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ARS Home » Southeast Area » Canal Point, Florida » Sugarcane Field Station » Research » Research Project #432517

Research Project: Identification of Resistant Germplasm and Markers Associated with Resistance to Major Diseases of Sugarcane

Location: Sugarcane Field Station

2020 Annual Report

1. Identify disease resistant sugarcane and energy cane clones for high yielding commercial production. 2. Develop methodologies to efficiently screen germplasm for resistance and to identify new molecular markers associated with resistance. 3. Identify pathogenic variation in sugarcane pathogens that are endemic and emerging within the United States.

Objective 1: Sugarcane clones in the cultivar development program for both sucrose and bio-energy will be screened for their disease reaction in artificial inoculation tests separate from the cultivar development plots with the major pathogens and ratings will be determined based on incidence and severity of disease. Objective 2: For Sugarcane Yellowleaf Virus (SCYLV) existing markers will be tested against our known historical clones having both resistance and susceptible reaction. Two new populations will be screened for SCYLV resistance and genotyping by sequencing (GBS) to identify new markers. For ratoon stunt disease and smut two populations will be screened for disease reaction and GBS to identify markers. For orange rust we will test published markers and perform fine mapping using a larger population and whole genome sequencing of the parents and bulked progeny to identify markers for screening the whole population. Objective 3: Orange rust spores have been collected over the last two years from different cultivars and locations in Florida and stored for determining possible pathogenic variation. Specifically, cultivars CP 80-1743 (the susceptible cultivar first observed with orange rust in Florida), CL 85-1040 (susceptible) and two cultivars CP 88-1762 and CP 89-2143 that were originally resistant but became susceptible will be used to evaluate pathogenic variation. The whorl inoculation technique (Sood et al. 2009) that is routinely used to evaluate brown and orange rust reactions will be used to evaluate variation between the isolates. The whorl technique has been shown to give consistent results, as seen by the sample data in Table 2 of the Appendix. These will be used along with the new collections described. Sentinel plots of orange rust susceptible and resistant sugarcane cultivars will be planted at 5 different locations and from grower’s fields where the “breaking down” of resistance is reported. Fields will be observed monthly from March to July when orange rust is most prevalent. Orange rust pustules that develop on previously resistant cultivars will be collected and compared using the whorl inoculation techniques to isolate the collected spores from different cultivars differing in susceptibility over several years. Mass rust spore collections from the field will be inoculated on plants of the same cultivar twice as a means to help purify the isolate. Variation in symptom development of isolates on specific cultivars will be used to identify pathogenic differences. Unfortunately, pathogen variation will be limited to isolates from Florida because isolates from outside the state are prohibited from introduction.

Progress Report
Economic losses caused by sugarcane diseases, especially sugarcane brown and orange rusts, mosaic, ratoon stunting disease (RSD), smut, and leaf scald, are substantial. Thus, the Canal Point (CP) cultivar development programs (CP programs) screen its germplasm and advanced breeding lines for resistance to the above diseases. Data are obtained from both natural infection and inoculated trials ensuring that resistant or disease tolerant clones are advanced and released from the program. Because, pathogenic variations occur over time, developing resistant cultivars is a continuous effort in the CP programs. Canal Point (CP) cultivar development programs have been releasing disease resistant high yielding cultivars for the Florida sugarcane industry. Thus, all susceptible clones in the seedling, Stage I, and Stage II are eliminated if exhibiting disease symptoms based on a natural infection. In two CP cultivar development programs (i.e., CP program for organic (muck) soils and CP program for mineral (sand) soils), clones in both the muck and sand programs were screened annually in inoculation tests at Stage III (135 clones), Stage III increase (40 clones) and Stage IV (13 clones) for their resistance to ratoon stunt, smut, brown rust, orange rust, leaf scald, and mosaic. Clones in Stage I and II are screened for resistance to rusts, mosaic, leaf scald, and smut only by natural infection. All clones with acceptable resistance levels were advanced to later stages of the programs. High-yielding and disease resistant or tolerant clones are released for commercial cultivation by the Florida Sugarcane Variety Committee. Sugarcane orange rust appeared in the Western hemisphere in 2007 and impacted the Florida sugarcane industry as well as the CP cultivar development program. Marker-assisted selection along with the phenotypic evaluation of disease resistance will be beneficial for the CP program therefore quantitative trait loci associated with orange rust as well as with brown rust resistance and SCYLV resistance have been identified. The developed diagnostic markers for orange rust resistance are being used in the CP program. To detect QTL for other diseases, we have inoculated leaf scald, RSD, and smut pathogen in the field and pot trials for the diseases screen in the first ratoon crop of a population from a cross of CP 72-2086 and CP 01-2390. Currently, we are analyzing the genotype data including marker filtering for mapping and QTL analysis. Seedlings of the population derived from a reciprocal cross between CP 01-2390 and CP 72-2086 were planted for seed increase. Since we have already identified and successfully validated the markers associated with orange rust, we decided not to further pursue the orange rust markers fine-mapping study. However, we have initiated transcriptome analysis through RNA-sequencing for identifying the candidate genes associated with orange rust. We were unable to test the effectiveness of the SCYLV reported markers by Debabakas et al. (2014) using our historical population as the corresponding author did not respond to our request to provide the marker sequence. However, we were able to analyze QTL associated with SCYLV resistance using another two populations derived from cross CP 95-1039 X CP 88-1762 and Green German X IND81-146. Genotyping of SCYLV and RSD population from a cross between CP 72-2086 and CP 01-2390 has been completed using NGS based Exon capture sequence-based technique. Also, phenotype data (leaf scald, sugar, and yield traits) from the plant cane and ratoon have been collected for the same population. Another validation population from the reciprocal using the same parents has been planted for seed increase. Genomic DNA and RNA were extracted from spores of P. kuehnii collected from sugarcane cultivars CL 85-1040 and CP 89-2143. These two rust isolates showed pathogenic variation. Pathogenic variation was also observed in the sugarcane mosaic virus.

