Selection of Host Differentials for Elucidating Pathogenic Variation in Magnaporthe grisea Populations Adapted to Finger Millet (Eleusine coracana).

Blast, caused by Pyricularia grisea (teleomorph: Magnaporthe grisea), is the most devastating disease of finger millet affecting production, utilization, and trade in Africa and Southeast Asia. An attempt was made to select a set of putative host differentials that can be used to determine virulence diversity in finger-millet-infecting populations of M. grisea. Thus, a differential set comprising eight germplasm accessions selected from finger millet core collection (IE 2911, IE 2957, IE 3392, IE 4497, IE 5091, IE 6240, IE 6337, and IE 7079) and a resistant ('GPU 28') and a susceptible ('VR 708') variety was developed. This differential set was used to study pathogenic variation in 25 isolates of M. grisea collected from Karnataka, Telangana, and Andhra Pradesh states in India. Based on the reaction (virulent = score ≥4 and avirulent = score ≤3 on a 1-to-9 scale) on host differentials, nine pathotypes were identified among 25 M. grisea isolates. Pathotype 9, represented by isolate Pg23 from Vizianagaram, was the most virulent because it could infect all of the host differentials except GPU 28. This study will be helpful in devising strategies for monitoring virulence change in M. grisea populations, and for identification of blast resistance in finger millet for use in disease resistance breeding programs.

Identification of Blast Resistance in a Core Collection of Foxtail Millet Germplasm.

Blast, also known as leaf spot, caused by Pyricularia grisea (teleomorph: Magnaporthe grisea), is a serious disease affecting both forage and grain production in foxtail millet in India. For the identification of new and diverse sources of blast resistance, a foxtail millet core collection comprising 155 accessions was evaluated against the Patancheru isolate (Fx 57) of M. grisea. In a field screen during 2009 and 2010, 21 accessions were identified with neck and head blast resistance against Fx 57. In a greenhouse screen, 11 of the 155 accessions exhibited seedling leaf blast resistance to the same isolate. Further evaluation of the selected 28 accessions (found resistant to neck and head blast under field conditions during 2009 and 2010 or leaf blast in the greenhouse screen) against four M. grisea isolates (Fx 57, Fx 58, Fx 60, and Fx 62 from Patancheru, Nandyal, Vizianagaram, and Mandya, respectively) led to the identification of 16 accessions with leaf, sheath, neck, and head blast resistance to at least one isolate. Two accessions (ISe 1181 and ISe 1547) were free from head blast infection and showed resistance to leaf (score ≤3.0 on a 1-to-9 scale), neck, and sheath blast (score ≤2.0 on a 1-to-5 scale) against all four isolates. In addition, ISe 1067 and ISe 1575 also exhibited high levels of blast resistance. Blast-resistant accessions with superior agronomic and nutritional quality traits can be evaluated in multilocation yield trials before releasing them for cultivation to farmers.

Pathogenic Variation in the Pearl Millet Blast Pathogen Magnaporthe grisea and Identification of Resistance to Diverse Pathotypes.

Blast, also known as leaf spot, caused by Pyricularia grisea (teleomorph: Magnaporthe grisea), has emerged as a serious disease affecting both forage and grain production in pearl millet in India. Pathogenic variation was studied in a greenhouse using 25 M. grisea isolates collected from four major pearl-millet-growing states in India (Rajasthan, Haryana, Maharashtra, and Uttar Pradesh) on 10 pearl millet genotypes (ICMB 02444, ICMB 02777, ICMB 06444, ICMB 93333, ICMB 96666, ICMB 97222, ICMB 99444, 863B, ICMR 06222, and ICMB 95444). Differential reactions to the test isolates were recorded on ICMB 02444, ICMB 93333, ICMB 97222, 863B, and ICMR 06222. The 25 isolates were grouped into five different pathotypes based on their reaction types (virulent = score ≥ 4 and avirulent = score ≤ 3 on a 1-to-9 scale). For the identification of resistance sources, a pearl millet mini-core comprising 238 accessions was evaluated under greenhouse conditions against five M. grisea isolates (Pg118, Pg119, Pg56, Pg53, and Pg45) representing the five pathotypes. Of 238 accessions, 32 were found to be resistant to at least one pathotype. Resistance to multiple pathotypes (two or more) was recorded in several accessions, while three accessions (IP 7846, IP 11036, and IP 21187) exhibited resistance to four of the five pathotypes. Four early-flowering (≤50 days) blast-resistant mini-core accessions (IP 7846, IP 4291, IP 15256, and IP 22449) and four accessions (IP 5964, IP 11010, IP 13636, and IP 20577) having high scores (≥7) for grain and green fodder yield potential and overall plant aspect were found to be promising for utilization in pearl millet improvement programs. Identification of five pathotypes of M. grisea and sources of resistance to these pathotypes will provide a foundation for breeding for blast resistance in pearl millet in India.

Resistance to Foliar Diseases in a Mini-Core Collection of Sorghum Germplasm.

