Discrepancies in Serology-Based and Nucleic Acid-Based Detection and Quantitation of Tomato Spotted Wilt Orthotospovirus in Leaf and Root Tissues from Symptomatic and Asymptomatic Peanut Plants

Abstract

Thrips-transmitted tomato spotted wilt orthotospovirus (TSWV) causes spotted wilt disease in peanuts. A serological test (DAS-ELISA) is often used to detect TSWV in peanut leaf samples. However, in a few studies, DAS-ELISA detected more TSWV infection in root than leaf samples. It was not clear if the increased detection was due to increased TSWV accumulation in root tissue or merely an overestimation. Additionally, it was unclear if TSWV detection in asymptomatic plants would be affected by the detection technique. TSWV infection in leaf and root tissue from symptomatic and asymptomatic plants was compared via DAS-ELISA, RT-PCR, and RT-qPCR. TSWV incidence did not vary by DAS-ELISA, RT-PCR, and RT-qPCR in leaf and root samples of symptomatic plants or in leaf samples of asymptomatic plants. In contrast, significantly more TSWV infection and virus load were detected in root samples of asymptomatic plants via DAS-ELISA than other techniques suggesting that DAS-ELISA overestimated TSWV incidence and load. TSWV loads from symptomatic plants via RT-qPCR were higher in leaf than root samples, while TSWV loads in leaf and root samples from asymptomatic plants were not different but were lower than those in symptomatic plants. These findings suggested that peanut tissue type and detection technique could affect accurate TSWV detection and/or quantitation.

Evaluation of Wild Peanut Species and Their Allotetraploids for Resistance against Thrips and Thrips-Transmitted Tomato Spotted Wilt Orthotospovirus (TSWV)

“…To determine TSWV infection status, plant tissue (ca 0.03 g) was collected from the first fully expanded leaf below the terminal and subjected to a double antibody enzyme-linked immune sorbent assay (DAS-ELISA). DAS-ELISA with TSWV-specific polyclonal antibodies (Adgia, Elkhart, IN, USA) was used to detect TSWV in peanut tissues in the laboratory using the protocol described in Lai et al, 2021 [ 38 ].…”

Characterization of gene expression patterns in response to an orthotospovirus infection between two diploid peanut species and their hybrid

“…The reaction was extended with a melting curve in a QuantStudio 3 System (applied biosystems by Thermo Fisher Scientific, Waltham, MA) to rule out non-specific binding. Two technical replicates for targeted transcripts and the reference gene (alcohol dehydrogenase class III) ( Lai et al., 2021b ), and water control were included in each RT-qPCR run. The log 2 fold change of each target transcript in infected plants against mock-inoculated plants was calculated after normalization to the reference gene.…”

Advances in molecular plant pathology, plant abiotic and biotic stress

“…The reaction was extended with a melting curve in a QuantStudio 3 System (applied biosystems by Thermo Fisher Scientific, Waltham, MA) to rule out non-specific binding. Two technical replicates for targeted transcripts and the reference gene (alcohol dehydrogenase class III) (Lai et al, 2021b), and water control were included in each RT-qPCR run. The log 2 fold change of each target transcript in infected plants against mockinoculated plants was calculated after normalization to the reference gene.…”