Field Assessment of Anthracnose Disease Response for the Sorghum Germplasm Collection from the Mopti Region

Problem statement: Sorghum anthracnose ( Colletotrichum sublineolum) is a highly variable pathogen and new sources of host plant res istance are required for the development of resistant sorghum varieties. Germplasm collections are an important source of host plant resistance and screening germplasm will be essential to identi fy new sources of resistance. Approach: The sorghum ( Sorghum bicolor) collection from the Mopti region of Mali was inoc ulated with Colletotrichum sublineolum and evaluated for foliar anthracnose disease respo nse in Isabela, Puerto Rico during the 2004 and 2005 growing seasons using a partially balanced lattice design with three replications. Results: A resistant response was observed for 45 of the 97 accessions in the collection and mean infection severity for the 52 susceptible accessions was 27.6%. An association was observed between resistance and the administrative district where the germplasm was collected. More than 50% of the accessions from the Bandiagara and Bankass districts showed a resistant response. The lowest frequency of resistant germpla sm was observed for the Mopti district with 25% of the accessions showing a resistant response. The susceptible accessions from the Mopti district, however, showed the lowest mean infection severity. Approximately 44% of the accessions from the Douentza district showed a resistant response with the susceptible accessions showing the highest mean infection severity. These results suggest an a ssociation between annual rainfall and anthracnose resistance, with sorghum accessions fro m d ier regions showing greater susceptibility. Anthracnose resistance also showed an association w ith sorghum race classification and race guinea accessions were more frequently resistant as compar ed to accessions classified as race durra or durra-bicolor. Conclusion: The results indicated that anthracnose resistant s orghum germplasm is frequent in the Mopti region of Mali and that ecoge raphic origin and sorghum characterization information can be used to aid in germplasm selecti on or germplasm acquisition to identify anthracnose resistant sources.


INTRODUCTION
Anthracnose is considered one of the most destructive diseases for sorghum (Sorghum bicolor (L.) Moench) production due to the rapid development of the disease on susceptible cultivars. Colletotrichum sublineolum (P. Henn. in Kabat and Bubák) is the fungal pathogen responsible for sorghum anthracnose (Crouch et al., 2006) and was first reported in Togo, West Africa in 1902 (Thakur and Mathur, 2000). The disease occurs worldwide, but is more commonly observed in tropical or subtropical environments where frequent rainfall, high relative humidity and warm temperatures enhance the development and spread of the disease (Casela et al., 2001;Hess et al., 2002;Marley et al., 2001;Néya and Le Normand, 1998;Ngugi et al., 2002;Thakur and Mathur, 2000;Thomas et al., 1996;Valério et al., 2005). In the United States, anthracnose is more prevalent in the Southern Plains and Southeastern States (Rosewich et al., 1998;Cardwell et al., 1989). Sorghum anthracnose will typically appear on infected leaves approximately 30 to 40 days after seedling emergence; although, infection can occur at every stage of plant development. Anthracnose infection can be observed on all above ground tissues of the sorghum plant, including the leaf, stalk, panicle and seed (Hess et al., 2002;Thakur and Mathur, 2000). Foliar infection is more commonly observed and can be a source of inoculum for infection of other tissues. Typical symptoms include circular, elliptical, or elongated lesions depending on host plant response. During sporulation of the fungus, acervuli, asexual fruiting bodies, will appear as black spots in the center of the lesions on susceptible cultivars.
Grain yield losses from foliar anthracnose infection are typically associated with a reduction in grain size as the infection will affect grain development. Yield losses greater than 50% may occur under epidemic conditions (Thakur and Mathur, 2000). In experimental evaluations, grain yield losses ranging from 30-67% have been report Thomas et al., 1996). In Puerto Rico, grain yield losses of 100% have been observed for highly susceptible sorghum germplasm accessions with plant death occurring prior to flowering. The disease can be successfully managed through the use of resistant cultivars. However, the long-term durability of resistant cultivars is hindered by variation in virulence within the pathogen population Cardwell et al., 1989;Marley et al., 2001Pande et al., 1991Thakur and Mathur, 2000;Valério et al., 2005). Pyramiding of resistance genes can aid in the development of resistant cultivars, but additional sources of resistance are needed for sorghum improvement.
The United States Department of Agriculture, Agricultural Research Service, National Plant Germplasm System (USDA-ARS, NPGS) sorghum collection is a valuable resource for the identification of new sources of anthracnose resistance. The NPGS maintains more than 43,000 sorghum accessions and field evaluation of germplasm from the collection has successfully identified anthracnose resistant sources (Erpelding and Prom, 2006;Erpelding and Wang, 2007). An anthracnose evaluation of a subset of sorghum germplasm from the Mali collection indicated resistance was frequent in this collection (Erpelding and Prom, 2004); however, it is unknown if resistance was associated with specific ecogeographic regions of Mali. Therefore, an anthracnose field evaluation was conducted for sorghum landraces from the Mopti region of Mali. The objectives of the anthracnose evaluation were: (1) determine the frequency of anthracnose resistance for the sorghum landraces from the Mopti region and (2) determine if resistance was associated with ecogeographic origin and sorghum phenotypic characteristics.

