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Module 5:

Breeding for Disease Resistance


I.
II.

Introduction
Some Concepts of Plant Pathology
a. Bacterial pathogens
b. Fungal pathogens
c. Nematode pathogens
d. Viral pathogens
e. Phytoplasm pathogens
f. Parasitic plants
g. Insect pathogens
III. Genetics of Resistance
IV. Breeding for Resistance to Rusts
V. Breeding for Resistance to Fusarium Head Blight
VI. Breeding for Resistance to Septoria tritici Blotch
VII. Breeding for Resistance to Karnal Bunt
VIII. Breeding for Resistance to Nematodes

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I. Introduction

"There is no good theory of disease which


does not at once suggest a cure."
Ralph Waldo Emerson

".when the cause of disease is discovered,


consider that the cure is discovered."
Cicero

Early civilizations were well aware that plants were attacked by diseases. There
are references in the Bible to blights, blasts, and mildews (Haggai 2:17, 2 Chronicles
6:28, Amos 4:9). Aristotle wrote about plant diseases in 350 B.C. Theophrastus, the
father of botany (372-287 B.C.), in his Historia Plantarum (History of Plants),
described rusts on grain crops, in meticulous details. During the Middle Ages in Europe,
ergot fungus infected grain and Shakespeare mentions wheat mildew in one of his plays
("He Mildews the White Wheat": King Lear). In India in about the year 500, the
situation arose in which plants were thought to be suffering from human ailments. Thus,
trees and other plants were thought to suffer from wind, bile, phlegm, jaundice and
indigestion. Mathieu Tillet, in 1755, while observing the smuts on wheat, discovered
that there were two types of smut: la carie or what the English called "Common Bunt" or
"Stinking Smut"; and the second type of smut le charbon or what the English called
"loose smut". The distinction between the two types of smut was verified a century later,
in 1847, by Louis and Charles Tulasne. To honor Tillet, they named the "Stinking Smut"
Tilletia caries. In 1807 Isaac Benedict Prevost was the first to provide definitive
evidence that the bunt disease of wheat is caused by a microorganism, smut fungus. He
completed detailed experiments on the germination of bunt spores and demonstrated by
direct inoculation that they could infect wheat. In 1853 Heinrich deBary published a
comprehensive paper that clearly implicated smut and rust fungi as causal organisms of
diseases affecting cereal crops.
In this and the last century, plant breeders have developed cultivars with genetic
resistance to possibly devastating plant pathogens. As this effort has curbed the
widespread famine formerly caused by wheat pathogens it can be regarded among the
most important contributions to agriculture. However, agriculture is a dynamic trade.
Changing agronomic practices and the evolution of new virulent races of pathogens,
requires a persistent and continuous effort in disease management.
An example of this needed vigilance is occurring now (2007) as a new form of
stem rust, has jumped from eastern Africa and is now infecting wheat in Yemen in the
Arabian Peninsula. Researchers with the Global Rust Initiative (GRI) and the
Agricultural Research Service of the United States Department of Agriculture (USDAARS) have confirmed conclusively the existence of the disease in Yemen. There is also

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evidence that the disease has spread into Sudan but more tests are needed to confirm the
finding. Until this discovery, this new strain of stem rust, known as Ug99, had only been
seen in Uganda, Kenya and Ethiopia.
There is precedence for this, from a virulent strain of another wheat disease,
called yellow rust, which emerged in eastern Africa in the late 1980s. Once it appeared in
Yemen, it took just four years to reach wheat fields of South Asia. On its way, this new
strain of yellow rust caused major wheat losses in Egypt, Syria, Turkey, Iran, Iraq,
Afghanistan, and Pakistan, exceeding USD 1 billion in value.
There is every reason to believe the new Ug99 strain of stem rust represents a
much greater risk to world wheat production. Annual losses of as much as $3 billion U.S.
in Africa, the Middle East and south Asia alone are possible.

II. Some Concepts of Plant Pathology


Diseases of wheat are numerous and are caused by: bacteria, fungi, nematodes,
viruses, phytoplasmas, a single parasitic plant, and insects. For a complete list of known
pathogens of wheat, excluding insect pests see:
http://www.apsnet.org/online/common/names/wheat.asp
(From the homepage follow ONLINE RESOURCES to COMMON NAMES OF PLANT DISEASES
to TABLE OF CONTENTS and to #96 WHEAT.)

In order to properly develop a defense against a pathogen, the wheat breeder must know
the attacker. The breeder must understand the pathogens life cycle (inoculation,
infection, proliferation, spread, and latency), its virulence during different
environmental conditions and varying stages of wheat growth, along with its
epidemiology. The breeder must be able to spot signs of the presence of a pathogen and
the breeder must be able to recognize, distinguish, and describe the symptoms that the
wheat plant is displaying. Finally, the breeder must understand the economic impacts of
a particular pathogen in order to determine the amount of resources that should be
directed to resolving the problem.
Inoculation - The introduction of a pathogen to a host.
Infection - The pathogen penetrates and establishes a parasitic relationship with the host.
Proliferation - The reproduction and growth of the pathogen.
Spread - The process by which the pathogen moves within a single host plant or the
process of moving from one infected plant to another.
Latency - A time of inactivity of the pathogen (dormancy in advance of proper
environmental conditions).
Virulence - The degree of pathogenicity of a given pathogen.
Epidemiology - The study of factors influencing the initiation, development, and spread of
infectious disease; the study of disease in populations of plants

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II a) Bacterial pathogens
Bacterial plant pathogens are unicellular microorganisms typically 1 to 3 m in
length. They have neither a well defined nucleus, nor a nuclear membrane. Bacteria are
spread by insects, air currents, splashing rain, and by mechanical means. Free moisture is
usually necessary for infection, and penetration of host tissue occurs through wounds
(created by hail, blowing soil particles, insects, or mechanical injury) or leaf openings
(stomata and hydathodes). Bacteria reproduce chiefly by binary fission, or cell division
yielding two identical daughter cells.
Symptoms caused by bacterial diseases may vary greatly, depending on the
pathogens involved. A bacterial origin is suggested by water-soaked spots or watersoaked margins around lesions consisting of water-congested green tissue in the early
stages of infection. The lesions are greasy and translucent in appearance and may
produce exudates. An exudate consists of droplets of bacterial slime emerging from the
leaf surface through natural openings (stomata, hydathodes).
Fig. 1 Example of a bacterial disease cycle

Duveiller et al. 1997

Further information on bacterial diseases of wheat can be found in the CIMMYT


publication: The Bacterial Diseases of Wheat: Concepts and Methods of Disease
Management. Edited by: E. Duveiller, L. Fucikovsky, and K. Rudolph. Available in
PDF on line at: http://www.cimmyt.org/

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II b) Fungal pathogens
Fungal pathogens of wheat are diverse and constitute the most problematic and costly of
the biotic stresses of wheat. Fungi are eukaryotic, non-vascular, saprophytes (taking
nourishment from non-living material), or parasites (removing nutrients in a harmful way
to a living host). Fungi reproduce by means of spores, both asexually and sexually
depending on the species and environmental conditions. Fungal spores spread from plant
to plant through wind, water splashing, and by mechanical means. Fungi infect their host
when a germ tube growing from a spore enters a plant stomata; or by piercing the cuticle
and epidermal cell wall by physical and/or enzymatic means. Within the host, fungi
spread by filamentous growth of hyphae forming a mycelium. The lifecycles of the
different fungal pathogens can be extremely different from one another; and in order to
develop a proper genetic defense against them, a solid understanding of the life cycle of
each pertinent fungal pathogen is important.
Fig. 2 Example of a fungal pathogens life cycle

Source: USDA ARS

Further information can be found on a number of fungal pathogens of wheat in the CIMMYT
publications: Bunt and Smut Diseases of Wheat: Concepts and Methods of Disease
Management. Edited by: R. Wilcoxson, and E. Saari; The Septoria Diseases of Wheat:
Concepts and Methods of Disease Management. Edited by: G. Hettel; Septoria and
Stagonospora Diseases of Cereals: A Compilation of Global Research. Edited by: M. van
Ginkel, A. McNab, J. Krupinsky. Available in PDF on line at: http://www.cimmyt.org/ ; Rust
Diseases of Wheat: Concepts and Methods of Disease Management. Edited by: G.P. Hettel.

