Crop Science 42:1135-1138 (2002)
© 2002 Crop Science Society of America
CROP BREEDING, GENETICS & CYTOLOGY
Analysis of Resistance to Ergot in Sorghum and Potential Alternate Hosts
J. D. Reeda,
B. A. Ramundob,
L. E. Claflinb and
M. R. Tuinstra*,a
a Dep. of Agronomy, Kansas State Univ., Manhattan, KS 66506
b Dep. of Plant Pathology, Kansas State Univ., Manhattan, KS 66506
* Corresponding author (mtuinstra{at}bear.agron.ksu.edu)
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ABSTRACT
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Ergot (caused by Claviceps africana Frederickson, Mantle & de Milliano) of sorghum [Sorghum bicolor (L.) Moench] recently has become a global disease problem and is a major threat to hybrid seed production. Host-plant resistance is one option for control; however, the genetic and physiological bases for ergot resistance are poorly understood. The objective of this study was to evaluate resistance to C. africana in 18 genetically diverse sorghum lines, including cultivated landraces and wild accessions, as well as in potential alternate hosts, including grassy weeds and range grasses commonly found in sorghum-producing areas in the central Great Plains of the USA. These entries were evaluated for ergot resistance in the greenhouse following spray inoculation with conidial suspensions during flowering. The results of this analysis indicated that only Sorghum ssp. were susceptible to ergot; however, within the sorghum germplasm pool, several wild accessions were identified with resistance to ergot. Two of these resistant entries, IS14131 and IS14257, were characterized further in male-sterile (A3 cytoplasm) genetic backgrounds to evaluate the physiological basis for their resistance. Parent lines, male-sterile hybrids, and susceptible checks were evaluated for ergot resistance following spray inoculation with ergot in experiments in a winter nursery at Guayanilla, Puerto Rico, and in a greenhouse at Manhattan, KS, during the winter and spring of 2000. The expression of ergot resistance in IS14131 and IS14257 and in corresponding male-sterile hybrids suggests that these sorghums may harbor genes for resistance to ergot.
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INTRODUCTION
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SORGHUM HAS BEEN CLASSIFIED into several different diploid and tetraploid species (Doggett, 1988). On the basis of patterns of geographical distribution and phenotypic variation, each species has been classified into distinct taxonomic units that are generally denoted by race or working group classification (Murty et al., 1967). Although sorghum is currently grown in many different parts of the world, Africa represents the center of origin for this crop (Doggett, 1988). Little is known about the genetic structure of sorghum; however, DNA fingerprinting studies have generally shown a significant amount of variation among and between cultivated and wild races (Cui et al., 1995).
Ergot of sorghum recently has become a plant disease of international concern. First identified in India in 1917 and Kenya in 1924, ergot for many years was characterized as an Old World disease (Frederickson and Mantle, 1988; Frederickson et al., 1991). However, ergot was reported in 1995 in hybrid-seed production plots in the states of Sao Paulo, Minas Gerais, and Goias, Brazil (Reis, 1996). In 1996, the pathogen was reported in Australia in a number of nurseries and seed production blocks (Ryley and Henzell, 1999). By 1997, the pathogen had progressed up the South American continent to Mexico. In March of 1997, ergot was reported in the USA in regrowth sorghum in Cameron and Hidalgo counties, Texas (Isakeit et al., 1998).
Only unfertilized ovaries are susceptible to ergot (Futrell and Webster, 1965). Claviceps africana attacks sorghum ovules and causes reduced seed set, yield loss, and harvesting problems because of honeydew secreted from the infected ovaries (Bandyopadhyay et al., 1998). The swift and unexpected spread of ergot in such a short period of time has threatened hybrid-seed production areas in the USA. Given the proper conditions, this disease also could be devastating to grain production fields in Kansas, Nebraska, Oklahoma, and Texas. Although fungicides have been identified to aid in controlling the disease, the most economical and effective strategies for control appear to be pollen management and/or host-plant resistance (McLaren, 1994).
Ergot-resistant genotypes have been reported in several studies; however, most sources of resistance appear to escape the disease through efficient pollination and fertilization. Pollen-based mechanisms of resistance are not very useful in developing resistant seed-parent lines (Frederickson et al., 1994; Bandyopadhyay et al., 1998). Only one source of physiological resistance to ergot has been identified to date. IS 8525 expressed high levels of resistance to ergot and male-sterile hybrids reportedly expressed intermediate levels of resistance (Dahlberg et al., 1998).
Sorghum ergot has been endemic in Africa and India for at least the past 75 yr and probably longer (Frederickson et al., 1991). The host and pathogen may have coevolved in this part of the world. The objectives of this study were to (i) evaluate resistance to ergot in genetically diverse sorghum lines, including cultivated landraces and wild accessions, and in other potential alternate hosts, including grassy weeds and range grasses commonly found in sorghum-producing areas of the central U.S. Great Plains and (ii) confirm and evaluate the expression of resistance shown by two wild sorghums in male-sterile (A3-cytoplasm) hybrids.