Anthracnose, leaf blight, and rust are important biotic constraints to grain and forage sorghum production worldwide and are best managed through host plant resistance. A sorghum mini-core collection, consisting of 242 germplasm accessions developed from a core collection of 2,246 landrace accessions originating from 58 countries, was evaluated to identify sources of resistance to foliar diseases. The mini-core accessions were evaluated in anthracnose- and leaf-blight-screening nurseries under artificial inoculation in the rainy and late rainy seasons, respectively, during 2009 and 2010. For rust resistance, screening was done under artificial inoculation in the greenhouse as well as in the field under natural infection. In all, 13 accessions were found resistant (score ≤3.0 on a 1-to-9 scale) to anthracnose and 27 to leaf blight in both 2009 and 2010. Six accessions exhibited resistance to rust in both the greenhouse and the field. In the resistant accessions, a wide range of diversity was observed for agronomic traits such as days to 50% flowering, plant height, and grain yield/plant, and morphological characteristics such as grain or glume color, glume coverage, endosperm texture, and panicle type (ear head compactness). Three mini-core accessions (IS 473, IS 23684, and IS 23521) exhibited resistance to all three diseases. These accessions with multiple disease resistance will be useful in sorghum disease resistance breeding programs.

Resistance to Grain Mold and Downy Mildew in a Mini-Core Collection of Sorghum Germplasm.

Grain mold and downy mildew are important biotic constraints to grain sorghum (Sorghum bicolor) production worldwide and are best managed through host plant resistance. A sorghum mini-core collection composed of 242 germplasm accessions developed from a core collection of 2,246 landrace accessions from 58 countries was evaluated to identify sources of grain mold and downy mildew resistance. Of the 242 accessions, 140 that flowered during the rainy season (the other 102 accessions were photoperiod sensitive) were screened for grain mold resistance in a grain mold nursery under field epiphytotic conditions during 2007 and 2008. All 242 accessions were screened for downy mildew in the greenhouse using a sandwich inoculation technique. Fifty accessions were resistant to grain mold (≤10% mean severity). These resistant accessions represented four basic and six intermediate races of sorghum that originated from 21 countries and exhibited considerable diversity for agronomic and morphological traits. Downy mildew resistance (mean incidence ≤10%) was observed only in six (IS 28747, IS 31714, IS 23992, IS 27697, IS 28449, and IS 30400) of the 242 accessions. One accession, IS 23992, exhibited resistance to both the diseases. The morphologically and agronomically diverse accessions that are resistant to grain mold or downy mildew should be useful to sorghum disease resistance breeding programs.

Fertility and Mating Type Frequency in Indian Isolates of Sclerospora graminicola, the Downy Mildew Pathogen of Pearl Millet.

Sclerospora graminicola, the downy mildew pathogen of pearl millet, is an oomycetous obligate parasite which reproduces by both sexual and asexual means. Fertility and mating type frequencies were studied in 70 single-zoospore isolates (SZIs) obtained from seven representative oo-sporic isolates (Sg 021, Sg 048, Sg 110, Sg 139, Sg 149, Sg 152, and Sg 153) of S. graminicola collected from major pearl millet-growing states of India. Of the 70 SZIs tested for fertility according to oospore production potential, 62 were self-sterile and 8 were self-fertile, indicating the low occurrence of homothallism in the S. graminicola populations. The sexual mating type test of the 70 SZIs, conducted by pairing each isolate with the two standard mating type tester isolates PT2 (Mat A) and PT 3 (Mat B), revealed 28 (40.0%) isolates of Mat A, 33 (47.14%) of Mat B, 8 (11.43%) of both Mat A and Mat B, and 1 (1.43%) as unknown. The frequencies of Mat A and Mat B were in approximately equal proportions among the isolates tested, except in three parental isolates. Implications of these results in understanding the dynamic genetic structure of S. graminicola population and potential for evolution of new virulence in the pathogen are discussed.

Effects of Wetness Duration and Grain Development Stages on Sorghum Grain Mold Infection.

Grain mold caused by a complex of fungi is an economically important disease of sorghum worldwide. Little is known about the epidemiology of sorghum grain mold, which is essential for its management. Studies were conducted to quantify the effects of wetness duration on grain mold development under controlled conditions at ICRISAT. Six major sorghum grain mold fungi determined from previous field experiments, Curvularia lunata, Cladosporium oxysporum, Bipolaris australiensis, Fusarium moniliforme, F. pallidoroseum, and Phoma sorghina, were used. Panicles of a pot-grown mold-susceptible sorghum line, IS 10513, were spray inoculated with each fungus at five growth stages: flowering (F), milk (M), soft dough (S), hard dough (H), and physiological maturity (P), and were incubated in dew chambers for 0, 16, 24, 40, 48, and 72 h. Then, the plants were placed on greenhouse benches at 25 ± 1°C to allow grain mold infection to develop. Eight days after treatments, grains from F, M, and S stages were plated onto potato dextrose agar, while those from H and P stages were incubated in blotter paper humid chambers at 28 ± 1°C. Fungal colonization of grains were scored after 7 days. Results indicated a significant (P < 0.01) correlation between wetness duration and grain mold development at different stages of inoculation. Generally, with increasing wetness duration, there was an increase in grain infection by all six fungi. However, infection frequency varied among fungi and grain development stages, indicating that individual fungi might have different windows for maximum infection during the grain development stages.