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II c) Nematode pathogens
Nematodes are a diverse group of worm-like animals. They are found in virtually
every environment, both as parasites and as free-living organisms. They are generally
minute, but some species can reach several meters in length. Plant parasitic nematodes
can be aerial parasites (those feeding on above-ground plant parts) or root and tuber
parasites (those feeding on below ground parts). Nematodes can further be classified as
migratory endoparasites (mobile, feeding inside the plant), sedentary endoparasites (cease
moving once a feeding location inside the plant has been reached), or ectoparasites (feed
on the outside of the plant). The nematode life cycle is typically divided into six stages:
the egg, four juvenile stages, and the adult. The duration of these stages and of the
complete life cycle differs for different species and depends on environmental factors.
Fig. 3 Example of a Typical Nematode Disease Cycle.

feeding on host
cells and tissue

Juvenile 3
Juvenile 4

Second stage
Juveniles hatch
and are attracted
to host root
exudates

Juvenile 2

Adult
Juvenile nematodes
develop inside the
egg
male and female
nematodes mate
and eggs are laid in
the soil

Source: American Phytopathological Society


Vicky Brewster

Further information on nematodes can be found in the CIMMYT publication: Common


Diseases of Small Grain Cereals: A Guide to Identification. Edited by: F. Zillinsky.
Available on line at Grain Genes (http://wheat.pw.usda.gov/GG2/index.shtml).

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II d) Viral pathogens
Viruses are sub-microscopic particles consisting of genetic material (either DNA
or RNA) contained within a protein coat known as the capsid. Viruses are obligate
parasites as they can replicate themselves only by the use of another organisms cellular
machinery and intermediate products. Viral infection occurs through wounds, generally
being caused by an insect or nematode vector. Viruses can also be spread by fungi, by
mechanical means or by being carried on in the seed of infected plants.
Fig. 4 Example of Viral Disease Cycle
Spring

Summer

Mites
Infected
winter wheat

Virus

Fall

Mites
Virus

Mites
winter wheat
seedlings

volunteer
wheat

hosts other than


wheat

Winter

Virus

Overwintering
winter
wheat
crop and
other
hosts

Arrows indicate
virus movement
Figure adapted from Nebraska Cooperative Extension

Further information on nematodes can be found in the CIMMYT publication: Common


Diseases of Small Grain Cereals: A Guide to Identification. Edited by: F. Zillinsky.
Available on line at Grain Genes (http://wheat.pw.usda.gov/GG2/index.shtml).

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II. e) Phytoplasm pathogens


Phytoplasmas are similar to bacteria, but unlike bacteria they have no cell wall.
They reproduce as bacteria do and are usually found in the water- and food-conducting
vessels of an infected plant. Phytoplasmas and spiroplasmas are obligate parasites.
Phytoplasmas are transmitted from plant to plant by insects that feed off of phloem tissue
Fig. 5 Example of a Phytoplasm Disease Cycle

Insect feeds on
annual or perennial
plants, Phytoplasm
is incubating and is
not yet transmitted

Phytoplasm present in
salivary glands in
large numbers and is
injected into plant
while insect is feeding

Insect vector feeds on


vein of infected plant
Insect vector feeds on
infected plant, picking up
the phytoplasm and
delivering it to a new plant
Phytoplasm
overwinters in
perennial
plant

Phytoplasm spreads by
phloem throughout the
plant

Picture by: S. Mahr; Drawings from: USDA-ARS; Information adapted from: Gianna Sassi Plant Pathology 401 Cornell University

Further information on phyoplasms can be found in the CIMMYT publication: Common


Diseases of Small Grain Cereals: A Guide to Identification. Edited by: F. Zillinsky.
Available on line at Grain Genes (http://wheat.pw.usda.gov/GG2/index.shtml).

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II f) Parasitic Plants
Parasitic plants are flowering plants that gain some or all of their sustenance from another
plant. They do this by penetrating host plant vascular tissue through modified roots
called haustoria. Striga spp. are obligate root parasites that infect many of the worlds
principal grain and legume crops. Although endemic to Africa, Striga spp. occur in over
40 countries worldwide, ranging from Asia to North America. In sub-Saharan Africa,
more than two-thirds of the 73 million ha of land used for cereal cultivation is severely
infested. Over time, the impact of Striga spp. has increased and it has been described as
one of the most serious biological constraints to food production in Africa (Vasey et al.
2005).

Fig. 6 Example of a
Parasitic Weed Disease
Cycle

Weeks 4-7
Striga emerges
above ground

Week 9-12
Flowering of
Striga

Week 11+
Capsules form
on Striga

Week 11+
Dissemination
of Striga seeds

Host
plant

Aerial Phase
Underground phase

Striga shoot grows


utilizing the host root
system for water and
nutrients
Day 6 to weeks 4-6

Germination
Days 1-2
Attachment to
host root
Day 2-6

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Adapted from Stringer, 2007

II g) Insect pathogens
The number of insect pests that feed on wheat is numerous. Insects cause injury
by feeding, and predispose weakened plants to disease. They produce wounds that serve
as infection sites, and often carry pathogens with them from plant to plant. Insects are by
far the most numerous and most efficient vectors of viruses and phytoplasmas. Insects
with chewing mouth parts remove tissue or cut off young plants. Insect larvae feed on,
above and below ground plant parts as well as inhabiting and feeding off of the inside of
the plant. Insects can also be problematic, feeding on the grain both pre- and postharvest.
Fig. 7 Example of an Insect Pathogen Disease Cycle
The pupae overwinter/oversummer
within puparia, the hardened skin of
the last instar larvae. These puaria,
known as the flaxseed stage are
located just below the surface near
the crown of the plant.

Flies emerge from the


flaxseed stage and
disperse throughout
the crop

The maggots enter the


flaxseed stage for
overwinter/oversummer

Flies deposit eggs on


wheat and die in 2 or 3
days

Maggots hatch from the eggs in 3 to 7 days, crawl down the leaves,
and feed at the crown or joints along the stem. The maggots develop
through three instars over a 25 to 30 day period.

Source: http://www.oznet.ksu.edu/hessianfly/

Further information on insect pathogens can be found in the CIMMYT publication:


Common Diseases of Small Grain Cereals: A Guide to Identification. Edited by: F.
Zillinsky. Available on line at Grain Genes (http://wheat.pw.usda.gov/GG2/index.shtml).

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III. Genetics of Resistance


Farmers have noticed for centuries that some individual plants in a given species
manage to survive disease or epidemics of insects relatively unscathed, while their
neighbors succumb to infection or insect predation. In 1905, Sir Roland Biffen of
Cambridge, England, wondered whether healthy plants inherited pest resistance, just as
they might inherit the tendency to be tall or short. His experiments on two varieties of
wheat showed that the ability to resist infection by a rust fungus was indeed inherited in
Mendelian fashion, a discovery that intensified attempts by farmers and plant breeders to
produce varieties of pest-resistant crop plants.
About 50 years later a plant scientist by the name of Harold Flor, while working
with flax and the rust fungus Melampsora lini, found that host resistance to a pathogen
was not only dependent on the host genetic makeup (resistance genes, R-genes), but also
on the genetic makeup of the pathogen (avirulence genes, Avr-genes). With these
findings Flor introduced the widely accepted concept of the gene-for-gene theory of
disease resistance (often referred to as race-specific or vertical resistance), which
predicts that a host will achieve successful disease resistance only if both a hosts Rgenes and the pathogens corresponding Avr-genes are present. Since that time many RAvr interactions have been studied and documented. Dozens of R-genes, against many
different pathogens have been identified from a variety of plants. These genes encode
proteins that can be grouped into superfamilies based on sequence and structure similarity
(McDowell and Woffenden 2003); and based on their structure it can be deduced that
some of these R-proteins are found in the cell cytoplasm, some are anchored to the cell
membrane, and some traverse the cell membrane (Dangl and Jones 2001). There are two
models proposed for R-Avr interaction and are depicted in figure 7.

