Breeding for disease resistance in maize new breeders course

1. Breeding for disease resistance in Maize
Stephen Mugo, George Mahuku, Dan Makumbi and C. Magorokosho, Suresh L.M.
Presentation made to the New Maize Breeder’s Training Course,
Lusaka, Zambia, 17 August – 4 Sept, 2015

2. Maize Diseases
 Maize production in sub-Saharan Africa is affected
by a wide array of diseases
 Environmental conditions prevalent in the different
agro-ecological zones are conducive to the growth
and spread of pathogens
 Different disease complexes affect maize production
in the lowlands and mid-high altitudes
 Diseases often reduce production and cause up to
100% yield loss under severe epidemics depending
on environmental conditions

3. Need to manage disease in maize
• Prevent economic losses
– Reduced yields
– Increased production
costs
– Poor quality grain
• Reduce or eliminate the
risks associated with
presence of a disease
• Guarantee food security

4. Constraints Lowlands Mid-altitude-highlands
Foliar diseases (reduce photosynthetic area)
Gray leaf spot (Cercospora zeae-maydis) x
Northern corn leaf blight (Exserohilum turcicum) x
Southern corn leaf blight (Bipolaris maydis) x
Common rust (Puccinia sorghi) x
Southern rust (Puccinia polysora) x
Downy mildew (Peronosclerospora sorghi) x
Maize streak virus disease x x
Ear rots (reduce quality of maize grain)
Diplodia ear rot (Stenocarpella maydis) x
Aspergillus ear rot (Aspergillus flavus) x x
Fusarium ear rots (Fusarium moniliforme) x x
Stalk rots (cause premature death of plants)
Diplodia stalk rot (Diplodis maydis) x
Fusarium ear rots (Fusarium moniliforme) x x
Charcoal rot (Macrophomina phaseoli) x
x =prevalent in the zone
Major maize diseases that significantly reduce maize
production in different ecological zones in SSA

5. NCLB RustSCLB
GLS FV AF
MSV
Focus Diseases

6. A field heavily infected with GLS

7. A field heavily infected with NCLB –Kakamega,
Kenya

8. Objective
• Identify superior disease resistant germplasm for
incorporation into breeding programs
• Collect good disease phenotypic data
• Use association mapping approaches to
understand the organization of disease resistance
genes in the maize genome
• Develop markers for marker assisted selection
breeding

9. Research Strategy
• High precision multi-location phenotyping:
– identify good sources of resistance for use as donors
– Validate stability of resistance genes
• Association mapping studies
– Leverage the DTMA AM set to identify chromosomal
regions involved in disease resistance
– Organization of disease resistance genes on maize
genome
• Fine mapping pipeline to develop functional
markers
– DH lines
– F2.3, BC1, BC2, BC3 populations

10. Breeding for resistance to diseases
 Use of disease resistant cultivars is the most valuable and
practical means to control diseases
 It is also inexpensive, effective, and simple to apply over a
wide area in a target production zone
Requirements for development of disease resistant maize
cultivars
1. Diverse germplasm
2. Screening tools
3. Test locations with consistently high disease pressure

11. Resistance is available for most of the economically important
diseases in maize
Resistance is controlled mainly by
 One or a few genes (monogenic or oligogenic)
 Many genes (polygenic)
 With additive and dominance effects
1.Vertical resistance
Complete resistance of a host to a specific race of a pathogen
The host plant exhibit hypersensitive reaction that prevents
the establishment and multiplication of the pathogen
Controlled by one (monogenic) or a few (oligoginic) genes
Plants show distinct resistant and susceptible categories
Selection is thus easy in segregation populations
Transfer from source to other germplasm is also easy
It is less durable
Has been used to control very few disease in maize
Types of resistance

12. 2. Horizontal resistance
Effect of resistance on the survival and reproduction of he
pathogen is less complete
Provides less selection pressure on the pathogen
It retards the infection process and slows down the spread of
the disease
Controlled by many genes (polygenic) each with small effect
Resistance shows continuous variation
It is more durable and stable due to the buffering effect of
polygenes
Has been used for controlling most diseases in maize
The two types of resistance can co-exist
Types of resistance

