1. Improving disease resistance
of maize in the developing
world: challenges and
opportunities
Santiago Mideros
17 February 2011
CIMMYT
WFP/Elmer Martinez

2. Outline
 Challenges and Opportunities
◦ General
◦ Specific  Aflatoxins
AFB1

3. Opportunities
 Ag. development is back
 Science & Technology
◦ Genetics, genomics, plant immunity
◦ Precise phenotype, barcoding, databases
 CIMMYT
◦ Germplasm
◦ World network
◦ Mandate

4. Challenges
 Climate Change
◦ Food security
◦ Maize pathology
 New or emerging pathogens/races
◦ Movement of germplasm
◦ Change on cultural practices
 Food quality, health concerns,
mycotoxins
 Science  Impact
 Policy

5. Climate Change
 Uncertainty in predictions
 Lack of historical data
 Fusarium species and
DON
Battisti and Naylor 2009

6. New and (re)emerging
pathogens or races
 Gray Leaf Spot
◦ Cercospora zeina
◦ Cercospora zeae-maydis
 Maize fine streak virus
 Maize streak
(gemini)virus
 Phaeosphaeria leaf
spot
Maize streak virus Shepherd et al

7. Policy
 Investment on science in developing countries
 Aid and technology from developed countries
◦ Budgetary concerns

8. Opportunities
 Ag. development is back
 Science & Technology
◦ Genetics, genomics, plant immunity
◦ Precise phenotype, barcoding, databases
 CIMMYT
◦ Germplasm
◦ World network
◦ Mandate

9. Opportunities
 Ag. development is back
 Science & Technology
◦ Genetics, genomics, plant immunity
◦ Precise phenotype, barcoding, databases
 CIMMYT
◦ Germplasm
◦ World network
◦ Mandate

10. Science
 Association Mapping  Yan et al, 2011
◦ Northern Leaf Blight,
◦ Southern Leaf Blight,
◦ Gray Leaf Spot
Wisser, Kolkman, Benson, Nelson, Balint-Kurti (in review)
 Glutatione S-transferase

11. Nested Association Mapping
Southern leaf blight (Kump et al, 2011)
 32 QTL
 51 SNP: Receptor like kinases, Glutathione S-
transferase
Northern leaf blight (Poland et al, in review)
 21 QTL
 60 SNP:
◦ Receptor like kinases
◦ Ethylene response factor
◦ Mlo-like
 Quantitative disease resistance  PTI (Basal
Resistance)
Gray leaf spot (Benson et al, in preparation)
 12 QTL
 21 SNP

12. Science (cont.)
 Mutant-Assisted Gene Identification and
Characterization (MAGIC)
• Fine mapping genes responsible for
resistance in maize

14. Nature Biotechnology Volume: 28, Pages: 365–369 Year published: (2010) DOI:
doi:10.1038/nbt.1613

15. Technology

16. Others
 Communication (Internet)
◦ Developing partnerships
◦ Collaboration
◦ Capacity building
 Genomic selection
 Genotyping by sequencing
◦ Next generation sequencing

17. Opportunities
 Ag. development is back
 Science & Technology
◦ Genetics, genomics, plant immunity
◦ Precise phenotype, barcoding, databases
 CIMMYT
◦ Germplasm
◦ World network
◦ Mandate

18. Germplasm
Photo by Suketoshi Taba, Yan et al
2011

19. Multiple Locations

20. Partners
 National agricultural
research institutes
 Other international
organizations
 Universities in
developing countries
 Universities in
developed countries
 Private companies
 NGOs

21. Sustainable Disease
Management
Disease resistance
 No silver bullets
◦ Deployment:
 Pyramids
 Refuge areas (Bt)
 Resistance +
management
 Multiple disease
resistance Effect of a single QTL. Chung et al,
2010

22. Outline
 Challenges and Opportunities
◦ General
◦ Specific  Aflatoxins
AFB1

23. Challenges and Opportunities
A practical example
Challenge:
Mycotoxin contamination
Opportunities:
Technology and genetics
Food systems

24. Mycotoxins AFB1
 Aflatoxins
 A. flavus and A. parasiticus 
aflatoxins
 Potent hepatocarcinogen
 Acute: death; liver damage
 Chronic:
◦ Immune suppression, growth
impairment
◦ Hepatocellular carcinoma
 Financial burden