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Apoplast

Cytoplasm
Host defenses
are suppressed
and disease
occurs

(a)

Signal
transduction

Defenses
activated

(b)

Signal
transduction

Defenses
activated

R
Adapted from McDowell and Woffenden 2003

(c)

Fig. 7 R-Avr Interaction. A pathogen (grey box) has come into contact with a potential host and is
expressing virulence proteins (red). Once inside host cytoplasm, the virulence genes target host defense
response proteins (green). (a) The plant cell does not hold an R-protein capable of recognizing any
virulence protein, and disease results. (b) Pathogen Avr-protein is recognized by a host R-protein,
activating a signal cascade and a defense response. (c) The guard hypothesis: a host defense response
protein has been altered by a virulence protein, this is detected by the host R-protein and a signal
cascade followed by defense mechanisms being activated soon follows.

Plant defense starts with non-self recognition and/or cellular intactness. During
host invasion, pathogens release exogenous as well as endogenous elicitors (Vorwerk et
al. 2006; Huckelhoven 2007). Elicitors are molecules produced by either the host or the
pathogen that in turn induce a response by either the host or the pathogen. Fungal
pathogen elicitors include enzymes which break down the cuticle or cell wall
polysaccharides. Chitin, the major constituent of fungal cell walls is also known to be an
elicitor which triggers a defense response in plants. The bacterial protein, flagellin, is also
an elicitor of host defense. Cuticle and cell wall fragments are examples of endogenous
elicitors of host defense. As these tissues are under physical or enzymatic attack,
released fragments are detected by R-proteins stationed in the cell membrane and a
defense response is initiated.
Within 15 minutes of pathogen recognition, host cells begin producing new
proteins in reaction to the attack (Dangl and Jones 2001). This first response to invasion
is known as a basal defense, and includes production of proteins that inhibit pathogen
enzymes, structural and chemical remodeling of the host cell wall at penetration sites, and
production of antimicrobial agents that will kill the intruder. A second, more extreme,
line of defense is known as the hypersensitive response (HR), or programmed cell
death (PCD) (Greenberg and Yao 2004; Lam et al. 2001; Day and Graham 2007;
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Glazebrook 2005). The hyper sensitive response is a strategy of scorched earth where
the plant cell introduces toxic molecules into the surroundings; creating a localized
environment that is incapable of sustaining life for both the pathogen and the cell itself.
The HR is an efficient means of defense against many pathogens, but is ineffective
against necrotrophic fungi.
Breeders have successfully developed lines resistant to diseases by integrating Rgenes into their cultivars for many years; but a durable (sometimes called Horizontal
Resistance, Race non-specific resistance, or Qualitative Resistance), long lasting
resistance in many cases has been difficult to achieve as pathogens quickly evolve and
develop counter resistance genes that circumvent the host cultivars resistance. Breeders
often spot this breakdown in resistance and hurriedly integrate a newly found effective Rgene into their populations. In time, the new R-gene loses its effectiveness and the boombust and induced co-evolution between crop and pathogen continues. An example of this
comes from the United States where in Indiana, 1955; a cultivar was released that held on
R-gene conferring resistance to Hessian fly (Meyetiola destructor) attack. Just six years
later the Hessian fly population had developed a substantial amount of counter-resistance.
By 1964, the Indiana breeders had introduced a second R-gene into their cultivars. This
new R-gene provided resistance for eight years before counter-resistance developed. A
third R-gene was introduced and released in cultivars in 1971; and again was overcome
by the pathogen, this time in a period of ten years (Rausher 2001).
Two methods are available to plant breeders in order to increase the durability of
their resistant cultivars. The first is known as the High-Dose/Refuge or Multiline
strategy (Rausher 2001; Pink 2002). Recall that a hosts resistance to a disease is
conferred by the interaction of its R-gene with the pathogens Avr-gene. A change due to
a mutation of the pathogens allele for an Avr-gene will allow the pathogen to circumvent
or suppress host defenses and cause disease. Returning to the concepts of population
genetics and selection, it is understandable how single R-genes held by the host can be
quickly overcome. Suppose p represents a pathogen populations allele frequency for an
Avr-gene that is recognized by a host plant R-gene, while q represents the allele
frequency of a new recessive allele formed due to mutation:
A1 A1 : P11 = 0.99 p = 0.99 + 0.005 = 0.995
A1 A2 : P12 = 0.01
A2 A2 : P22 = 0.00 q = 0.00 + 0.005 = 0.005
and the selection coefficients for each genotype are:
s11 = 0.90
s12 = 0.70 .
s22 = 0.00
Following the equation for allele frequencies after selection:
q 1 ( ps12 + qs22 )
q1 =
,
1 ( p 2 s11 + 2 pqs12 + q 2 s22 )
one can see that the rare mutant allele becomes predominant in just a small number of
generations:

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A multiline or refuge will reduce the selection intensity against the A1A1 and A1A2
genotypes by providing an acceptable host for the pathogen; and maintaining (for an
extended period) a higher frequency of the Avr-gene, recognizable by the wheat cultivars
R-gene, within the pathogen population. Suppose 10 to 20 % of the wheat field contains
susceptible cultivars to the pathogen A1A1 and A1A2 genotypes; the selection intensities
are decreased and the number of generations necessary for the new allele to become
predominant increases dramatically:

The multiline strategy requires that some level of disease is acceptable and that
the pathogen reproduces by sexual means. Also, multilines may not hold a necessary
uniformity that many cropping systems require.
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A second option for the plant breeder in generating a cultivar with durable
resistance is know as gene pyramiding. In this strategy several R-genes are deployed in
the same cultivar (McDowell and Woffenden 2003; Pink 2002). In theory, pyramiding
several undefeated R-genes into a single cultivar will provide a more durable resistance
as several mutations would need to take place, one at each of the pathogens
corresponding Avr-loci. With modern molecular biology techniques (See the module:
Biotechnology and Wheat Breeding) it is possible to use markers and probes to track the
introgression of several R-genes into a single cultivar from various sources during a
crossing program.
Although many disease resistances often follows the gene-for-gene model some
follow a race-non-specific, or qualitative mode of resistance (also known as horizontal
resistance) where resistance is controlled by genes with minor to intermediate and
additive effects. Combinations of 3 to 5 of these minor genes can result in a high level of
resistance (Singh et al. 2000).
A third type of resistance is known as partial resistance and most commonly is
associated with slow rusting cultivars. Slow rusting as defined by Caldwell (1968) is a
type of resistance where disease progresses at a retarded rate, resulting in intermediate to
low disease levels against all pathotypes of a pathogen. Partial resistance is a form of
incomplete resistance characterized by a reduced rate of epidemic development despite a
high susceptible infection type. The components that cause slow rusting of a cultivar are
longer latent period, low receptivity or infection frequency, as well as smaller uredial size
and reduced duration and quantity of spore production. All of which can affect disease
progress in the field. Slow rusting resistance has dominated in CIMMYTs bread wheat
improvement program for more than 25 years.
Wheat cultivars susceptible to a disease may in fact show no significant yield
reduction (Zuckerman et al. 2006). Protection of yield, or tolerance, in infected plants
was defined by Caldwell and Shafer (1958) as the ability of a crop to endure severe
epidemics by the pathogen while sustaining only insignificant yield losses as compare
with an infected non-tolerant cultivar. The tolerance that some cultivars demonstrate
over others is not yet well understood, but it has been proposed that storage of
carbohydrates and their mobilization to the sink under stress conditions; or that tolerant
plants have the ability to compensate for the loss of photosynthetic leaf area by increasing
the photosynthetic capacity of the unaffected leaf area contribute to the tolerance to
disease (Zuckerman et al. 1996).