13. Factors affecting disease occurrence
* Climate change impacts the host, agent (pathogen) and environment
Environment
(favorable)
Pathogen
(virulent)
Host (Susceptible)
Man
•Temperature
•Relative humidity
•Rainfall
•Dew
•Solar radiation

14. Mechanisms for disease resistance
1. Resistance to pathogen establishment
•Immunity
– Prevent pathogen from establishing itself due to innate
structural or functional properties of the host
•Hypersensitivity
– Prevents pathogen survival and reproduction due to rapid
death of the host plant cells
2. Resistance to an established pathogen
– Restricts the ability of the pathogen to spread and
reproduce after becoming established in a host
3. Tolerance
– The plant exhibits severe disease symptoms without a
serious loss in yield

15. Availability of diverse germplasm for screening as
sources of resistance
• Sufficient genetic variation exists for most of the
diseases in maize
• Locally adapted or introduced maize germplasm
• Old varieties and breeding stocks
• Landrace collections
• Resistance alleles in these genetic resources can occur
at low or high frequencies
• Resistance genes occurring at low frequencies can be
gradually increased
• Genes at high frequency are easy to transfer

16.  Based on reliable identification of good sources of
resistance
 This can be done through:
 Development of standardized, highly efficient inoculation techniques and
disease rating systems for major maize diseases that include:
 Fusarium ear rot (FSR); Fusarium stalk rot (FSR); Gray
leaf spot (GLS); Tar spot complex (TSC); and Turcicum
leaf blight (TLB)
 Establishment of misting systems at key and crucial sites to create
microclimatic conditions suitable for disease development.
 Identification of disease hot spot sites, known for consistent, uniform and
reliable disease incidence and pressure.
Approach to minimizing production losses from
diseases

17. Resistance screening methods
• Field, greenhouse (screenhouse) and laboratory-based
screening techniques are available for the major
diseases of maize
• Use established screening techniques
Effective
Cheap
Easy to handle depending on available facilities and
personnel
High throughput for screening a large number of
breeding materials
• Field screening of breeding nurseries at hot-spot
locations with consistently high disease pressure is
also effective
• Evaluate selected resistant genetic materials in one

18.  Test entries are exposed to adequate and uniform
disease pressure.
 Guarantee greatest differentiation of genotypes.
Objective of disease evaluations

19.  Rate of progress to develop stable and durable
disease resistance or marker development depends
on:
 the use of reliable screening techniques
 use of as wide a spectrum of the pathogen as possible and at an
appropriate disease pressure
 Take note that:
 Low disease pressure
 Unreliable results that slow down rate of genetic
gain
 High or severe disease pressure
 Eliminate low level resistance inherent in adapted
germplasm and may drastically narrow the
germplasm base
Disease screening methods

20. Two major groups:
1. Naturally occurring epidemics
2. Artificially created epidemics
Disease screening techniques

21. Naturally occurring epidemics
 Hot spots
 Use of a location known for its high level of infection for a
particular disease
 Used for a pathogen with a local concentration of alternate
hosts
Advantages
 Cheap and easy to manage
 Test materials are exposed to all pathogen races
Disadvantages
 Success depends on year– to– year consistent expression of
epiphytotics
 adequate and uniform natural infections can rarely be
achieved in most locations
 Disease might not be evenly distributed within the field

22. Naturally occurring epidemics
 Enhanced natural infections to ensure adequate
disease levels
 Manipulation of planting dates
 Create favorable environmental conditions (e.g.
irrigation, enhanced drought etc)
 Use of spreader rows & use susceptible checks every
few rows (e.g. every tenth row)
 Sufficient replications (minimum of three)
 Multiple locations

23. Naturally occurring epidemic of common rust
Puccinia sorghi

24. Parameter Batan
(2008)
Batan
(2009)
Batan
(2010)
Combined
location
Entry Variance 0.92 0.69 1.28 0.45
Residual variance 0.39 0.31 0.17 0.28
Heritability 0.82 0.87 0.96 0.85
LSD -05 1.24 0.92 0.66 0.91
CV 22.53 25.50 12.12 20.40
Resistant Susceptible
Replications =3
Number of entries = 300 genotypes
Good phenotyping data across locations –
Common Rust (Puccinia sorghi)
Oxalis sp. – an alternative host for Puccinia sorghi