MATERIALS AND METHODS
Ninety-seven landraces from the Mopti region of Mali maintained in the NPGS sorghum collection were identified using available passport information and seed samples were obtained from the USDA-ARS Plant Genetic Resources Conservation Unit, Griffin, Georgia .   Three anthracnose resistant control genotypes, NSL  365745, PI 148097 and SC748-6 and six susceptible  control genotypes, PI 257599, PI 276842, PI 561472, PI  564163, PI 609251 and PI 609634, were included in the evaluation. PI 609251 was also included in the evaluation as a non-inoculated control. The anthracnose field evaluations were conducted at the USDA-ARS Tropical Agriculture Research Station in Isabela, Puerto Rico. The first evaluation was planted on 6 July 2004 with the second evaluation planted on 7 March 2005 using a partially balanced lattice design with three replications. The 97 sorghum accessions and 10 controls were planted in single rows 1.8 m in length with 0.9 m row spacing. A border row of anthracnose susceptible genotypes was planted around each experimental field. At planting, fertilizer was applied at a rate of 560 kg ha −1 (15-5-10 NPK) and to prevent seed loss from fire ants, Lorsban 15G (Chlorpyrifos) granular insecticide (Dow AgroSciences, Indianapolis, IN) was applied at a rate of 8 kg ha −1 . Supplemental irrigation was applied after planting for stand establishment and three times before inoculation for the evaluation conducted in 2004 and five times before inoculation for the evaluation conducted in 2005. No irrigation was applied after inoculation. Weeds were controlled with mechanical tillage.
The anthracnose inoculum used for disease evaluation was prepared from anthracnose infected sorghum leaves randomly collected from research plots in Isabela, Puerto Rico before establishment of experiments to represent the pathotypes present at the research location. Preparation of anthracnose cultures, field inoculation and disease evaluation were as described by Erpelding and Prom (2006). Plants were inoculated with anthracnose-colonized sorghum seed 37 days after planting in 2004 and 33 days after planting in 2005. Anthracnose infection response was evaluated at 18, 31, 56 and 81 days after inoculation in 2004. The third and final evaluations were delayed by a tropical storm and the evaluations were conducted after the plants recovered. Field evaluations in 2005 were conducted at 35, 48 and 63 days after inoculation. Anthracnose infection response was evaluated using a 1-5 rating scale based on disease response observed on inoculated leaves and disease progression on noninoculated leaves (Erpelding and Prom, 2004). Resistant plants were rated as 1 or 2, moderately susceptible plants as 3, susceptible plants as 4 and highly susceptible plants as 5. Plants rated as 1 showed no disease symptoms and plants rated as 2 showed reddening of inoculated leaves and no acervuli development. Susceptible plants showed the development of acervuli on inoculated leaves with chlorotic lesions observed on plants rated as 3 and necrotic lesions observed on plants rated as 4 or 5. Plants rated as 4 showed infection spreading to non-inoculated leaves with infection observed on most leaves including the flag leaf for plants rated as 5. The percentage of infected leaf area was also determined for the susceptible accessions during the final evaluation, with infection severity based on a visual estimate of leaf infection for the susceptible plants within a row. Statistical analysis was conducted on the disease severity data from the final evaluation using the Statistix software package (Analytical Software, Tallahassee, FL).