25. Other Ear Rots
Gibberella Ear Rot Fusarium Ear Rot
Fusarium graminearum Fusarium verticillioides
Deoxynivalenol = DON = Vomitoxin Fumonisin
Photo: A. Robertson

26. Aflatoxin  child stunting

27. Regulation & Exposure
Country Consumptio Max. aflatoxin Exposure
n (ppb) (ng/bw/day)
(g/p/day)
Western 33 4 0.3 – 1.3
Europe
USA 86 20 0.26
Southern 148 4 0-4
Europe
Kenya 248 20* 3.4 - 133
Mexico 300 20 14 - 85
Sources:
Liu, Y. and Wu, F. 2010. EHP. 118: 818-824
Reddy et al. 2010. Toxin Reviews. 29: 3-26
1ppb = 1 ng per g or 1mg per metric ton

28. 2010 aflatoxin
outbreak, Eastern
Province, Kenya

29. Survey in western
Kenya 09
Source of maize N % >10 % >20
ppb ppb
Gifts from friends or 30 27 7
relatives
Purchased 34 11 2
9
Own-grown 18 5 2
3
Food aid S. Mutiga; V.14
7 0
Hoffman; and R.J. Nelson
unpb
Total 56 14 3
9

30. Strategy
Components of
Resistance
Map disease
resistance

31. CML69 Sc212m
Components of
resistance
 Tissue specific Mp339 Tx303
 Components of
resistance
◦ Latent period (LP)
Picture: K. Loeffler
◦ Sporulation intensity
(SP)
◦ Infection frequency
(IF)
 Colonization
◦ qPCR

32. qPCR Development
 A. flavus Primers:
◦ Internal Transcribed Spacer 1 (ITS1), 5.8S
rDNA, and ITS2
◦ Genbank
 A. flavus, A. oryzae, A. parasiticus, A. sojae, A.
tamarii, Emericella nidulans

33. A. flavus biomass in maize is strongly
correlated with aflatoxin concentration
A. flavus Aflatoxin
Pedigree/Line log (IC+1) IC log (ng/g+1) ng/g 4
CML103 0.78 a 1.18 10.03 a 22735
Mo17 0.62 ab 0.85 9.39 ab 11982
3
Coefficient (log IC)
B73 0.51 abc 0.67 9.54 ab 13917
B97 0.33 bcd 0.39 9.22 abc 10118
Infection
MS71 0.28 bcd 0.33 8.21 abcde 3689 2
Oh43 0.27 bcd 0.31 7.84 bcde 2542
Oh7B 0.21 cd 0.23 8.16 abcde 3483
IBM54 0.11 d 0.12 7.93 bcde 2790 1
NC358 0.11 d 0.11 6.75 defg 854
IBM262 0.09 d 0.10 8.75 abcd 6283
Ky21 0.08 d 0.08 6.96 defg 1056 0
2 3 4 5 6 7 8
Tx303 0.06 d 0.06 7.20 cdef 1333
Aflatoxin (log ng/g)
Ki3 0.05 d 0.06 7.46 bcde 1739
Mp339 0.05 d 0.05 6.79 defg 886
M37W 0.02 d 0.02 6.43 efgh 622 Correlation between infection
CML52 0.00 d 0.00 4.55 h 94 coefficient (IC) and aflatoxin
Mp313E 0.00 d 0.00 4.84 gh 126
CML247 0.00 d 0.00 5.15 fgh 172 = 0.85 **
Mideros et al 2009 Plant Disease 93:
1163

34. Experiments
New York Mississippi

35. Components of resistance for
diverse inbreds: 3 years of studies (P-
values)
Maize Tissue
Component of
resistance Developing Mature
Silk Kernel Kernel
0.062
Latent Period 4 ns
Infection 0.001
Frequency 8 ns
In-vitro
0.006
Sporulation 7 0.0963
Colonization
(qPCR) - 0.2097
Colonization 0.031
Field (qPCR) 2 - 0.0301
While significant,
Aflatoxin - - 0.0006
not correlated
with aflatoxin

36. Kernel resistance correlations
Field In-vitro Flint-Garcia's traits
Colonization
Colonization
Sporulation
%endsprm
Moisture
Carbhdrt
sd wght
Protein
(qPCR)
(qPCR)
Fiber
Ash
Fat
Aflatoxin 0.70*** 0.02 0.06 -0.03 -0.45 -0.25 -0.68** -0.57* 0.52* 0.57* 0.12
Field
Colonization -0.16 -0.01 -0.02 -0.21 -0.17 -0.47 -0.50 0.27 0.46 0.09
Colonization 0.00 0.33 -0.38 0.03 -0.07 0.03 0.23 0.14 -0.05
In-vitro
Sporulation -0.53* 0.39 0.27 0.03 0.26 -0.38 -0.41 0.15