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MATERIALS AND METHODS
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Screening for Ergot Resistance
Eighteen sorghum accessions, four millets, and five range grasses commonly found in sorghum-producing areas of the central Great Plains (Table 1) were evaluated for susceptibility to ergot (Ramundo et al., 1999). The sorghum accessions represent breeding lines and cultivars from the world sorghum collection maintained at the International Crop Research Institute for the Semi-Arid Tropics (Hyderabad, Andhre Pradesh, INDIA) and the USDA-ARS National Plant Germplasm System (Fort Collins, CO, USA). The millet varieties were obtained from Dr. David Andrews (University of Nebraska). The range grasses were obtained from the Manhattan Plant Materials Center USDA-NRCS (Manhattan, KS). Plants were evaluated for ergot resistance in a greenhouse at Manhattan, KS, during the winter and spring of 1998. Entries were planted in 8-L pots with two plants per pot using a 1:1:1 peat-perlite-soil potting medium. Pots were arranged in a randomized complete block design with four replications. Entries were grown at 27°C with natural lighting and an overhead misting system was used to maintain high humidity.
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Table 1. Analysis of host range for Claviceps africana among various sorghum, millet, and range grass species commonly found in the Central Great Plains.
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The C. africana strain used for inoculation was collected from ergot infected fields in Nebraska by Dr. Stan Jensen (USDA-ARS, Lincoln, NE). The U.S. isolates of C. africana were shown to be identical genetically or very similar (Pa
outovà et al., 2000; Tooley et al., 2000), and this isolate presumably represents a typical North American strain of the pathogen. The C. africana inocula were prepared as described by Tegegne et al. (1994) and Musabyimana et al. (1995). A conidial suspension was prepared by washing infected sorghum panicles containing fresh honeydew in water. The resultant suspension was filtered through two layers of cheesecloth and adjusted to 1 x 106 macroconidia mL-1 with a hemacytometer. When terminal florets began to flower, panicles were inoculated to runoff with a hand sprayer and then covered with clear plastic bags. Five days after the initial inoculation, bags were removed and florets were reinoculated. Panicles were not covered with bags after the second inoculation. After 30 d, inoculated panicles were evaluated for symptoms of ergot infection. Disease incidence ratings were based on the percentage of florets per panicle exhibiting honeydew exudate and presence of sphacelia as described by McLaren and Wehner (1992).
Evaluation of Wild Sorghum Germplasms and Male-Sterile Hybrids
The most ergot-resistant wild sorghums were crossed to A3TX430 to produce male-sterile testcross hybrids. Sufficient hybrid seed was produced for replicated experiments to analyze ergot resistance in IS14131 and IS14257 and their respective male-sterile hybrids. Male-fertile parent lines were contrasted with male-sterile hybrids and susceptible checks for ergot resistance to evaluate the role of male-fertility in expression of ergot resistance in these entries.
These entries were evaluated for ergot resistance under field conditions in a winter nursery at Guayanilla, Puerto Rico, during the spring of 2000. The parent lines IS14131 and IS14257 and their male-sterile hybrids produced in crosses with A3TX430 were included in a trial with AN123 as a male-sterile, susceptible check. The experiment was conducted using a randomized complete block design with three replications. Herbicide and fertilizer were applied to the soil and incorporated prior to planting. These treatments included 1.41 kg ha-1 S-metolachlor [2-chloro-6'-ethyl-N-(2-methoxy-1-methyl-ethyl)acet-o-toluidide], 1.79 kg ha-1 atrazine [2-chloro-2',6'-diethyl-N-(methoxymethyl) acetanilide], and 150 kg ha-1 nitrogen in the form of urea. Entries were planted on 27 Nov. 1999 in single-row plots (4.61 x 0.76 m) with 75 seeds per row. Plots were irrigated throughout the growing season as needed using drip irrigation. The C. africana strain used for inoculation was collected from ergot-infected sorghum fields in Puerto Rico. Ergot inoculum was prepared as described by Tegegne et al. (1994). A conidial suspension was prepared by washing infected sorghum panicles containing fresh honeydew in water. The resultant suspension was filtered through two layers of cheesecloth and was then diluted to approximately 1 x 106 macroconidia mL-1. At anthesis, all panicles were spray-inoculated to runoff with the conidial suspension with a hand sprayer every other day until anthesis was complete. Approximately 1 mo after inoculation, ergot incidence was recorded as percentage of florets infected per panicle on 10 plants in each plot as described above.
The expression of ergot resistance in these wild sorghums was also evaluated in a greenhouse at Manhattan, KS, during the winter and spring of 2000. The parent lines IS14131 and IS14257 and their male-sterile hybrids produced in crosses with A3TX430 were included in the trial with A3TX430 and ATX623 as susceptible checks. These entries were evaluated in 8-L pots with two plants per pot and a randomized complete block design with five replications was used. Plants were grown in a 1:1:1 peat-perlite-soil potting medium at 27°C with supplemental lighting (140 W m-2 photosynthetically active radiation). An overhead misting system was used to maintain high humidity. The C. africana strain used in this experiment was the same as used in the initial greenhouse experiments. The inoculation and disease rating protocols were identical to those previously described in the greenhouse evaluation of sorghum accessions.
