Concept note-DBT/10Jul09

Name of the Institute/University/Organisation submitting the Project Proposal:

ICRISAT (International Crops Research Institute for the Semi-Arid Tropics), Patancheru,
Hyderabad, 502 324

1. Title of the concept: Combining downy mildew and blast resistance in pearl millet
hybrid parental lines through marker-assisted breeding

Scientists (PI): Rajan Sharma and T Nepolean

Collaborating Scientists: R P Thakur, C T Hash and M Butterfield

2. Introduction/preliminary proof:
Pearl millet (Pennisetum glaucum (L.) R. Br.) is a staple cereal grown on about 29
million ha in the arid- and semi-arid tropical regions of Africa, Asia and Latin America
with India having the largest area of 9.3 million ha
(http://www.icrisat.org/PearlMillet/PearlMillet.htm). The crop is grown as a nutrient-rich
food source for humans as well as a forage/fodder crop for livestock. During the past
three decades single-cross F1 hybrids based on cytoplasmic-nuclear male-sterility (CMS)
systems have contributed significantly in increasing pearl millet productivity in India.
However, diseases such as downy mildew, blast, rust, smut and ergot account for severe
crop damage in pearl millet. Increased yield and yield stability of pearl millet, which
could be achieved by improving disease resistance of improved cultivars, would
contribute to improving standards of living for people in the harsh production regions
where pearl millet is grown.
Among several diseases that affect pearl millet, downy mildew caused by Sclersopora
graminicola (Sacc.) Schroet has been a major problem of pearl millet hybrids. The
disease is highly destructive and widespread in major pearl millet growing areas of the
world (Jeger et al., 1998; Williams, 1984). Sclerospora graminicola is an obligate
pathogenic oomycete and reproduces both asexually by producing sporangia and sexually
by means of oospores. The fungus is largely heterothallic but homothallism has also been
reported (Idris and Ball, 1984; Michelmore et al., 1982). Existence of mating types and
their frequency greatly contribute towards the development of new recombinants in the
pathogen populations (Pushpavathi et al., 2006a). These characteristics of the fungus
make it highly variable. Evolution of host-specific virulence in pearl millet downy
mildew is well documented (Thakur et al., 1992; Sastry et al., 2001; Pushpavathi et al.,
2006b). As a result of evolution of host specific virulence, resistant genotypes lose their
effective resistance within a short period and leads to the development of new
pathotypes/races in the pathogen populations (Kolmer et al., 2006).
Now blast, also known as leaf spot caused by Pyricularia grisea (Cooke) Sacc.
[teleomorph: Magnaporthe grisea (Herbert) Barr] has emerged as another serious
disease affecting both forage and grain production in pearl millet. The disease appears as
grayish, water-soaked foliar lesions that enlarge and become necrotic, resulting in
extensive chlorosis and premature drying of young leaves (Wilson et al., 1989). This
disease becomes more severe during humid weather conditions especially with dense
plant stands. Leaf blast on pearl millet has been found to be negatively correlated with
green-plot yield, dry matter yield and digestive dry matter (Wilson and Gates, 1993). In

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India, the disease was first reported from Kanpur, Uttar Pradesh (Mehta et al., 1953).
Although blast was considered a minor disease of pearl millet in India, the disease
incidence has increased alarmingly during the recent years (Lukose et al., 2007;
Anonymous, 2009) and requires identification and utilization of blast resistance in pearl
millet breeding program.

Considerable developments in biotechnology have led plant breeders to develop more

efficient selection systems to replace traditional phenotypic-pedigree-based selection
systems. Marker-assisted selection (MAS) is indirect selection process where a trait of
interest is selected not based on the trait itself but on a marker linked to it. In pearl millet
breeding program at ICRISAT, MAS has been used for transferring downy mildew
resistant QTLs into the hybrid parental lines (Hash and Witcombe 2001). Several DM
resistant lines, such as IP 18292, 7042R and 700651 have been used in developing hybrid
parental lines. A number of DM resistant QTLs effective against diverse Indian
pathotypes of S. graminicola have been mapped on the pearl millet linkage groups and
some of them have been transferred to the commercial B-lines (843B, 81B) and R-lines
(H 77/833-2, ICMP 451). Development and commercial deployment of DM resistant
version of HHB 67 is the first successful story of MAB in field crops in public domain in
India (Hash et al. 2006).