37. Strategy
Dissect the trait into
multiple components
Map disease
resistance

38. Model selection QTL mapping (B73 x
CML322)
Trait Year Marker Position p-value source
DvK Sporulation 2007 m758 6.07 1.8E-03 B73
DvK Sporulation 2008 m758 6.07 3.7E-05 * B73
DvK Sporulation 2008 m838 7.06 7.8E-05 * B73
DvK Sporulation 2008 m209 2.03 1.0E-04 * CML322
Aflatoxin, MS 2008 m500 4.08 4.4E-06 ** CML322
Aflatoxin, MS 2008 m746 6.06 7.0E-05 * CML322
Aflatoxin, MS 2008 m35 1.03 2.2E-04 * CML322
Silk LP, NY 2008 m316 3.04 2.1E-10 ** CML322
Silk LP, NY 2008 m421 3.09 7.1E-10 ** B73
Silk LP, NY 2008 m540 4.10 2.6E-08 ** CML322
Silk LP, NY 2008 m154 1.11 2.7E-07 ** CML322
Silk LP, NY 2008 m706 6.03 1.0E-06 ** B73
Silk LP, NY 2008 m849 8.01 2.6E-06 ** B73
Silk LP, NY 2008 m168 1.11 1.3E-04 * B73
Silk LP, NY 2008 m857 8.03 1.6E-04 * CML322
Silk SP, NY 2008 m911 8.06 2.1E-08 ** CML322
Silk SP, NY 2008 m766 7.01 3.7E-08 ** B73
Silk SP, NY 2008 m428 3.09 1.0E-06 ** CML322
Silk SP, NY 2008 m763 7.01 4.9E-04 * CML322
Silk IC, MS 2008 m774 7.02 4.4E-05 ** B73
Silk IC, MS 2008 m775 7.02 5.0E-06 ** CML322
Aflatoxin, MS 2009 m821 7.04 1.0E-07 ** B73
Aflatoxin, MS 2009 m1096 10.07 1.0E-07 ** CML322
Aflatoxin, MS 2009 m216 2.03 9.3E-05 * CML322
Aflatoxin, MS 2009 m746 6.06 2.7E-03 CML322
Sporulation SP
25 Latent Period LP
QTL Infection Freq IF

39. QTL studies
Plant Disease Germplasm Source
Aspergillus Ear Rot Mp313E x Va35 Wilcox et al. (unpublished)
Mp313E x B73 Brooks et al. 2005
Mp717 x Nc300 Warburton et al. 2009
Mp715 x T173 Warburton et al. 2010
CML322 x B73 Mideros et al. (unpublished)
Gibberella Ear Rot CO387 x CG62 Ali et al. 2005
Fusarium Ear Rot 3 x 18 Perez-Brito et al. 2001
5 x 18 Perez-Brito et al. 2001
87-1 x Zone3 Ding et al. 2008
GE440 x Robertson-Hoyt et al. 2006
FR1064
NC300 x B104 Robertson-Hoyt et al. 2006