Statistical Analysis
Ergot resistance was computed on a plot means basis in each experiment. Analyses of variance (ANOVA) for ergot resistance were conducted by means of the PROC GLM procedure of the SAS Statistics package (SAS, 1990). Blocks were treated as random effects, and entries were treated as fixed effects. Variation among entry means was tested by the least square difference (LSD) at the 5% level of significance.
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RESULTS
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Analysis of Ergot Resistance
Only sorghum accessions were found to be susceptible to C. africana (Table 1). Thirteen sorghum accessions representing several races of diploid (2N = 20) and tetraploid (2N = 40) species were found to be susceptible to ergot. Infection scores varied among sorghum entries with ATX623, the male-sterile check, being the most susceptible. Five wild sorghum accessions did not develop ergot. IS14257 and IS 14357 representing accessions of S. bicolor subsp. verticilliflorum (Steud.) Piper., PI 185574 and IS 14301 representing S. bicolor subsp. arundinaceum (Steud.) de Wet., and IS14131 representing S. bicolor subsp. drummondi (Steud.) de Wet. expressed high levels of resistance to ergot and exhibited no visible signs of sphacelial growth or honeydew exudation.
None of the nonsorghum entries evaluated in this test were susceptible to C. africana. On the basis of this limited sampling, these millets and native range species would not serve as alternate hosts to the pathogen in the U.S. ergot disease cycle.
Evaluation of Ergot Resistance in Wild Sorghum Accessions
Comparison of ergot resistance in wild sorghums and their male-sterile testcross hybrids with standard ergot-susceptible checks indicated significant differences among entries in Puerto Rico and the greenhouse (Table 2). In Puerto Rico, no significant differences were detected among the wild sorghums, but they had significantly lower mean ergot scores than the susceptible check. More important, ergot scores for male-sterile hybrids were also significantly lower than those of the susceptible check. The male-sterile hybrids had ergot scores similar to the resistant parent lines. Data from inoculations in the greenhouse study were similar. IS14131 exhibited a lower incidence of ergot than IS14257; however, both wild sorghum accessions had significantly lower mean ergot scores than the susceptible checks. The ergot scores for male-sterile hybrids also were significantly lower than those of the susceptible checks, but did not differ significantly from each other.
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Table 2. Variation in ergot resistance among wild sorghum accessions, male-sterile hybrids, and standard susceptible checks.
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DISCUSSION
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The analysis of host range for C. africana among crop species and range grasses commonly found in the central Great Plains indicated that all nonsorghum species tested were immune to this disease. These results are generally consistent with previous reports; however, some studies have suggested that C. africana may infect other species (Frederickson et al., 1991; Bandyopadhyay et al., 1998). Among sorghum accessions, shattercane (S. bicolor subsp. drummondi) and Johnson Grass [S. halepense subsp. halepense (L.) Pers.] were shown to be susceptible to ergot and could serve as over-wintering hosts of C. africana. Both species often grow abundantly along roadsides and in waste areas as unwanted weeds and could be problematic in the case of early-season conidial dissemination. The role of these species in mid- and late-season infection deserves further study.
Several germplasm characterization studies have indicated variation in resistance to ergot in sorghum (Tegegne et al., 1994; Frederickson et al., 1994; Musabyimana et al., 1995; Dahlberg et al., 1998). In most cases, the expression of resistance was associated with rapid pollination characteristics that allow fertilization prior to infection (Bandyopadhyay et al., 1998). Although this type of resistance would be functional in varieties and hybrids, it cannot be used in male-sterile seed production fields, which are at greatest risk to ergot infection. When these sources of genetic resistance are crossed to A3-cytoplasm male-sterile testers, the pollen-based resistance mechanism is circumvented, and these hybrids become highly susceptible to ergot. Prior to this study, IS 8525 was the only reported germplasm source of physiological resistance to ergot in male-sterile genetic backgrounds (Dahlberg et al., 1998). Male-sterile hybrids produced by means of IS 8525 were shown to be significantly more resistant to ergot than susceptible checks; however, the level of infection in these hybrids was still quite high. Given this limited genetic basis for resistance to ergot in sorghum, the identification of new sources of resistance in IS14131 and IS14257 is highly significant. On the basis of the low level of infection observed in male-sterile hybrids, particularly under field conditions, resistance exhibited in these accessions appears to be physiological and not pollen-mediated resistance. These results suggest that this form of resistance may be useful for controlling ergot in commercial seed production fields. Further study of the inheritance and physiological basis for resistance in these sorghums is warranted.
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ACKNOWLEDGMENTS
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We thank Amanda Hall for the excellent technical assistance in greenhouse experiments. The Schultz Company (St. Louis, MO) provided fertilizer for this study. This research was supported by the Sorghum/Millet Collaborative Research Program (INTSORMIL) (AID/DAN-1254-G-00-1022-00), the Kansas Agriculture Experiment Station (North-Central Regional Research Project, NC-227), and a USDA-ARS Specific Cooperative Research Agreement (No. 58-6202-9-155).
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NOTES
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Contribution No. 01-57-J from the Kansas Agric. Exp. Stn.
Received for publication October 30, 2000.
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ABSTRACT
INTRODUCTION