25. Artificially created epidemics
 Environmental conditions favorable for optimal
disease development rarely occur every year
 Great variation in the severity of the disease within a
location in a year
 Ensure adequate epidemic development
 Versatile - Can be done in laboratory, greenhouse
and field

26.  Inoculum production in
the maize pathology
laboratory in CIMMYT.
The inoculum is
produced on colonized
sorghum grain and
used for artificial
inoculations of leaf
diseases of maize (TLB,
MLB).
 Inoculations done at
the 6-8 leaf stage.
Colonized sorghum
seed serves as sources
of inoculum for 7
weeks under field.

27. In the case of Fusarium ear rot -
Steps for preparing inoculum

28. Inoculating with Fusarium ear rot -
Silk channel inoculation technique Kernel puncher inoculation technique

29.  Disease phenotyping hubs
 Harare
 Kakamega / Embu
 Misting system
 Environment conditions that
inherently favors expression of
plant diseases
 Protocols
 Well developed and available
for many disease
 Issues
 Lack of standardization
 Common checks
 Limits harmonization of data
from different organizations
Protocols for reliable disease phenotyping

30. Evaluation – Resistance vs. susceptible
Data loggers Inoculation technique
Fieldbook and fieldlog
Parameters for reliable disease phenotyping

31. Various degree of TLB infection on
different genotypes of maize despite of
using same inoculum, same inoculation
and observation time. Therefore, it
requires standardized disease rating
scale.
Disease evaluation

32. Rating scale 1-5 for common rust
(single leaf-based)

33. 1 2 3 4 5
Foliar disease
Diseases rating keys / scales

35. Evaluating ear rots

36. Multi-location Disease Phenotyping
Phenotyping Site MSV GLS Et Ear rots Ps PP BM
Harare, Zimbabwe X X X
Mpongwe, Zambia X
Kakamega, Kenya X X X
Embu, Kenya X X X
Kibos, Kenya X X
Catalina, Colombia X X
El Batan, Mexico X X X
Agua Fria, Mexico X X X X
San Pedro , Mexico X
Acatic, Mexico X
= Natural condition; = Artificial condition

37. P = 0.0001
EM-KN BA-Mex1 BA-Mex2 SL-Mex
EM-KN 1
BA-Mex1 0.86 1
BA-Mex2 0.87 0.96 1
SL-Mex 0.99 0.99 0.98 1
Phenotypic correlations between sites for
common rust

38. Parameter San Pedro
Lagunillas
(Nayarit)
Acatic
(Jalisco)
Santa
Catalina
(Colombia)
Paraguacito
(Colombia)
Entry
Variance
0.80 0.6 0.99 0.80
Residual
variance
0.25 0.15 0.36 0.11
Heritability 0.86 0.89 0.89 0.95
LSD -05 1.13 0.76 0.98 0.55
CV 22.60 14.14 19.84 9.61
Replications =3
Number of entries = 300 genotypes
Good phenotyping data across locations – Gray
Leaf Spot (GLS)

39. Good phenotypic data across locations
GLS (6) MSV (3) E. Turcicum
(12)
P. Sorghi
(5)
Var (Entry) 0.27 0.209 0.188 0.492
Var(LocxEntry) 0.45 0.121 0.182 0.079
Var(Resid) 0.22 0.795 0.269 0.320
Grand_Mean 2.66 2.780 2.605 2.153
LSD_0.5 0.87 2.510 0.729 1.399
H 0.76 0.62 0.91 0.91
#Sites 6 3 17 3
Combined Analysis Across Location: four adaptive diseases

