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

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

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

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

NCLB RustSCLB

GLS FV AF

MSV

Focus Diseases

A field heavily infected with GLS

A field heavily infected with NCLB –Kakamega, Kenya

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

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

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 germplasm2. Screening tools3. Test locations with consistently high disease pressure

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 resistanceComplete resistance of a host to a specific race of a pathogenThe host plant exhibit hypersensitive reaction that prevents

the establishment and multiplication of the pathogenControlled by one (monogenic) or a few (oligoginic) genesPlants show distinct resistant and susceptible categories

Selection is thus easy in segregation populationsTransfer from source to other germplasm is also easy It is less durableHas been used to control very few disease in maize

Types of resistance

2. Horizontal resistanceEffect of resistance on the survival and reproduction of he

pathogen is less completeProvides 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 variationIt is more durable and stable due to the buffering effect of

polygenesHas been used for controlling most diseases in maize

The two types of resistance can co-exist

Types of resistance

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

Mechanisms for disease resistance1. 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 cells2. 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

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

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

Resistance screening methods • Field, greenhouse (screenhouse) and laboratory-based

screening techniques are available for the major diseases of maize

• Use established screening techniquesEffectiveCheapEasy to handle depending on available facilities and personnelHigh 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 location

(greenhouse, laboratory) at multiple-locations to expose them to different populations (races) of the pathogen

Test entries are exposed to adequate and uniform disease pressure.

Guarantee greatest differentiation of genotypes.

Objective of disease evaluations

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

Two major groups:

1. Naturally occurring epidemics

2. Artificially created epidemics

Disease screening techniques

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

hostsAdvantages Cheap and easy to manage Test materials are exposed to all pathogen racesDisadvantages 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

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

Naturally occurring epidemic of common rust Puccinia sorghi

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 =3Number of entries = 300 genotypes

Good phenotyping data across locations – Common Rust (Puccinia sorghi)

Oxalis sp. – an alternative host for Puccinia sorghi

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

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.

In the case of Fusarium ear rot -

Steps for preparing inoculum

Inoculating with Fusarium ear rot -

Silk channel inoculation technique Kernel puncher inoculation technique

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

Evaluation – Resistance vs. susceptible

Data loggers Inoculation technique

Fieldbook and fieldlog

Parameters for reliable disease phenotyping

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

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

1 2 3 4 5

Foliar disease

Diseases rating keys / scales

Evaluating ear rots

Multi-location Disease Phenotyping

Phenotyping Site MSV GLS Et Ear rots Ps PP BMHarare, Zimbabwe X X XMpongwe, Zambia XKakamega, Kenya X X XEmbu, Kenya X X XKibos, Kenya X XCatalina, Colombia X XEl Batan, Mexico X X XAgua Fria, Mexico X X X XSan Pedro , Mexico X

Acatic, Mexico X

= Natural condition; = Artificial condition

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

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 0.95LSD -05 1.13 0.76 0.98 0.55CV 22.60 14.14 19.84 9.61Replications =3Number of entries = 300 genotypes

Good phenotyping data across locations – Gray Leaf Spot (GLS)

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 0.079Var(Resid) 0.22 0.795 0.269 0.320

Grand_Mean 2.66 2.780 2.605 2.153LSD_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

Best - bet Sources of disease resistanceMean 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 2.13DTMA-11 CIMCALI8843/S9243-BB-#-B-5-1-BB-4-1-3 1.74 1.41 1.41 1.26DTMA-12 CIMCALI8843/S9243-BB-#-B-5-1-BB-4-3-3 1.71 1.72 1.79 1.63DTMA-13 CIMCALI8843/S9243-BB-#-B-5-1-BB-4-3-4 1.93 1.60 1.70 1.38DTMA-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 1.59DTMA-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 1.71DTMA-268 La Posta Sequia C7-F33-1-2-1-B-B 2.25 2.23 1.99 1.58DTMA-293 La Posta Seq C7-F153-1-1-1-2-B-B-B 2.50 2.35 2.33 2.43DTMA-40 [CML144/[CML144/CML395]F2-8sx]-1-2-3-

2-B*5 2.01 2.03 1.70 1.52DTMA-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 1.23DTMA-26 P502SRC0-F2-54-2-3-1-B 1.71 1.60 1.76 1.51

Disease resistance in best-bet DT linesMean Disease rating (1-5)

Stock ID Pedigree GLS (6 loc) NCLB(12 loc)

MSV (3 Loc)

Rust (5)

DTMA238DTPYC9-F46-1-2-1-2-B-B-B-B 2.87 2.67 2.41 2.28

DTMA-261La 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-176CLQ RCWQ103=

(CML150xCML254)-B-16-2-2-2-B-B-B-B-B

2.87 2.88 2.9 2.78

DTMA-233DTPYC9-F46-3-4-1-1-B-B-B-B-

B 2.46 2.97 2.39 1.38

DTMA-193CL-RCY015 2.16 - 2.93 2.36

DTMA-165S87P69Q(SIYF) 109-1-1-4-B 2.73 1.37 3.65 3.36

DTMA-217DTPWC9-F24-4-3-1-B-B-B-B-B 3.86 2.69 3.29 3.14

DTMA-62CLA44-B-B 3.74 3.17 3.45 4.59

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.

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

Thank you for your interest!