IV. Breeding for Resistance to Rusts By Ravi Singh


The three rust diseases, stem (or black), leaf (or brown) and stripe (or yellow) are
caused by fungi Puccinia graminis f. sp. tritici, P. triticina and P. striiformis f. sp. tritici,
continue to cause losses, often major, in various parts of the world and hence receive
much higher attention in breeding. The wheat crop can be protected from rust, or at least
the occurrence of epidemics can be reduced, by emphasizing the following three
strategies: 1) regional cooperation in monitoring the evolution and migration of new races
of rust fungi, 2) enhanced information on the genetic basis of resistance in important
wheat cultivars for their deployment, and 3) shift towards breeding and deploying wheat
cultivars with durable resistance.

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The South American cultivar Frontana is considered the best-known source of


durable resistance to leaf rust (Roelfs 1988). The Mexican-Rockefeller Program first
used this variety in the 1950s. Later its derivatives, such as Penjamo 62, Torim 73,
and Kalyan/Bluebird, showed slow rusting characteristics possibly derived from
Frontana. Genetic analysis of Frontana and several CIMMYT wheats possessing
excellent slow rusting resistance to leaf rust worldwide has indicated that such adult plant
resistance is based on the additive interaction of Lr34 and two or three additional slowrusting genes (Singh and Rajaram 1992). Leaf rust severity observed in Mexico on most
slow-rusting cultivars could be related to the number of minor genes they carry. When
susceptible cultivars display 100% leaf rust severity, cultivars with only Lr34 display
approximately 40% severity; cultivars with Lr34 and one or two additional minor genes
display 10-15% severity; and cultivars with Lr34 and two or three additional genes
display 1-5% severity. Leaf rust may increase to unacceptable levels on cultivars
carrying only Lr34, or Lr34 and one or two additional genes. However, cultivars with
Lr34 and two or three additional genes, referred to as near immune (Singh et al. 2000)
showed a stable response in all environments tested so far, with final leaf rust ratings
lower than 10% (Navabi et al. 2003). The presence of Lr34 can be indicated by the
presence of leaf tip necrosis in adult plants, which is closely linked with it (Singh 1992a).
Slow-rust resistance to leaf rust is common in spring wheat germplasm and at least 10-12
slow-rusting genes are involved in the adult plant resistance of CIMMYT wheat
germplasm (Singh and Rajaram 2002). Lines where Lr34 is absent but still possess high
level of slow-rusting resistance are also identified indicating that durable resistance is
feasible even in the absence of Lr34. The second slow-rusting resistance gene Lr46 was
recently identified in wheat cultivar Pavon 76 and located in the short arm of
chromosome 1BL (Singh et al. 1998; William et al. 2003b).
Singh (1992b) and McIntosh (1992) have indicated that the moderate level of
durable adult-plant resistance to stripe rust (caused by Puccinia striiformis) of the
CIMMYT-derived U.S. wheat cultivar Anza and winter wheats, such as Bezostaja, is
controlled in part by the Yr18 gene. This gene is completely linked to the Lr34 gene. The
level of resistance it confers by itself is usually not adequate. However, combinations of
Yr18 and 3-4 additional slow-rusting genes result in adequate resistance levels in most
environments (Singh and Rajaram 1994). Genes Lr34 and Yr18 occur frequently in
germplasm developed at CIMMYT and in various countries. Recently identified slowrusting gene Yr29 is completely linked to slow leaf rusting gene Lr46 (William et al.
2003b).
Low disease severity to stripe rust is usually associated with at least some
reduction in infection type because stripe rust grows systematically in leaf tissues. This
phenomenon results in chlorotic or necrotic stripes and therefore creates difficulties in
distinguishing slow rusting resistance from race-specific resistance. Durability and
acceptance of adult plant resistance can be expected if the cultivars low disease severity
is due to the additive interaction of several (4 to 5) partially effective genes (Navabi et al.
2004).
Stem or black rust, caused by Puccinia graminis tritici (Pgt), is historically known
to cause severe losses to wheat production. However, it has been controlled effectively
through the use of genetic resistance in cultivars associated with the green revolution
during the 1960s and 1970s. Over 80% of the spring wheat area in developing countries

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is currently sown to cultivars either derived directly from CIMMYT germplasm or from
CIMMYT germplasm used as parents. For more than 30 years, a major proportion of the
CIMMYT wheat germplasm and germplasm developed by other breeding programs have
remained resistant to stem rust. Resistance gene Sr31, located on rye translocation 1B.1R
contributed to high level of resistance in several wheat cultivars developed worldwide in
recent years. Consequently stem rust disease is often not considered important and in
many countries wheat breeding is currently done in the absence of stem rust and research
in this area in the last two decades also declined substantially.
Detection in 1999 of Pgt race Ug99 in Uganda with broad virulence, including the
virulence for Sr31, and its migration to Kenya and Ethiopia has been recognized as a
highly significant event and led to the launch of Global Rust Initiative during 2005. All
major cultivars currently grown in North Africa, the Middle East and Asia are moderately
or highly susceptible to this race. Predominant wind patterns or human errors are likely
to introduce this race to above regions and beyond. One of the major challenge to wheat
breeding is identifying or developing and diffusing adapted resistant cultivars before this
migration occurs. Although some race-specific resistance genes, mostly of alien origin,
viz., Sr22, 24, 25, 26, 27, 29, 32, 33, 35, 36, 39, 40, 44, R (1A.1R translocation) and
Tmp, can provide effective control; not all can be used in developing cultivars as some of
these alien translocations are associated with negative effects on grain yield or quality.
Shortening of these alien segments could help their utilization. Improved wheat
germplasm that carry Sr24, Sr25, Sr26, SrTmp and SrR genes is already identified and
can be used in breeding. Gene Sr24 currently occurs with a relatively high frequency in
wheat cultivars and has become ineffective in India and South Africa soon after the
release of cultivars that carried this resistance gene. The best strategy therefore is to
reconstitute durable adult-plant resistance that once protected the green revolution and
subsequent wheat cultivars. If race-specific genes need to be used, they must be
deployed in combination to enhance their longevity.
Durable stem-rust resistance of some older US, Australian and CIMMYT spring
wheats is believed to be due to the deployment of Sr2 in conjunction with other unknown
minor, additive genes. McFadden transferred Gene Sr2 to hexaploid wheat in the 1920s
from tetraploid emmer wheat cultivar Yaroslav. The slow rusting gene Sr2 confers by
itself only moderate levels of resistance. Its presence can be detected through its complete
linkage with the pseudo-black chaff phenotype. A large number of wheat lines were
evaluated during 2005 in Kenya under the stem rust epidemic caused by Ug99.
Genotypes with pseudo-black chaff phenotype showed varying degrees of disease
severity with a maximum severity reaching to about 60% compared with 100% severity
for highly susceptible materials. Moreover, the host reaction for these genotypes on the
same internode varied from moderately resistant to susceptible. These observations
clearly indicated that although slow rusting resistance gene Sr2 continues to confer at
least some resistance, the level of resistance was not sufficient when this gene is present
alone under high disease pressure in Kenya. Sr2 was detected in several highly resistant
old, tall Kenyan cultivars, including Kenya Plume (Singh and McIntosh 1986), and
CIMMYT-derived semidwarf cultivar Pavon 76. These cultivars have shown a
maximum disease score of 15MR (15% disease severity with moderately resistant
reaction). Because Pavon 76 is susceptible to race Ug99 at the seedling stage, its
resistance, as speculated earlier (Rajaram et al. 1988), is based on multiple additive genes