40. Best - bet Sources of disease resistance
Mean Disease rating (1-5)
Stock ID Pedigree GLS (6 loc) MSV (3 Loc) NCLB(12 loc) Rust (5)
DTMA-3
[(CML395/CML444)-B-4-1-3-1-
B/CML395//DTPWC8F31-1-1-2-2]-5-1-2-2-
BB 1.43 1.12 1.74 1.30
DTMA-10 CIMCALI8843/S9243-BB-#-B-5-1-BB-2-3-4 2.06 1.60 1.67 2.13
DTMA-11 CIMCALI8843/S9243-BB-#-B-5-1-BB-4-1-3 1.74 1.41 1.41 1.26
DTMA-12 CIMCALI8843/S9243-BB-#-B-5-1-BB-4-3-3 1.71 1.72 1.79 1.63
DTMA-13 CIMCALI8843/S9243-BB-#-B-5-1-BB-4-3-4 1.93 1.60 1.70 1.38
DTMA-17
[CML312/CML445//[TUXPSEQ]C1F2/P49-
SR]F2-45-3-2-1-BBB]-1-2-1-1-2-BBB-B 1.87 1.12 1.80 1.59
DTMA-90 CML311/MBR C3 Bc F112-1-1-1-B-B-B-B-B 2.24 2.37 2.50 1.59
DTMA-146 [CML-384 X CML-176]F3-107-3-1-1-B-B-B 2.25 2.45 1.94 1.71
DTMA-268 La Posta Sequia C7-F33-1-2-1-B-B 2.25 2.23 1.99 1.58
DTMA-293 La Posta Seq C7-F153-1-1-1-2-B-B-B 2.50 2.35 2.33 2.43
DTMA-40
[CML144/[CML144/CML395]F2-8sx]-1-2-3-
2-B*5 2.01 2.03 1.70 1.52
DTMA-19
[CML312/CML445//[TUXPSEQ]C1F2/P49-
SR]F2-45-3-2-1-BBB]-1-2-1-1-1-BBB-B 2.20 1.61 1.77 1.23
DTMA-26 P502SRC0-F2-54-2-3-1-B 1.71 1.60 1.76 1.51

41. Disease resistance in best-bet DT lines
Mean Disease rating (1-5)
Stock ID Pedigree GLS (6 loc) NCLB(12
loc)
MSV (3
Loc)
Rust (5)
DTMA238
DTPYC9-F46-1-2-1-2-B-B-B-B
2.87 2.67 2.41 2.28
DTMA-261
La Posta Sequia C7-F180-3-1-
1-1-B-B-B-B-B 2.02 2.38 2.73 1.98
DTMA-79
(A.T.Z.T.R.L.BA90 5-3-3P-1P-
4P-2P-1-1-1-B x G9B C0
R.L.23-1P-2P-3-2P-3-2P-1P-B-
B-B)-B-16TL-3-1-4-B-B-B
2.19 2.94 2.44 2.2
DTMA-176
CLQ RCWQ103=
(CML150xCML254)-B-16-2-2-
2-B-B-B-B-B
2.87 2.88 2.9 2.78
DTMA-233
DTPYC9-F46-3-4-1-1-B-B-B-B-
B 2.46 2.97 2.39 1.38
DTMA-193
CL-RCY015
2.16 - 2.93 2.36
DTMA-165
S87P69Q(SIYF) 109-1-1-4-B
2.73 1.37 3.65 3.36
DTMA-217
DTPWC9-F24-4-3-1-B-B-B-B-B
3.86 2.69 3.29 3.14
DTMA-62
CLA44-B-B
3.74 3.17 3.45 4.59

42. Conclusions
• A large number of inbred lines, open-pollinated
varieties, hybrids, and source population with
resistance to the major diseases are available for
use
– As sources of alleles to breed maize for resistance to the
major diseases
• Other disease for which artificial inoculations are
conducted / protocols available
– Ear rots (Fusarium and Aspergillus), Stalk Rots, turcicum
blight, southern corn leaf blight, common rust.

43. Conclusions
 Information on sources of disease resistance on
CIMMYT website & available to collaborators
 Establishing diseases phenotyping network comprised
of different institutions and seed companies etc.
 Build capacity of collaborators on use of harmonized
disease evaluation protocols

44. Thank you
for your
interest!