40. 5
1 2 3 4
5.00
1.00 umc1292 2.00 phi109642 3.00 umc2105 4.00 umc1008 5.01
umc1746 5.02 umc1761
2.01
umc1727 2.02 umc1185
1.01
umc1079
3.01 umc1970 4.01 umc1682
bnlg371 5.03 bnlg1046
umc1166
1.02 bnlg1429 umc1065 3.02 bnlg603
umc1070 2.03 5.04
bnlg371 4.02 umc1757
bnlg1144
3.03 mmc0081
bnlg1325 5.05
1.03 umc2019
5.06 mmc0481
phi001 4.03 bnlg1126
1.04 bnlg1598 2.04 umc1783 4.04 nc004
phi036
3.04 phi029
2.05
1.05 4.05
nc005 5.07 bnlg1306
2.06 3.05 sts001
umc1590
1.06 4.06
bnlg1057 bnlg2291 5.08 umc2136
2.07 umc1637
umc1035
1.07 bnlg1596 4.07 bnlg1137
3.06 bnlg1063
1.08 umc2047 umc1667
3.07 bnlg1350
1.09 bnlg197 4.08 5.09 umc1153
bnlg400
bnlg2162
2.08 umc1526 3.08
1.10 umc1051
umc1361
umc1173
umc1421 umc1062
1.11 3.09 4.09 umc1631
bnlg1182
umc1630
3.10 umc2048
1.12 umc1118 4.10 umc2011
umc1065
6
10
phi075 7
6.00 bnlg238 9
8 10.00
umc1498 umc1291
10.01
7.00 umc1642 9.01 umc1867
6.01
bnlg1165 8.00 9.02
umc1632 phi027
8.01 umc1139 9.03
6.02 bnlg1451
umc1688
Mp313Ea 6.03
umc1006
bnlg1194
umc1033
10.02
phi062
umc1191
Mp313Eb 6.04 umc1014
7.01 8.02
8.03
9.04 bnlg1209
umc2121 10.03
10.04
bnlg640
umc1246
bnlg1352 umc1053
Mp717 6.05
bnlg1154
8.04 bnlg1863
9.05
umc1654
10.05
umc1506
umc1898
umc1066
bnlg1702 bnlg666
Mp715
umc1477
bnlg1200 8.05
umc1149
6.06 umc2170 7.02 umc1978 bnlg240 9.06 umc1789
bnlg1350 10.06 umc1061
Oh516 umc1463
7.03
7.04
phi114
bnlg1070
8.06 umc1997
bnlg1191
Tex6 umc1001 8.07
bnlg1759 umc1029 9.07 bnlg128 umc1084
6.07
bnlg1828 bnlg1450
umc1248 7.05 phi260485
B73 umc2222 8.08
10.07
6.08 phi089 umc1407
umc2059 8.09 9.08 umc1277
7.06 phi116 dupssr14 bnlg1185
CML322 6.09 umc1127 umc1799
Collaboration with M. Warburton
10.08

41. QTL Meta-analysis
Consensus Genetic Map
Projection of QTL and CI
Select number of QTL in chromosome
Position of metaQTL
Veyrieras et al. 2007 BMC Bioinformatics 8
Truntzler et al. 2010 TAG 114

42. Consensus Genetic
Map
Mp313E x B73
Genetic 2008
Consensus B73 x CML322

43. QTL Meta-analysis
Consensus Genetic Map
Projection of QTL and CI
Select number of QTL in chromosome
Position of metaQTL
Goffinet et al. 2000 Genetics 155
Veyrieras et al. 2007 BMC Bioinformatics 8
Truntzler et al. 2010 TAG 114

44. 1 2 3 4 5
6 7 8 9 10
Aspergillus Ear Rot
Aflatoxin
Fusarium Ear Rot
Fumonisin
Ear rot (w)
Ear rot
Gibberella Ear Rot
Silk disease severity
Kernel disease severity

45. QTL Meta-analysis
Consensus Genetic Map
Projection of QTL and CI
Select number of QTL in chromosome
Position of metaQTL
Goffinet et al. 2000 Genetics 155
Veyrieras et al. 2007 BMC Bioinformatics 8
Truntzler et al. 2010 TAG 114

46. 1.01
2.5 Mb
1.06
23.2 Mb

47. Sustainable Disease
Management
Disease resistance
 Resistance +
management Effect of a single QTL Chung et al,
2010
Pre-harvest
resistance
Post-
Field Lower
Drought Better harvest
manageme aflatoxin
tolerance varieties manageme
nt s
nt
Yield •Resistance
Biological •Storage
control •Testing
•Detoxification

48. Acknowledgments R. Nelson Lab
o Chia-Lin Chung
 Rebecca Nelson o Jesse Poland
 Tiffany Jamann o Samuel Mutiga
o Judy Kolkman
 Paola Zuluaga
o Wenzhe Li
 Margaret Smith
o Chris Mancuso
 Jaci Benson o Nelson Chepkwony
 Bill Holdsworth o Ariel Fialko
 Jose Luis Zambrano o Oliver Ott
o Sara Heinz
 Marilyn Warburton o Lin-Si Hsieh
 Gary Windham o Kristen Kennedy
 Paul Williams o Kerri Lyons
o Gregory Flint o Joy Longfellow
o Ellie Walsh
o Ladonna Owens
o Mike Alpe
o Victoria Scott