The project aims at identifying DNA marker linked to blast resistance gene(s) (mapping
population segregating for blast resistance developed at ICRISAT will be used) in pearl
millet and marker-assisted backcrossing to transfer major QTLs associated with
pathotype specific QTLs for downy mildew resistance from different donor parents to
several genetically diverse, agronomically elite, hybrid parental lines. Finally blast
resistance gene will be introgressed in elite lines carrying QTLs for downy mildew
resistance.

References cited:

Anonymous 2009. Annual Report, All India Coordinated Pearl Millet Improvement, Indian Council of
Agricultural Research. Mandore, Jodhpur.
Hash CT, Witcombe JR. 2001. Pearl millet molecular marker research. International Sorghum and Millets
Newsletter 42:8–15.
Hash CT, Sharma A, Kolesnikova-Allen MA, Singh SD, Thakur RP, Raj AGB, Ratnajirao MNV, Nijhawan
DC, Beniwal CR, Sagar P, Yadav HP, Yadav YP, Srikant, Bhatnagar SK, Khairwal IS, Howarth CJ,
Cavan GP, Gale MD, Liu C, Devos KM, Breese WA, Witcombe JR. 2006. Teamwork delivers
biotechnology products to Indian small-holder crop-livestock producers: pearl millet hybrid “HHB
67 Improved” enters seed delivery pipeline. Journal of SAT Agricultural Research 2:16–20.
(www.ejournal.icrisat.org)
Idris MO, Ball SL. 1984. Inter- and intracontinental sexual compatibility in Sclerospora graminicola. Plant
Pathol 33: 219-223.
Jeger MJ, Gilijamse E, Bock CH, Frinking HD. 1998. The epidemiology variability and control of the
downy mildews of pearl millet and sorghum, with particular reference to Africa. Plant Pathol 47:
544-569.
Kolmer JA, Long DL, Hughes ME. 2006. Physiologic specialization of Puccinia triticina on wheat in the
Unites States in 2004. Plant Dis 90: 1219-1224.
Lukose CM, Kadvani DL, Dangaria CJ. 2007. Efficacy of fungicides in controlling blast disease of pearl
millet. Indian Phytopathology 60:68-71.

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Mehta PR, Singh B, Mathur SC. 1953. A new leaf spot disease of bajra ( Pennisetum typhoides Staph and
Hubbard) caused by a species of Piricularia. Indian Phytopathology 5:140-143.
Michelmore RW, Pawar MN, Williams RJ. 1982. Heterothallism in Sclerospora graminicola.
Phytopathology 72: 1368-1372.
Pushpavathi B, Thakur RP, Chandrashekara Rao K. 2006a. Fertility and mating type frequency in Indian
isolates of Sclerospora graminicola, the downy mildew pathogen of pearl millet. Plant Dis 90: 211-
214.
Pushpavathi B, Thakur RP, Chandrashekara Rao K, Rao VP. 2006b. Characterization of Sclerospora
graminicola isolates from pearl millet for virulence and genetic diversity. Plant Pathol J 22: 28-
35.
Sastry JG, Sivaramakrishnan S, Rao VP, Thakur RP, Singru RS, Gupta VS, Ranjekar PK. 2001. Genetic
basis of host specificity in Sclerospora graminicola, the pearl millet downy mildew pathogen. Indian
Phytopathol 54: 323-328.
Thakur RP, Shetty KG, King SB. 1992. Selection for host-specific virulence in asexual populations of
Sclerospora graminicola. Plant Pathol 41: 626-632.
Williams RJ. 1984. Downy mildew of tropical cereals. Adv Plant Pathol 3:1-103.
Wilson JP, Gates RN. 1993. Forage Yield losses in hybrid pearl millet due to leaf blight caused primarily
by Pyricularia grisea. Phytopathology 83:739-743.
Wilson JP, Wells HD, Burton GW. 1989b. Inheritance of resistance to Pyricularia grisea in pearl millet
accessions from Burkino Faso and inbred Tift 85DB. Journal of Heredity 80:499-501.