Virulence Characterization of Single-Zoospore Isolates of Sclerospora graminicola from Pearl Millet.

Sclerospora graminicola, the causal agent of downy mildew in pearl millet, is well-known for variation in its virulence pattern. Nine single-zoospore isolates (Sg 026-Z-1 to Sg 026-Z-9) derived from an oosporic isolate Sg 026 from a pearl millet F1 hybrid cultivar Nath 4209 grown in a farmer's field in a village, Veelad, in Maharashtra state, India, and three controls (Sg 026, Field-1, and Field-2) were evaluated for their virulence in two experimental runs in a greenhouse. The isolates were maintained on pot-grown seedlings of a highly susceptible pearl millet line, 7042S, in a greenhouse through asexual (sporangial) generations. Pot-grown seedlings of six pearl millet potential differential lines/cultivars (7042S, NHB 3, MBH 110, ICMH 451, 843B, and 852B) were spray-inoculated with a sporangial suspension (5 × 105 sporangia ml-1) and maintained in a greenhouse at 25 ± 2°C. Data were recorded for latent period (days) and disease incidence (%), from which a virulence index (incidence × latent period-1) was calculated to quantify disease-causing potential of isolates. Results indicated significant variation in latent period, incidence, and virulence index among isolates. The isolates were classified into four distinct pathotype groups based on their virulence indices on six pearl millet lines. Because of the significant variation for virulence in the S. graminicola population infecting Nath 4209, it is recommended that the hybrid be regularly monitored for downy mildew infection in farmers' fields, and be replaced by a resistant cultivar that is genetically unrelated to the parental lines of Nath 4209. This will help delay or avoid development of downy mildew epidemics and the resulting heavy loss to pearl millet farmers in the region.

Genetic Resistance of Pearl Millet Male-Sterile Lines to Diverse Indian Pathotypes of Sclerospora graminicola.

Single-cross F1 hybrid cultivars based on cytoplasmic-nuclear male-sterility (CMS) system have contributed significantly to increasing productivity of pearl millet (Pennisetum glaucum). Genetic resistance to downy mildew (Sclerospora graminicola) in parental lines is critical for successful commercial cultivation of a hybrid cultivar. In this study, 46 genetically diverse male-sterile lines (A-lines), including 42 test A-lines, four control A-lines, a commercial hybrid, and a highly susceptible line, were evaluated in disease nurseries at four diverse locations in India and compared with pathotype isolates from the same locations under greenhouse environments. Variability in downy mildew incidence (0 to 100%) due to genetic differences among lines, among pathotypes, and that due to line × pathotype interaction were all highly significant (P < 0.001). In the field experiment, eight of the 42 test A-lines, including 841A (control), that recorded ≤10% disease incidence, were identified as resistant compared with 84 to 100% incidence on the control susceptible line 7042S. Resistance in eight of these test A-lines (863A, ICMA 88004, -94333, -98222, -98111, -92777, and -96666) and 841A was confirmed against the four pathotypes in greenhouse experiments. Cluster analysis of downy mildew incidence data from field and greenhouse experiments, using the Euclidian distance, classified the 48 lines into four distinct groups with the above eight A-lines in the resistant group. These resistant A-lines would be useful in the development of F1 hybrids with stable resistance to diverse pathotypes of downy mildew in India.

Detection of genetic variability in pearl millet downy mildew (Sclerospora graminicola) by AFLP.

Downy mildew, caused by Sclerospora graminicola, is an economically important disease of pearl millet in the semiarid regions of Asia and Africa. Amplified restriction fragment length polymorphism (AFLP) was used to detect the extent of genomic variation among 19 fungal isolates from different cultivars of pearl millet grown in various regions of India. Fourteen AFLP primer combinations produced 184 polymorphic bands. An unweighted pair-group method of averages cluster analysis represented by dendrogram and principal coordinate analysis separated the mildew collections into four distinct groups. Isolates having characteristic opposite mating abilities, geographic relatedness, virulence, common host cultivars, and changes through asexual generations reflected heterogeneity of the pathogen. The use of AFLP to detect genetic variation is particularly important in selecting mildew isolates to screen breeding material for identification of resistant millet and monitoring changes in S. graminicola in relation to changes in host for effective disease management.

DNA fingerprinting detects genetic variability in the pearl millet downy mildew pathogen (Sclerospora graminicola).

Genetic variability in six host genotype-specific pathotypes of pearl millet downy mildew pathogen S. graminicola was studied at the molecular level using mini- and micro-satellites. Our results indicated that microsatellites (GAA)6, (GACA)4, and especially (GATA)4 were quite informative and showed high levels of polymorphism among the pathotypes. The six pathotypes could be classified into five groups based on the cluster analysis of their genetic similarities, thereby confirming the existence of distinct host genotype-specific virulence in S. graminicola pathotypes. We demonstrate, for the first time, the use of DNA fingerprinting to detect genetic variation in downy mildew fungus of pearl millet.