- 17 -

where Sr2 is an important component. Wide testing of improved wheat germplasm also
has helped in identifying additional sources of adult-plant resistance. These sources are
being used at CIMMYT to incorporate durable stem-rust resistance into high yielding,
widely adapted wheat cultivars using the methodology described below.
Breeding for durable resistance based on minor additive genes has been
challenging and often slow, for several reasons: 1) a sufficient number of minor genes
may not be present in a single source genotype, 2) a source genotype may be poorly
adapted, 3) there may be confounding effects from the segregation of both major and
minor genes in the population, 4) crossing and selection schemes and population sizes are
more suitable for selecting major genes, 5) reliable molecular markers for several minor
genes are unavailable, and 6) the cost associated with identifying and utilizing multiple
markers is high. One suggested approach is to use recurrent selection schemes to
accumulate several minor genes in a single genetic background. Such selection schemes
have often been more of a scientific interest than actually being applied in breeding.
Selection for resistance alone will not generate important popular cultivars, unless it is
simultaneously combined with other traits, such as high yield and quality. However, such
germplasm carrying combinations of minor genes should be very useful in transferring
these genes to modern cultivars.
A successful example of breeding for resistance based on minor genes is the
resistance to leaf and stripe rusts in wheat, which took about 30 years of continuous effort
at CIMMYT. In the early 1970s, S. Rajaram, influenced by the concept of slow-rusting
resistance in wheat proposed by R. Caldwell and partial resistance to late blight of potato
put forth by J. Niederhauser, made a strategic decision: to initiate selection for slowrusting resistance to leaf rust in CIMMYT spring wheat germplasm. In the early phase of
breeding he maintained plants and lines in segregating populations that would show 2030% rust severity with compatible infection type. This strategy led to the release of
several successful wheat cultivars, such as Pavon 76 and Nacozari 76, in Mexico and
other countries. These slow-rusting lines were used heavily in the crossing program and
resulted in the wide distribution of minor genes within CIMMYT spring wheat
germplasm.
The genetic basis of such resistance started to become clear in the early 1990s.
High-yielding lines that combine four or five additive, minor genes for both leaf and
stripe rusts and show near-immune levels of resistance were developed in the 1990s
(Singh et al. 2000). Three or four lines carrying different minor genes were crossed (3way and 4-way crosses), and plants in large segregating populations were selected under
artificially created rust epidemics. Races of pathogens that have virulence for racespecific resistance genes present in the parents were used to create the epidemics. The
resulting highly resistant lines are now being used in a planned manner to transfer these
minor resistance genes to well adapted, farmers choice cultivars that are currently
grown across large areas but have become susceptible to rust races in Mexico. Based on
genetic information on the number of additive, minor genes that must be transferred to
achieve the desired level of resistance, the crossing and selection scheme described below
was developed and applied. This strategy has allowed simultaneous transfer not only of
resistance genes but also other minor genes with small effects that increase the yield
potential or improve the grain quality of an adapted cultivar.

- 18 -

To transfer minor gene-based resistance into a susceptible adapted cultivar or any


selected genotype, we use a single backcross-selected bulk scheme (figure 10), where
the cultivar/genotype is crossed with a group of about 8-10 resistance donors; 20 spikes
of the F1 plants from each cross are then backcrossed to obtain 400-500 BC1 seeds.
Selection is practiced from the BC1 generation onwards for resistance and other
agronomic features under high rust pressure. Because additive genes are partially
dominant, BC1 plants carrying most of the genes show intermediate resistance and can be
selected visually. About 1600 plants per cross are space-grown in the F2, whereas
population sizes of about 1000 plants are maintained in the F3-F5 populations. Plants
with desirable agronomic features and low to moderate terminal disease severity in early
generations (BC1, F2 and F3), and plants with low terminal severity in later generations
(F4 and F5) are retained. We use a selected-bulk scheme where one spike from each
selected plant is harvested as bulk until the F4 generation, and plants are harvested
individually in the F5. Bulking of selected plants poses no restriction on the number of
plants that can be selected in each generation, as harvesting and threshing are quick and
inexpensive, and the next generation is derived from a sample of the bulked seed.
Because high resistance levels require the presence of 4 to 5 additive genes, the
level of homozygosity from the F4 generation onwards is usually sufficient to identify
plants that combine adequate resistance with good agronomic features. Moreover,
selecting plants with low terminal disease severity under high disease pressure means that
more additive genes may be present in those plants. Selection for seed characteristics is
carried out on seeds obtained from individually harvested F5 plants. Small plots of the
F6 lines are then evaluated for agronomic features, homozygosity of resistance, etc.,
before conducting yield trials.
Resistant derivatives of several cultivars and genotypes were recently developed
using the above methodology. In each case we could identify derived lines that not only
carry high levels of resistance to leaf rust or yellow rust or both, but also show about 515% higher yield potential than the original cultivar. We believe this approach to wheat
improvement allows us to maintain the characteristics of the original cultivar while
improving its yield potential and rust resistance. It should be noted that having minor
gene-based resistance in several backgrounds should ease future selection for these
resistance genes.

- 19 -

Fig. 10 Strategy for Breeding Durable Rust Resistance

- 20 -

V. Breeding for Resistance to Fusarium head blight


Fusarium Head Blight (FHB, Fusarium head scab) is one of the most destructive
diseases of wheat in areas with warm and humid climatic conditions. This disease poses a
grave threat to wheat and barley production and industries throughout the world since it
brings about mycotoxin contamination. The mycotoxins, such as deoxynivalenol (DON)
and nivalenol (NIV), are produced by Fusarium fungi and cause acute food poisoning in
people who eat infected wheat and barley products. The mycotoxins also affect animals
fed on diseased grains. Over the years FHB has become widespread, partly because of the
climatic changes brought about by global warming and also due to the increase in
reduced-tillage practices. Genetic and breeding studies on FHB resistance have been
carried out in Japan since the 1960s. CIMMYT also started a breeding program for FHB
resistance approximately 20 years ago.
Several species of the genus Fusarium cause fusarium head blight (FHB) or scab,
which is a main production constraint in environments where humid and semi-humid
conditions, caused by frequent rainfalls, coincide with flowering in wheat. The Yangtze
River basin of China with about 7 million ha has traditionally been known to be highly
prone to the disease epidemics. Disease incidences leading to epidemics are now frequent
in various other developing countries, e.g., Argentina, Brazil and Uruguay, where residue
retention is a common practice for conservation agriculture. Introduction of maize-wheat
crop rotation further increases the disease build-up.
Sources of resistance to FHB have been divided into three groups: China and
Japan, Argentina and Brazil and Eastern Europe (Singh and Rajaram 2002). More
recently additional sources, including some hexaploid synthetic derived wheat lines have
also been identified to carry moderate resistance. Earlier genetic analysis indicated that a
few additive genes confer resistance in Chinese and Brazilian wheats, and genes present
in Chinese sources are different from those in Brazilian sources (Singh et al. 1995, Van
Ginkel et al. 1996). Although several genomic regions are now known to contribute
quantitative resistance (Buerstsmayr et al. 2002, Anderson et al. 2001), a gene from
Chinese cultivar Sumai 3 in the short arm of chromosome 3B has shown the largest and
consistent effect in reducing disease severity and mycotoxin accumulation (Anderson et
al. 2001). The Chinese sources are probably the best resistances currently available and
must be combined with other sources of resistance. The Chinese cultivars that best
combined with CIMMYT materials to transmit scab resistance are Sumai#3, Ning
7840, Shanghai#5, Yangmai#6, Suzhoe#6, Wuhan#3 and Chuanmai 18.
Further progress in enhancing the level of resistance beyond the current level can
come from a breeding strategy that would favor the accumulation of multiple minor
genes from various sources into a single genotype. This would involve intercrossing
parents where resistance is located in different genomic regions, followed by growing
large segregating population and using flanking markers linked to resistance locus for
selection in early segregating generations. Conventional field screening must wait until
the F4 or F5 generations when homozygosity has increased significantly. Progenies that
show higher levels of resistance than the parental sources could be used for a targeted
transfer to high yielding cultivars that would already by then carry moderate levels of
resistance from the ongoing breeding efforts. This strategy is currently practiced at
CIMMYT to accumulate resistance genes from different sources.
- 21 -

In 2006 CIMMYT modified their FHB screening system for greater screening
capabilities, accuracy and precision. These changes have included the following:
Changing the primary screening site from Toluca to El Batan (headquarters),
Mexico
Implementing an automated, programmable misting system
Using precision CO2 sprayers for liquid inoculum application

Figure 11: CIMMYT Primary FHB Screening Site, El Batan, Mexico: A) Micro-sprinkler of misting
system, B) CO2 backpack sprayer used for inoculations C) Examples of symptoms observed.

