Date of Graduation

8-2023

Document Type

Thesis

Degree Name

Master of Science in Plant Pathology (MS)

Degree Level

Graduate

Department

Entomology and Plant Pathology

Advisor/Mentor

Rupe, John C.

Committee Member

Rojas, Alejandro

Second Committee Member

Faske, Travis

Keywords

Target spot; Soybean; Greenhouse inoculation; Susceptibility

Abstract

Target spot, caused by the fungus Corynespora cassiicola, was once considered a minor foliar disease of soybean in the US. However, a severe outbreak of target spot occurred in the southeastern U.S in 2016. Due to the lack of information on resistance in soybean cultivars, and the challenges in screening cultivars in the field, a detached leaf assay was developed to evaluate resistance. The objectives of the effort were to 1) develop a greenhouse inoculation method of soybean with C. cassiicola 2) use that method to compare reactions of soybean germplasm lines to target spot 3) collect and characterize isolates of C. cassiicola from Arkansas soybean leaves. Initially, soybean seedlings were spray-inoculated with either a mycelial suspension or spores of the pathogen and placed in a dew chamber for three or more days. Small lesions developed but did not develop into typical target spot lesions. However, a detached leaf assay that placed a droplet of either inoculum on wounded leaves resulted in typical target spot lesions that continued to expand. There were significant differences between cultivars, with some cultivars showing little disease development. The detached leaf assay involved the use of detached unifoliate leaves. Wounded leaves were inoculated with a conidial suspension (1 x 105), incubated in Petri dishes on moist filter paper, and incubated at 23ºC under 12-hour light. Disease was assessed based on percent leaf area with symptoms and recorded between 7-17 days using the Bioleaf foliar analysis mobile application. Area under the disease progress curve (AUDPC) after inoculation was calculated. The assay was used to evaluate 37 breeding lines and the commercial cultivars DG48E49, LS4299XS and a moderately susceptible check, Hutcheson. Reactions ranged from resistant to susceptible and most lines were consistent between runs. A two-location field trial was conducted to determine target spot susceptibility. Although disease in both locations was low, there were significant differences between cultivars and between locations. However, there was low correlation between the field and detached leaf results. While the detached leaf assay consistently separates soybean cultivars based on disease development, further testing is required to address variability within and between tests, and additional field data is needed to determine the capability of the assay to predict field performance. To compare isolates of C. cassiicola, 28 single-spore isolates were collected from symptomatic soybean leaves at multiple Arkansas locations in 2020 and 2021. Isolates were compared morphologically in culture and the internal transcribed spacer (ITS) region of each isolate was sequenced. All isolates produced colonies and conidia consistent with C. cassiicola; isolates differed in the amount of sporulation. Based on sequence analysis in BLAST, the isolates were confirmed to be C. cassiicola with a ≥ 98% match with sequences of C. cassiicola in the National Center for Biotechnology Information (NCBI) database. The isolates were assessed for aggressiveness in the detached leaf assay by measuring the percent leaf area and calculating AUDPC. Isolate Cory 20-03 was selected to evaluate the soybean cultivars for resistance in the detached leaf assay. Despite the initial comparison of isolates showing a high level of consistency in culture and the ITS region, the variation in aggressiveness and sporulation ability suggests further sequencing of additional genes is required.

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