem rust) Rain is an efficient mean to monitor spore movement RealTime PCR assay developed 5 rust species Sensitive to approximately 3 urediniospores Developing assay for rain samples