Fine-Misting System
Approximately 1.9 hectares were placed under a programmable misting system to
provide uniform humidity conditions for favorable FHB conditions. This misting system
includes about 1,600 DAN modular micro-sprinklers (Figure 1A) spaced at distances of 3
x 4 meters, and operated automatically by a programmable timer to provide humid
conditions 24 hours a day. The system is programmed for misting both day and night,
with misting intervals and duration varying depending on the time of day (more frequent
misting during the driest periods of the day).
CO2 Sprayers
CO2 backpack sprayers were purchased from R&D Sprayers with an 8002VS flat
fan nozzle for precise application of inoculum (Figure 1B). Liquid inoculum was applied
at a concentration of 100,000 macroconidia/ml of a mixture of three isolates of F.
graminearum at a pressure of 40psi and a rate of 39ml of inoculum per meter. Wheat
plots were first sprayed at anthesis, and barley plots were first sprayed at heading.
Inoculum was re-applied at the same rate three days after the first inoculation. Grain
spawn inoculum (colonized maize grains) was also spread in the FHB evaluation plots to
ensure adequate levels of the pathogen for the generation of the disease.
By screening germplasm using the method above, recommendations were made to
CIMMYT breeders on germplasm for further use of these materials by their programs
and development of new FHB resistant wheat (Table 1).

- 22 -

Table 1 Accessions That Have Been Recommended for Further Use Based on
Recent (2006) FHB Trials in a Disease Nursery at CIMMYT
Cross

Selection History

SHANGHAI

-33GH-0M-0Y-0Y-0SCM-0FGR-0FGR-0FGR

GONDO/TNMU

CMSS92M01425S-015M-0Y-0Y-050M-16Y-2M-0Y-2SJ-0Y 0FGR0FGR

IVAN/6/SABUF/5/BCN/4/RABI//GS/CR
A/3/AE.SQUARROSA (190)

CMSS00M00007S-030M-1Y-4SCM-010Y-0FGR

IVAN/6/SABUF/5/BCN/4/RABI//GS/CR
A/3/AE.SQUARROSA (190)

CMSS00M00007S-030M-1Y-8SCM-010Y-0FGR

IVAN/6/SABUF/5/BCN/4/RABI//GS/CR
A/3/AE.SQUARROSA (190)

CMSS00M00007S-030M-1Y-24SCM-2Y-0FGR

GONDO/FINSI

CMSS00Y02909S-030Y-030M-030Y-2M-1M-0Y

TNMU/6/CEP80111/CEP81165/5/MRN
G/4/YKT406/3/AG/ASN//ATR

CMBW91Y01692S-13Y-2AL-3AL-010Y-3M-0Y-3PZ-0Y

MAYOOR//TK SN1081/AE.SQUARROSA
(222)/4/CS/LE.RA//CS/3/PVN/5/PRINIA

CSSS00B00011S-2Y-10M-2Y-8FGR-1Y-0FGR-0BI

80456/YANGMAI 5//SHA5/WEAVER/3/PRINIA

CMSS98M00896T-040Y-0100M-040Y-040M-030Y-53M-1Y-0M

SUM3/3/CS/LE.RA//CS/4/YANGMAI 158

-0FGR

WUH1/VEE#5//CBRD

CMSS92M01863S-015M-0Y-050M-0Y-13M-0Y-0FGR-0FGR-0FGR
CMSS97M01288S-030M-020Y-030M-015Y-29M-2Y-2M-0Y020SCM
CMSS97M01295S-040M-020Y-030M-015Y-40M-1Y-2M-0Y020SCM
CMSS97M01318S-040M-20Y-010M-010Y-5M-2Y-2M-0Y-020S
CMSS97M01333S-030M-100Y-010M-010Y-6M-1Y-2M-0Y020SCM
CMSS98M00761T-040Y-0100M-040Y-040M-030Y-15M-2Y-0M

80456/YANGMAI 5//SHA5/WEAVER
NG8675/CBRD//SHA5/WEAVER
GONDO/CBRD
BAU/MILAN//CBRD
EMB16/CBRD//CBRD
80456/YANGMAI
5/3/PF70354/BOW//DUCULA/4/DULUS

