Location: Crop Genetics Research
2023 Annual Report
Objectives
1. Characterize the genetic mechanisms for controlling nematode resistance for diploid cotton species.
1.A. Genetic characterization of reniform nematode resistance for selected G. arboreum germplasm accessions.
2. Transfer novel nematode resistance identified for diploid cotton species to tetraploid upland cotton.
2.A. Introgression of reniform nematode resistance from G. arboreum germplasm accessions into upland cotton cultivars.
3. Determine environmental influences on nematode infection, development, and reproduction on cotton lines.
3.A. Determine soil temperature effects on nematode infection, development, and reproduction on cotton lines.
3.B. Determine soil moisture effects on nematode infection, development, and reproduction on cotton lines.
Approach
Develop populations by crossing resistant accessions with one or more Gossypium (G.) arboreum accessions classified as susceptible or highly susceptible to determine the inheritance of the resistance. Ovule culture will be used to introgress resistance from G. arboreum accessions into G. hirsutum varieties. Gossypium accessions with high levels of resistance to reniform nematode will be evaluated in growth chamber experiments to measure the effects of soil temperature and soil moisture stress on the number of infections, rate of development of females after infection, and production of eggs.
Progress Report
Reniform nematodes cause hundreds of millions of dollars loss in U.S. cotton production annually. Resistance to this important parasite was not found in Upland cotton, but in some diploid cotton species. The genetic barriers of uneven chromosome numbers make it very difficult to introduce resistant genes from diploid cotton species with two sets of chromosomes to the tetraploid upland cotton with four sets of chromosomes. Cotton bolls produced from direct crossing diploid and tetraploid cotton varieties rarely reach maturity, and the derived triploid hybrid plants with three sets of chromosomes will be sterile. To introduce useful genes from diploid cotton species to upland cotton by directly crossing these parents, two critical processes are necessary: embryo rescue by in vitro tissue culture, and chromosome doubling by treating cotton plants with chemicals such as colchicine.
Objectives 1 and 2 are focused on identifying and transferring resistance from the diploid cotton species Gossypium arboreum into tetraploid Upland cotton. The geneticist position on this project is leading the work on these objectives, but this position was vacant for most of the year. As a result, there will be a delay in meeting the milestones for these objectives. In early May of 2023, the position was filled and work on these objectives was initiated.
For in vitro embryo rescue purpose, five media with different concentrations and combinations of plant growth regulators have been tested. After surface sterilization, hundreds of embryos were dissected from young bolls two to three days post anthesis, placed on different media and incubated in a growth chamber at 27 C and 12 hours light/12 hours dark light regime. Embryos can grow on each of the five media. Calli were induced from some of the embryos cultured.
For chromosome doubling, two concentrations and different durations of colchicine treatments were tested by treating the meristem of cotton branches and seedlings. Cotton branches and seedlings may survive after they were treated with 0.05% and 0.1% of colchicine for 24 hours. On 27 triploid plants derived from the crosses of tetraploid upland cotton with some A or D genome diploid species, 48 out of 68 branches (70%) treated with 0.05% colchicine, and 29 out of 48 branches (60%) treated with 0.1% colchicine survived. The chromosome numbers of these treated branches will be inspected. If the chromosome number of a branch has been doubled, seed will be saved from this branch, and be used in the future as bridge species to cross with resistant G. arboreum accessions to produce fertile tetraploid progenies with resistance to reniform nematodes.
Alternatively, the diploid plants can be treated with colchicine to generate tetraploid plants, which should be able to cross with upland cotton and produce fertile progenies. In this way it would avoid the embryo rescue procedure and further chromosome doubling processes. So far, seedlings of three G. arboreum accessions (A2-190, A2-272, and A2-514) resistant to reniform nematodes were treated with 0.05% or 0.1% colchicine for 24 hours; eight, two and four plants from these lines, respectively, survived after the treatments and are growing in the greenhouse. The chromosomes of these plants will be counted before they are used to cross with upland cotton.
Given the challenges with introducing resistance to reniform nematode into Upland cotton from related species, work was initiated to better characterize the efficacy of the resistance when the plants are challenged with changing environmental conditions. Work on Objective 3 of the project has proceeded as planned during the first 12 months of the project. The initial work on this objective is focused on determining soil temperature effects on reniform nematode infection, development, and reproduction on cotton lines that are resistant to the nematode. During this reporting period, three growth chamber tests were conducted to characterize the resistant Gossypium barbadense accessions GB713 and TX110, with susceptible Gossypium hirsutum cultivar Deltapine 16 included as a control. Two of the tests were conducted at a soil temperature of 24ºC (cooler than normal), and one test was conducted at a soil temperature of 28ºC; 28ºC is the temperature at which all of our germplasm screening work was conducted (normal temperature). Data collection for the two tests conducted at 24ºC is complete, and data collection for the test conducted at 28ºC is still in progress. The second run of the 28ºC test is scheduled to begin in July 2023.
This project includes a congressionally-mandated subordinate project (6066-22000-094-001S). Research on this subordinate project during the past 12 months focused on determining the impact of naturally-occurring reniform nematode populations on nematode reproduction and cotton yield of three USDA reniform resistant cotton lines, three commercial Phytogen reniform nematode resistant varieties, and three commercial reniform susceptible cotton varieties in a field trial. On June 1, 2022 plots were planted to compare the effects on nematode populations and cotton yield. Soil samples were collected at planting, mid-season and post-harvest for nematode quantification. Plots were harvested in October 2022 and yield was calculated. Results were presented at the 2023 Beltwide Cotton Conference poster session held in New Orleans, Louisiana January 10-12, 2023. A second field season was initiated on May 30, 2023 to include the identical set of treatments from 2022 to confirm results. To confirm a third iteration of results and prepare for publication, a growth chamber experiment will be initiated in July of 2023 using the identical treatments from field trials. All results will be analyzed and a manuscript will be prepared to fulfill the requirements listed in the project outputs.
Accomplishments
1. Development of a nondestructive method to assess reniform nematode resistance in Gossypium species. The reniform nematode, a microscopic worm that feeds on plant roots, is damaging to cotton. To rapidly determine whether or not a plant has resistance to this nematode, the plant’s entire root system is cut off, stained to show the nematodes attached to it, and the nematodes are counted. The purpose of this work was to find a way to rapidly assess the number of nematodes on the roots without killing the plant. In one set of experiments conducted by ARS researchers in Stoneville, Mississippi, the root system was cut off at either 0, 1, 2.5, or 5 cm below the soil line, and used to count the number of nematodes infecting this portion of the root system. Resistant plants could be accurately identified while leaving up to 5 cm of roots with the shoot. In a second set of experiments, the rate of plant recovery and development of bolls and seeds was evaluated using a combination of root retention and shoot retention treatments. Plants rapidly recovered using a treatment combination in which the top leaves and 2.5 cm roots were kept. This combination saved more than 20 days to harvest compared to some other treatment combinations. This method will be useful for cotton breeders to screen their plants more effectively for resistance to reniform nematode while retaining their best materials to produce the next generation of plants.
Review Publications
Stetina, S.R., Erpelding, J.E. 2023. Nondestructive evaluation of cotton genotypes for resistance to reniform nematode. Journal of Cotton Science. 27:102-109. https://doi.org/10.56454/TDDZ4193.
Singh, B., Chastain, D.R., Stetina, S.R., Gardiner, E.S., Snider, J.L. 2022. Early-season growth responses of resistant and susceptible cotton genotypes to reniform nematode and soil potassium application. Agronomy. 12(11):2895. https://doi.org/10.3390/agronomy12112895.