3. Major objectives/Milestones

 Map QTLs for blast resistance from different genetic backgrounds

Segregating mapping population for blast resistance developed (at least two).

QTLs for blast resistance identified in various (at least two) genetic backgrounds.

 Identify the putative downy mildew resistance QTLs mapped on different linkage
groups of pearl millet in various genetic backgrounds from earlier results

Putative downy mildew resistance QTLs mapped on various pearl millet groups
in different genetic backgrounds identified.

 Use marker-assisted backcrossing to transfer major QTLs associated with

pathotype-specific downy mildew resistance from donor parents to several (ten)
genetically diverse, agronomically elite, hybrid parental lines

Several QTL introgression lines having downy mildew resistance in different

genetic backgrounds identified.

 Use marker-assisted backcrossing to transfer blast resistance to several (ten)

genetically diverse, agronomically elite, hybrid parental lines

Several QTL introgression lines having blast resistance in different genetic

backgrounds identified.

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  Combine blast resistance with QTLs associated with pathotype specific downy
mildew resistance through QTL pyramiding in common genetic backgrounds of
several genetically diverse, agronomically elite, hybrid parental line

Several QTL introgression lines having combined downy mildew and blast
resistance in different genetic backgrounds identified.

4. Expected deliverables/outcomes:

 Information on resistance gene(s)/QTLs for blast resistance mapped in various

genetic backgrounds available
 Hybrid parental lines having pathotype-specific resistance against downy mildew
available for national researchers
 Hybrid parental lines for blast resistance available for national researchers
 Hybrid parental lines having combined resistance for downy mildew and blast
available for national researchers

5. Proof of expertise/Publications

Bidinger FR, Nepolean T, Hash CT, Yadav RS, Howarth CJ. 2007. Identification of
QTLs for Grain Yield of Pearl millet [Pennisetum glaucum (L.) R. Br.] in
Environments with Variable Moisture during Grain filling. Crop Science 47:969-
980.
Hash CT, Rizvi SMH, Nepolean T et al. 2007. Replicated Success In Marker-Assisted
controlling yield and stover quality traits in pearl millet. International Sorghum and
Millets Newsletter 47:149-152.
Jones ES, Liu CJ, Gale MD, Hash CT, Witcombe JR. 1995. Mapping quantitative trait
loci for downy mildew resistance in pearl millet. Theoretical and Applied Genetics
91: 448-456.
Nepolean T, Blummel M, Bhasker Raj AG, Rajaram V, Senthilvel S, Hash CT. 2006.
QTLs controlling yield and stover quality traits in pearl millet. International
Sorghum and Millets Newsletter 47:149-152
Pushpavathi B, Thakur RP, Chandrashekara Rao K. 2006. Fertility and mating type
frequency in India isolates of Sclerospora graminicola, the downy mildew
pathogen of pearl millet. Plant Disease 90: 211-214.
Rathour R, Sharma R, Sharma V. 2006. Genetic differentiation of rice and non-rice
populations of Magnaporthe grisea from north-western Himalayas using native
protein and isozyme polymorphisms. Journal of Phytopathology 154: 641-647.
Senthilvel S, Jayashree B, Mahalakshmi V, Sathish Kumar P, Nakka S, Nepolean T,
Hash CT. 2008. Development and mapping of simple sequence repeat markers for
pearl millet from data mining of Expressed Sequence Tags. BMC Plant Biology
8:119.
Sastry JG, Ramakrishna W, Sivaramakrishnan S, Thakur RP, Gupta VS, Ranjekar PK.
1995. DNA fingerprinting detects genetic variability in the pearl millet downy
mildew pathogen (Sclerospora graminicola). Theoretical and Applied Genetics 91:
856-861.