CMSS99Y03242T-040M-040Y-040M-030Y-030M-17Y-2M

YANGMAI 5*2/4/MOR/VEE#5//DUCULA/3/DUCULA

CMSS99Y03149F-040M-040Y-040M-030Y-030M-15Y-2M-2M-0Y

SHA4/CHIL/4/CAR422/ANA//TRAP#1/3/STAR

CMSS99M02404S-040M-030Y-030M-2Y-1M-2M-0Y

SHA3/SERI//G.C.W1/SERI/3/SHA3/SERI//YANG87142

CMSS00Y02958S-030Y-030M-030Y-5M-1M-0Y

NG8675/CBRD//MILAN/3/NG8675/CBRD

CMSS95M01814T-050Y-0100M-050Y-050M-040Y-030M-3Y-0M0SCM-0Y-0FGR-0FGR

NING MAI 9558

-0CHN-0SCM-0Y-0SCM-0Y-0FGR-0FGR

TINAMOU

CM81812-12Y-06PZ-5Y-5M-0Y-3AL-0Y-2M-010Y-0M-3PZ-0Y

TRAP#1/BOW//TAIGU
DERIVATIVE

CMSS94Y01180S-0300M-0100Y-050Y-050M-41Y-030M-3PZ0Y-3M-0Y-0SCM-0Y-0FGR-0FGR

TRAP#1/BOW//TAIGU
DERIVATIVE

CMSS94Y01180S-0300M-0100Y-050Y-050M-41Y-030M-1SJ0Y-2M-0Y-0SCM-0Y-0FGR-0FGR

SHA3/CBRD

-0SHG-1GH-0FGR-0FGR-0SCM-0FGR-0FGR-0FGR

SUM3/3/CS/LE.RA//CS/4/YANGMAI 158

-0FGR-0FGR-0FGR

YANGMAI 5

-0CHN-0SCM-0Y-0SCM-0FGR-0FGR-0FGR

TRAP#1/BOW//TAIGUDERIVATIVE

CMSS94Y01180S-0300M-0100Y-050Y-050M-41Y-030M-3PZ0Y-0FGR-0FGR

EMB27/KLORI

F37023-901F-904F-902F-903F-901F-453F-901F-900F

BR23/EMB27

-7TSB-0Y-0SCM-0Y-0SCM-0Y-0FGR-0FGR

- 23 -

VI. Breeding for Resistance to Septoria tritici Blotch


Septoria tritici blotch (STB), caused by Mycosphaerella graminicola (anamorph
Septoria tritici), is currently the most serious foliar disease of wheat grown under
temperate (1520C) and humid climates in Europe, South America, North Africa, and
Central Asia. Severe infections can cause up to 60% losses. Resistant cultivars reduce
the use of costly fungicide treatments some of which have become obsolete as some
pathogen isolates have become immune (Arraino and Brown 2006). The population of M.
graminicola is highly diverse genetically (McDonald and Linde 2002) and the fungus
reproduces sexually several times during the wheat growing season, which increases the
risk that the pathogen overcomes host resistance (Zadoks 2003). Likewise, this high
evolutionary rate probably explains the widespread failure of strobilurin fungicides after
most M. graminicola isolates appeared to spread the mutation G143A in a very few years
in the United Kingdom (Lucas 2003).
Useful sources of resistance include Bobwhite, Kavkaz-K4500, Corydon, Catbird,
and Milan (Singh and Rajaram 2002). Kavkaz-K4500, Veranopolis, and Catbird have
isolate-specific resistance. This suggests that several resistance genes can be pyramided
in a single cultivar (Chartrain et al. 2004; Chartrain et al. 2005). Isolate specific
resistance is near-complete, oligogenic, and follows a gene for gene relationship, whereas
quantitative or partial resistance is incomplete, polygenic, and isolate nonspecific.
Twelve genes for resistance to STB have been identified (Chartrain et al. 2004). Specific
interactions between cultivars and isolates can easily be assessed by using a detached
seedling leaf technique to study resistance in wheat (Arraiano et al. 2001).
The detached seedling leaf technique has a number of advantages over field and
glasshouse trials:
The test can be carried out on several pathogen isolates at one time.
It is conducted under controlled conditions at any time of year.
Little glasshouse space is required.
The labor required to set up the trail is offset by no labor being needed
for watering, fertilizing, or other maintenance of plants.
Results are highly correlated with results from the field.
The following protocol for the detached seedling technique is abbreviated from Arraiano
et al. (2001):
Seeds of the cultivars to be tested are pre-germinated on wet filter paper in
darkness at 25C for 24 h.
Seeds moved to the refrigerator at 5C for 48 h.
Seeds moved back to 25C for 24 h.
Germinated seeds are grown in potting soil, in a glasshouse until first or
second leaf is expanded (12-15 days following germination).
Inoculum is produced from sporulating cultures of M. graminicola, grown on
potato dextrose agar for 7 days under near-ultraviolet light for 16 h per day at
15C.
Cultures are flooded with sterile distilled water and scraped to release conidia.
The concentration of conidia suspension is adjusted to107 spores/mL and
Tween 20 is added to 0.15% v/v.

- 24 -

Wheat seedlings are evenly sprayed with spore suspension to run off.
Leaves are left to dry for 30 min.
Three 5 cm sections cut from the middle of the primary leaves.
Water agar is prepared (10g/L) containing 100mg/L benzimidazole (used to
retard senescence)
50 mL dispensed into an appropriate number of 8x12x2 cm clear polystyrene
boxes.
Rectangular sections (3x9 cm) are cut from the center of the agar.
8 to 10 Seedling leaf sections per box are laid, top surface up, across the
gap(created by cutting out of the rectangular sections) so that the cut ends rest
on the agar.
Strips of agar are laid over the cut edges of the leaves.
The boxes are closed and covered with black plastic or foil.
The boxes are incubated at 20C for 48 h in darkness.
The boxes, then, are uncovered and left under white phosphorescent light for
12 days at 20C.
Boxes are then moved to conditions with near-ultraviolet light at 15C to
promote sporulation (14 days after inoculation)
The percentage of leaf area covered by lesions bearing pycnidia is scored 4 to
5 times at intervals of 2-4 days for 19-28 days after inoculation. (Assessments
carried out under a 40x dissecting microscope).
New computer tools may lend themselves useful in scoring of diseases. Image analysis
software can be a faster and more reliable method for scoring of diseases than that which
can be done with the eye which is more subjective and can tire. For further information
on how image analysis software can help quantify disease incidence see Mirik et al.
(2006).
In field studies at CIMMYT disease is rated using a double digit scale:
After anthesis, spot blotch severity is evaluated using the double-digit scale (00-99)
developed as a modification of Saari and Prescotts scale for assessing severity of foliar
diseases of wheat (Saari and Prescott, 1975; Eyal at al., 1987). The first digit (D1)
indicates disease progress in canopy height from the ground level, while the second digit
(D2) refers to severity measured based on diseased leaf area. Both D1 and D2 are scored
on a scale of 1 to 9. Since the rate of disease progress in the field can be extremely high
in some regions, it is often needed to take repeated scores to properly assess the level of
resistance (Dubin et al., 1998; Duveiller et al., 1998). It is recommended to take several
individual disease scores per plot at 3- to 7-day intervals over a 3- to 4-week period
between anthesis and dough stage depending on seeding date. For each score, percent
disease severity is estimated based on the following formula:
% severity = ((D1/9) x (D2/9) x 100)
The area under the disease progress curve (AUDPC) is calculated using the
percent severity estimates corresponding to the three to four ratings shown below:
,

- 25 -

where, xi = severity on the ith date, ti = ith day, and n = number of dates on which disease
is recorded. The AUDPC (%-day) measures, the amount of disease as well as the rate of
progress.
It may be appropriate to standardize the AUDPC to take into account different
epidemics situations and planting dates. In this case AUDPC should be divided by the
total number of days in the evaluation period (AUDPC/day) to better compare genotypes
or among epidemics.
A good example of how to identify sources of race-specific resistance is
underway at the CIMMYT station in Toluca. Ten nurseries, each containing the same 30
wheat cultivars replicated twice in each nursery have been planted. Each nursery has been
inoculated with a different strain of Mycosphaerella graminicola, and disease has been
scored as described above four times over the season. By doing this, the breeder can
identify which cultivars hold R-genes for specific pathogen isolates. Cultivars holding
resistance to different races of the pathogen can then be put into a double cross scheme in
an attempt to introgress a number of R-genes into a single cultivar.
A number of cultivars that can be used as a source for resistance are listed in table
2 taken from a study done by Chartrain et al. (2004).

VII. Breeding for Resistance to Karnal Bunt


Karnal (Tilletia indica) bunt or partial bunt is the most recently described smut
disease of wheat, being first reported around Karnal, India in 1931. Since that time it has
been reported in several countries including India, Nepal, Pakistan, Afghanistan, Iran,
Iraq, South Africa, Mexico, and U.S.A. Although high disease severity does not cause a
significant loss in grain yield, a crop containing just 1-4% of infected kernels is sufficient
to render wheat grain unpalatable; and disease incidence of 5% causes distinct
deterioration in flour quality.
Since the 1940s wheat cultivars have been reported to be resistant to Karnal Bunt
under field conditions in India. Resistant cultivars have also been reported in to originate
in China, Brazil and from synthetic varieties derived by T. turgidum x A. tauschii
(Fuentes- Davila et al. 1994; Multani et al, 1988; Villareal et al. 1996). Studies on the
mode of inheritance and allelic relationship among genes conferring Karnal bunt
resistance in wheat have identified two partially recessive and four partially dominant
genes (Fuentes-Davila et al. 1995). Other studies suggest resistance is oligogenic and
additive in nature; requiring two or three genes for resistance and as many as nine genes
available from a variety of sources (Sharma et al. 2005; Harjit-Singh et al.1999).
Inoculation of resistant lines with different Karnal bunt isolates reveal variable levels of
disease incidence; suggesting that available resistance to Karnal bunt is race-specific
which may require R-gene pyramiding for durable resistance (Datta et al. 1999). As
sufficient resistance appears to rely on just a few genes, a backcross strategy as the one
described above for the rusts would be an effective breeding method in developing a
locally adapted and resistant cultivar

- 26 -

Table 2 Sources of Resistance to STB and Percentage of Leaf Area Covered by Lesions Bearing Pycnidia of 12 Different
Mycosphaerella graminicola isolates
Wheat line

Origin of line

CA30
USA

IPO001
NLANDS

IPO323
NLANDS

IPO87019
URUGUAY

IPO88004
ETHIOPIA

IPO89011
NLANDS

IPO90004
MEXICO

IPO90012
MEXICO

IPO92006
PORTUGAL

IPO94269
NLANDS

ISR398
ISRAEL

ISR8036
ISRAEL

SENAT
GENE
TE9111
MILAN
ISRAEL493
CHAUCER
EQUINOX
ARINA
TONIC
MENTANA
FLAME
VERNOPOLIS
REAPER
OLAF
RIBAND
LONGBOW
BULGARIA88
FRONTANA
CATBIRD
BALDUS
SHAFIR
CHINESE
SPRING
KK (L6.A.4)
COURTOT
Isolate mean