1. Development of disease-resistant cultivars. Sugarcane diseases cause significant yield losses in susceptible varieties. The most economical and environmentally sustainable approach to limit these losses is to grow disease tolerant cultivars. The six cultivars were developed by scientists at ARS Canal Point, Florida, with the collaboration of the University of Florida and Florida Sugar Cane League. The disease-resistant CP cultivars have contributed greatly to the sustainable production of sugarcane in Florida. Also, these disease-resistant high yielding sugarcane cultivars help Florida sugarcane growers to economically grow sugarcane and produce approximately 20% of the sugar consumed in the United States.

2. Quantitative trait loci (QTL) associated with disease resistance. Brown and orange rusts are devastating diseases for the sugarcane industry in Florida as well as in other sugarcane growing areas. The selection of disease resistance by the molecular marker is more reliable and should complement inoculation and natural infection trials. QTL associated with orange rust resistance as well as with brown rust resistance have been identified by ARS researcher at Canal Point, Florida. Diagnostic markers have been developed for orange rust resistance. QTL associated with SCYLV resistance has been detected and published. RNA sequencing has been performed from 30 samples (2 clones X 5-time points X 3 replications) for transcriptome and candidate gene analysis associated with orange rust. We identified 217 nonredundant markers and 225 candidate genes to be significantly associated with the yield traits, which can serve as a comprehensive genetic resource database for future gene identification, characterization, and selection for sugarcane improvement. This study also suggests that genomic inbreeding has led to negative impacts on sugarcane yield.

Review Publications
Coto, A.O., Sandhu, H.S., Zhao, D., Gordon, V.S., Comstock, J.C., Sood, S.G., McCorkle, K.M., Davidson, W.R., Baltazar, M., McCord, P.H., Singh, M.P. 2020. Registration of ‘CP 09-1385’ sugarcane. Journal of Plant Registrations. 14(3):328-339.
Zhao, D., Davidson, W.R., Gordon, V.S., Islam, M.S., McCord, P.H., Sandhu, H.S., Sood, S.G., Comstock, J.C., Baltazar, M., Singh, M.P. 2020. Registration of ‘CP 11-2248’ sugarcane for the Florida organic soils. Journal of Plant Registrations. 14(3):318-327.
Yang, X., Luo, Z., Todd, J.R., Sood, S.G., Wang, J. 2020. Genome-wide association study of multiple yield components in a diversity panel of polyploid sugarcane (Saccharum spp.). Genetics. Plant Genome 13(1).
Boukari, W., Wei, C., Tang, L., Hincapie, M., Naranjo, M., Nuessly, G., Beuzelin, J., Sood, S.G., Rott, P. 2020. Lack of transmission of sugarcane yellow leaf virus in Florida from Columbus grass and sugarcane to sugarcane with aphids or mites. PLoS One. 15(3), 1-10.
You, Q., Yang, X., Peng, Z., Islam, M.S., Sood, S.G., Luo, Z., Comstock, J.C., Xu, L., Wang, J. 2019. Development of an Axiom sugarcane 100K SNP array for high-resolution genetic map construction and QTL identification. Journal of Theoretical and Applied Genetics.
Islam, M.S., McCord, P.H., Sandhu, H.S., Zhao, D., Davidson, W.R., Gordon, V.S., Sood, S.G., Comstock, J.C., Singh, M.P., Baltazar, M. 2019. Registration of 'CP 10-2195' sugarcane. Journal of Plant Registrations. 13:368-376.
Gordon, V.S., McCord, P.H., Sandhu, H.H., Zhao, D., Davidson, W.R., Comstock, J.C., Sood, S.G., Abbott, T., Singh, M.P., Islam, M.S. 2019. Registration of 'CP 10-1619' sugarcane use on sand soils. Journal of Plant Registrations. 13:353-361.
Singh, M., Edme, S.J., Zhao, D., Comstock, J.C., Davidson, W.R., Gordon, V.S., Sandhu, H., McCord, P.H., Sood, S.G., Baltazar, M. 2019. Registration of 'CP 09-4153', 'CPCL 09-4160', and 'CP 09-4758' sugarcane for sand soils in Florida. Journal of Plant Registrations. 13:345-352.
Davidson, W., Scott, A., Hernandez, E., Gordon, V.S., McCord, P.H., Sandhu, H., Zhao, D., Comstock, J.C., Sood, S.G., Singh, M., Islam, M.S., Baltazar, M., McCorkle, K.M. 2019. Registration of 'CP 08-1968' sugarcane. Journal of Plant Registrations. 13:178-186.