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Sharma R and Kaushal RP. 2004. Generation and characterization of pea (Pisum
sativum) somaclones for resistance to Ascochyta blight and powdery mildew.
Indian Journal of Biotechnology 3: 400-408.
Sharma R, Rao VP, Varshney RK, Prasanth VP, Kannan S, Thakur RP. 2008
Characterization of pathogenic and molecular diversity in Sclerospora
graminicola, the causal agent of pearl millet downy mildew. Archives of
Phytopathology and Plant Protection (accepted).
Sharma R. 2007. Genetic differentiation of host limited forms of Colletotrichum
truncatum from northwestern Himalayas. Archives of Phytopathology and Plant
Protection DOI: 10.1080/03235400701543848
Thakur RP, Rai KN, Rao VP, Rao AS. 2001. Genetic resistance of pearl millet male-
sterile lines to diverse Indian Pathotypes of Sclerospora graminicola. Plant Disease
85: 621-626.
Thakur RP, Rao VPR, Wu BM, Subbarao KV, Shetty HS, Singh G, Lukose C, Panwar
MS, Sereme P, Hess DE, Gupta SC, Dattar VV, Panicker S, Pawar NB, Bhangalre
GT, Panchbhai SD. 2004. Host resistance stability to downy mildew in pearl millet
and pathogenic variability in Sclerospora graminicola. Crop Protection 23: 901-
908.

6. Minimum budget:
Rs. 1,50,00,000 (US$300,000) (for 5 years)

7. Projects in hand:
 Marker-assisted improvement of iron and zinc grain density in pearl millet,
funded by DBT.
 Development of contigouos segmental lines in pearl millet, funded by DBT.
 Integrating genomics and mapping approaches to improve pearl millet
productivity in drought prone regions of Africa and Asia, funded by BBSRC.
 Fortification of downy mildew resistance of a Proagro millet restorer parent,
PP738, funded by Proagro, India (Completed).
 Identification of grain mold resistance genes and their introgression into elite
hybrid parents of sorghum by marker-assisted selection, funded by Sehgal
Foundation Endowment Fund.

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Work plan

Year Season Activity

1 1 Genotyping of two segregating mapping population completed for blast resistance
Phenotyping two segregating population for blast reaction under greenhouse
conditions
1 Making donor parent and recurrent parent crosses for downy mildew (DM) resistance
Making donor parent and recurrent parent crosses for blast resistance
2 Identification of QTLs for blast resistance
Collection of QTL information for downy mildew resistance
2 Sowing F1 seeds of various plant x plant crosses for DM resistance
Sowing F1 seeds of various plant x plant crosses for blast resistance
2 3 Generation advancement of BC1F1 families for DM resistance
Generation advancement of BC1F1 families for blast resistance
4 Generation advancement of BC2F1 families for DM resistance
Generation advancement of BC2F1 families for blast resistance
Marker genotyping to find QTL heterozygote for DM resistance
Marker genotyping to find QTL heterozygotes for blast resistance
Greenhouse screening for DM resistance
Greenhouse screening for blast resistance
3 5 Generation advancement of BC3F1/F2 families for DM resistance
Generation advancement of BC3F1/F2 families for blast resistance
Marker genotyping to find QTL heterozygotes for DM resistance
Marker genotyping to find QTL heterozygotes for blast resistance
Greenhouse screening for DM resistance
Greenhouse screening for blast resistance
6 Screening BC3F2 families for DM resistance
Screening BC3F2 families for blast resistance
Marker genotyping to find QTL homozygotes for DM resistance
Marker genotyping to find QTL homozygotes for blast resistance
Greenhouse screening for DM resistance
Greenhouse screening for blast resistance
6 Pyramiding BC3F2 QTL homozygotes of DM and blast resistance in common genetic
backgrounds
4 7 Generating QTL pyramided F1 families for DM and blast resistance
8 Screening QTL pyramided F2 families for DM and blast resistance
Marker genotyping to find common QTL homozygotes for DM and blast resistance
Greenhouse screening for DM and blast resistance
5 9 Generation advancement of QTL pyramided F2 families for DM and blast resistance
10 Seed multiplication

8. Association with industry: none

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