Denmark
USA
Potugal
CIMMYT
Israel
UK
UK
Switzerland
UK
Italy
UK
Brazil
UK
USA
UK
UK
Bulgaria
Brazil
CIMMYT
Netherlands
Israel

1
6
17
9
6
18
3
45
25
20
25
7
16
29
49
23
21
35
3/
71
18

10
8
10
48
24
7
18
37
24
29
59
3
45
44
34
50
38
4
51
64

0
0
0
0
7
71
65
0
63
39
3
0
64
1
71
57
5
53
1
30
2

1
20
0
10
21
1
17
37
31
49
37
9
22
30
34
27
51
39
3
44
66

0
53
4
77
0
0
22
7
8
42
30
0
54
3
0
0
44
71
13
2
95

6
1
33
1
12
23
36
8
6
5
59
58
41
47
38
38
37
16
43
63
55

17
20
50
12
7
27
24
8
23
38
33
3
36
0
30
42
36
37
81
54
58

7
37
1
19
6
6
28
82
84
47
66
3
48
1
56
72
57
67
89
78
87

3
39
17
25
44
5
61
78
59
44
64
53
69
52
59
54
65
69
80
66
59

25
0
12
9
16
30
30
44
24
3
27
17
34
15
32
35
28
5
55
55
24

6
10
0
34
6
30
28
19
14
29
15
18
41
18
24
42
18
35
0
26
44

2
0
1
29
30
2
10
6
9
41
2
29
9
24
13
24
37
17
49
9
46

China

38

44

15

96

55

90

99

57

42

16

39

CIMMYT
France

68
59
20

10
88
33

1
81
55

0
34
26

78
76
44

35
4
26

78
89
39

93
100
64

79
79
53

13
63
29

0
66
24

4
73
14

Grey shaded figures indicate significance for specific interactions of wheat line with M. graminicola isolates Wheat lines in bold are those with best resistance.

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It has been reported that selection for resistance to Karnal bunt is exceptionally difficult
as environmental conditions play a significant role in this pathogens virulence and
nurseries are both expensive and difficult to maintain (Datta et al. 1999; SukhwinderSingh et al. 2003).

VIII. Nematodes
It has been estimated that plant-parasitic nematodes account for 10% of
worldwide crop loss. Cereal Cyst Nematodes are a complex group of 12 described and
several undescribed species with Heterodera avenae being the most economically
important. Yield losses from H. avenae vary from 15-20% in Pakistan to 40-92% in
Saudi Arabia. The damage threshold is dependent on a number of variables including
wheat cultivar, soil type, the pathotype and ecotype of the nematode, and climactic
conditions of the geographical region. Potential damage from nematodes is increased
where wheat is grown in stressful environments such as poor soil nutrition, temperature
stress, water stress, or where there is pressure from other pathogens (Nicol et al. 2003). .
Identification of Cereal Cyst and Lesion nematodes is difficult, and has
historically been accomplished through comparative morphology. A key to the
identification of nematodes can be found at: http://nematode.unl.edu/nemakey.htm and
may be useful in identifying a specific nematode pest. More recently laboratory
techniques including AFLP markers have been developed to identify and discriminate
both cyst and lesion nematodes (Andrs et al. 2001; Mokabli et al. 2001; Orui and
Mizukubo 1999; Curran 2002).
A handful of Cyst resistance genes (Cre) have been identified in wheat and have
been deployed to provide effective resistance in parts of the world. In Europe and North
Africa, Cre1, provides good resistance against H. avenae; but has proven ineffective
against those races specific to Asiatic and Australian wheat growing regions. Greater
regional usability is offered by Cre2 and Cre4 which come from Aegilops spp. Table 2
contains a number of sources that could be used to introgress nematode resistance genes
into a breeding population. Host resistance to nematodes is still poorly understood and
collaboration between research institutions like CIMMYT and country programs will be
necessary to improve and develop more reliable host resistance.
Table 3 suggests that both horizontal and vertical resistance to nematodes is
available in existing germplasm. If nematode resistance is a part of a programs breeding
objectives, cultivars with proven resistance should be chosen as a donor parent in a
backcross strategy with a locally adapted recurrent parent. Stated earlier, nematode
pressure depends on many variables and successfully screening and selecting for
cultivars holding resistance genes may be difficult as nematode pressure will vary from
site to site and from year to year. Establishing a disease nursery for nematode resistance
selection would be helpful. In order to keep nematode populations at a maximum an
understanding of the problem nematodes preferred environment is necessary. By
maintaining an optimal soil ecosystem for the nematodes, and offering them susceptible
hosts (also useful as checks) season after season in the same location; a large population
of nematodes will be maintained, offering better disease pressure and an opportunity for
more effective selection.

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Table 3. Principal sources of genes used for breeding resistance to Heterodera avenae in cerealsb
Cereal species

Cultivar or line

Origin

Remarksc

Genetic information

Used

Wheat
Triticum aestivum

Loros, AUS 10894

-, Australia

1 dominant gene, Cre1 (formerly Ccn1) S, India; pR to several pathotypes


on chromosome 2BL

NW. Europe, Australia

Katyil

Australia

Ccn1

S, India

Australia

CreF on chromosome 7L?

pR in cv Molineux

Australia

R, to several cereal cyst pathotypes Australia, France, CIMMYT (under


and species and Pratylenchus thomei evaluation)

Festiguay
AUS4930
48'
T. durum

Australia
=

'Iraq Iraq

Psathias

S, to some pathotypes

also pR
7654,
Sansome,

7655, -

S, to some pathotypes

Khapli

France

also pR

Wild grass relatives


Aegilops tauschii
(T. tauschii)

CPI 110813

Central Asia

On chromosome 2DL, Cre4

Ae. tauschii
(T. tauschii)

AUS 18913

1 dominant gene on chromosome 2DL, R, Australia (Ha13) and several other Australia advanced breeding lines
Cre3
countries

T. variabilie

West Asia

Gene Rkn-mn1 on chromosome 3U or R, to various pathotypes and France, Algeria, Spain, India, Syria
Meloidogyne naasi and H. latipons
3Sv

T. longissimum

18

R and pR to several pathotypes

France (under evaluation)

T. ovatum

79

Mediterranean
basin

R and pR to several pathotypes

France (under evaluation)

T. triunciale
(Ae. triuncialis)

TR-353

Spain

1 dominant
CreAet)

T. geniculata
(Ae. geniculata)

Spain, Bulgaria,

T. ventricosum
(Ae. ventricosa)

VPM 1

Jordan, Tunisia

On chromosome 2AS, Cre5 (formerly populations and H. latipons R, to France, Australia (under evaluation)
CreX)
French pathotype (Ha12)

T. ventricosum
(Ae. ventricosa)

11, AP-1, H-93-8

Mediterranean
basin

On genome Nv, Cre2

T. ventricosum
(Ae. ventricosa)

11, AP-1, H-93-8, H- Mediterranean


93-35
basin

gene,

Cre7

R, Australia (Ha 13) and several other Australia synthetic hexaploid lines
countries

(formerly R, to several pathotypes (French, Spain (under evaluation)


Swedish, Spanish)
R, to several H. avenae

France, CIMMYT (under evaluation)

R, to Spanish, French and UK (Ha11) Spain (under evaluation)


pathotypes

1 dominant gene on chromosome 5Nv, R, to Australian pathotype (Ha13), not Spain, Australia (under evaluation)
Cre6
effective against Spanish (Ha71)

b Information unavailable from reference = -; no published scientific studies conducted =?


c R = resistant; pR = partially resistant; S = susceptible

- 29 -

Questions to Test Understanding:


1. Describe symptoms of bacterial diseases.
2. What are R-genes and Avr-genes?
3. How is a plant defense mechanism triggered and how does it operate?
4. What strategies do breeders use to increase the durability of their resistant
cultivars?
5. What strategy is CIMMYT using to develop Ug99 resistant cultivars?
6. What are the challenges for selecting for resistance to Septoria Tritici
Blotch (STB)?

- 30 -

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