RESULTS
The anthracnose disease response for the 97 sorghum accessions from the Mopti region of Mali is presented in Table 1   In 2004, the presence of acervuli on inoculated leaves for the susceptible controls was observed for the evaluation conducted 12 days after inoculation and disease rapidly progressed to non-inoculated leaves with the controls rated as susceptible or highly susceptible 31 days after inoculation.
The Mopti region is divided into eight administrative districts, Bandiagara, Bankass, Djenne, Douentza, Koro, Mopti, Tenenkou and Youvarou and sorghum landraces from the Bandiagara, Bankass, Douentza and Mopti districts are maintained in the NPGS sorghum collection (Table 1). Approximately 54% of the sorghum accessions from the Bandiagara and Bankass districts showed a resistant response with a similar infection severity for the susceptible accessions observed for the two districts. Mean infected leaf area was 24.4% for the susceptible accessions from the Bandiagara district and 22.3% for the susceptible accessions from the Bankass district. Approximately 44% of the accessions from the Douentza district showed a resistant response and 40% of the landraces evaluated for the sorghum collection from the Mopti region were from the Douentza district. Mean infected leaf area was 35.2% for the susceptible accessions from the Douentza district. For the Mopti district, 25% of the accessions showed a resistant response and mean infected leaf area was 19.3% for the susceptible accessions. Approximately 12% of the landraces included in the anthracnose evaluation were from the Mopti district. The differences in disease severity for all accessions from the four districts, however, were not significant.
Bankass is the southern-most district and would receive the highest annual rainfall for the region. The frequency of anthracnose resistant accessions and the infection severity for the susceptible accessions from the Bankass district was similar to the drier, northern Bandiagara district. Annual rainfall for the Mopti district would be similar to the Bandiagara district; although, the lowest frequency of anthracnose resistant accessions was observed for the Mopti district. However, the susceptible accessions from the Mopti district showed the lowest infection severity. The Douentza district receives the lowest annual rainfall in the Mopti region. The highest infection severity for the susceptible accessions was observed for landraces from the Douentza district. A lower frequency of anthracnose resistant landraces was also observed for the Douentza district.
The landraces from the Mopti region were classified into seven sorghum phenotypic races (Table 1). A small number of landraces were classified as bicolor, caudatum, guinea-bicolor and guineacaudatum and thus no comparisons could be made for these races. Race durra accessions showed greater disease severity with a mean infected leaf area of 42.6%. Only five of the 20 race durra accessions showed a resistant response. Race durra accessions were present in the germplasm collections from the four administrative districts, but were more frequent in the collections from the Bankass and Douentza districts. Mean infection severity for the race durra accessions was similar between the four administrative districts, but infection severity was generally higher for the durra landraces from the Douentza district. Sorghum landraces that were classified as race durra-bicolor also showed greater disease severity with a mean infected leaf area of 31.2%. Three of the 15 landraces classified as race durra-bicolor showed a resistant response and nearly all race durra-bicolor accessions were collected from the Douentza district. More than 60% of the durra and durra-bicolor landraces were rated as highly susceptible. Race guinea accessions were the most frequent and comprised approximately 59% of the accessions from the Mopti region. Approximately 61% of the race guinea accessions showed a resistant response and mean infected leaf area was 13.4% for the susceptible race guinea accessions. The mean infection severity was significantly lower for the race guinea accessions as compared to the race durra and durrabicolor accessions. Race guinea accessions were present in the collections from the four administrative districts, but nearly all accessions from the Mopti district were classified as race guinea and more than 65% of the accessions from the Bandiagara and Bankass districts were race guinea. The Douentza district had the lowest frequency of race guinea accessions with approximately 38% of the accessions classified as race guinea. Also, the highest mean infection severity, 25.5%, was observed for the race guinea accessions from the Douentza district; whereas, the lowest mean infection severity, 3.6%, was observed for the race guinea accessions from the Bankass district. Additionally, most of the accessions that showed variation for infection response within and between experiments were classified as race guinea. The race guinea accessions from the Mopti region can be further classified into two sorghum working groups, guineense and margaritiferum (data not shown). Anthracnose resistance was more frequently associated with guineense landraces. Approximately 72% of the guineense landraces showed a resistant response as compared to 31% of the margaritiferum landraces. Infection severity was similar for the susceptible guineense and margaritiferum landraces with a mean infected leaf area of 10.9% for the guineense landraces and a mean infected leaf area of 15.0% for the margaritiferum landraces; however, the guineense landraces were significantly more resistant than the margaritiferum landraces. The lowest mean infection severity, 0.6%, was observed for the four susceptible guineense landraces from the Bankass district. In comparison, mean infection severity was 15.2% for the six susceptible guineense landraces from the Mopti district.

DISCUSSION
Approximately 46% of the landraces from the Mopti region showed a resistant response suggesting that this region of Mali could be an important source of anthracnose resistant germplasm. The Mopti region receives less than 800 mm of annual rainfall (Hess et al., 2002). Several studies have indicated that anthracnose infection severity is greater in regions receiving higher annual rainfall (Hess et al., 2002;Néya and Le Normand, 1998;Ngugi et al., 2002); thus, selection pressure may favor a higher frequency of anthracnose resistant germplasm in these wetter regions. Presumably, sorghum landraces from regions receiving low annual rainfall, such as the Mopti region, would show greater disease severity and a lower frequency of anthracnose resistant germplasm. Mean infection severity for the susceptible accessions was 27.6% and nearly 40% of the accessions were rated as highly susceptible with approximately 54% of the accessions showing susceptibility to anthracnose infection. Even though a greater percentage of the landraces from the Mopti region showed anthracnose susceptibility, the frequency of anthracnose resistant landraces in the collection would suggest resistance is being maintained in the germplasm from this region of Mali. This may suggest that climatic conditions favorable for the pathogen could occur in the Mopti region contributing to a higher frequency of anthracnose resistance in the sorghum landrace population. It is also possible that sorghum landraces introduced from other regions more favorable for anthracnose disease development would have contributed to the greater frequency of resistance observed for the landraces. Additionally, the greater frequency of anthracnose resistance that is being maintained in the landrace collection from a region of low annual rainfall may contribute genetic diversity for disease resistance.
Although latitude and longitude passport information is lacking, the data that is available would suggest an association between disease response and ecogeographic origin, with anthracnose resistant accessions more frequently observed in regions receiving higher annual rainfall. However, the overall drier climatic conditions and lower variation in annual rainfall may influence selection for anthracnose resistance for the sorghum landraces cultivated in this region. The acquisition of additional sorghum germplasm from the Mopti region would enhance the ecogeographic assessment of anthracnose resistance for this region.
The results of this study would also suggest an association between resistance and sorghum race classification. Durra landraces are generally associated with regions receiving low annual rainfall. Compact panicles with large, white seed are common characteristics of race durra and these traits are also associated with higher susceptibility to grain mold, as such, durra landraces are commonly grown in regions where rainfall is infrequent during seed maturation. If selection pressure for anthracnose resistance is lower in drier regions, then race durra accessions should show greater susceptibility to anthracnose as was observed in this study. In contrast, race guinea accessions are typically associated with regions receiving higher annual rainfall. Selection pressure for anthracnose resistance should be greater in the wetter regions and race guinea accessions should show a greater frequency of anthracnose resistance with lower infection severity observed for the susceptible accessions. The majority of the race guinea accessions from the Mopti region showed a resistant response and infection severity was significantly lower for the susceptible guinea accessions suggesting greater selection for anthracnose resistance.

CONCLUSION
Anthracnose resistant germplasm was frequent in the sorghum collection from the Mopti region, even though this is a drier region of Mali. Additionally, resistance was associated with ecogeographic origin and sorghum race classification, which could be successfully used to select germplasm for anthracnose evaluation to increase the likelihood of identifying a greater frequency of resistant accessions.

ACKNOWLEDGEMENT
Mention of trade names or commercial products in this publication is solely for the purpose of providing specific information and does not imply recommendations or endorsement by the US Department of Agriculture.