Evaluation of Tomato Germplasm against Tomato Brown Rugose Fruit Virus and Identification of Resistance in Solanum pimpinellifolium

Abstract

The tomato is one of the most important vegetable crops grown worldwide. Tomato brown rugose fruit virus (ToBRFV), a seed-borne tobamovirus, poses a serious threat to tomato production due to its ability to break the resistant genes (Tm-1, Tm-2, Tm-2²) in tomatoes. The objective of this work was to identify new resistant source(s) of tomato germplasm against ToBRFV. To achieve this aim, a total of 476 accessions from 12 Solanum species were tested with the ToBRFV US isolate for their resistance and susceptibility. As a result, a total of 44 asymptomatic accessions were identified as resistant/tolerant, including thirty-one accessions of S. pimpinellifolium, one accession of S. corneliomulleri, four accessions of S. habrochaites, three accessions of S. peruvianum, and five accessions of S. subsection lycopersicon hybrid. Further analyses using serological tests identified four highly resistant S. pimpinellifolium lines, PI 390713, PI 390714, PI 390716, and PI 390717. The inheritance of resistance in the selected lines was verified in the next generation and confirmed using RT-qPCR. To our knowledge, this is a first report of high resistance to ToBRFV in S. pimpinellifolium. These new genetic resources will expand the genetic pool available for breeders to develop new resistant cultivars of tomato against ToBRFV.

Keywords: Solanum lycopersicum; genetic resistance; tobamoviruses; tomato; tomato brown rugose fruit virus (ToBRFV).

Figure 1

Symptom classes of ToBRFV infection in tomato germplasm used to calculate a disease severity index (DSI). 0: no symptoms; 1: mild mosaic; 2: mosaic; 3: mosaic, leaf deformed; 4: severe mosaic, leaf deformed, mottling; and 5: severe mosaic, leaf deformed, mottling, and string leaves.

Figure 2

Re-evaluating individual plants from selected Solanum pimpinellifolium lines confirmed their resistance to ToBRFV but also revealed a segregating population. Absorbance values at OD_405nm for two individual plants derived from seeds generated from self-pollination (S₁) of four putative ToBRFV-resistant lines, (A) PI 390711 (line 326); (B) PI 390714 (line 329); (C) PI 390716 (line 331); and (D) PI 390717 (line 332) along with buffer, healthy tomato, and ToBRFV-infected tomato as controls for comparisons. Data (absorbance readings measured at OD_405nm) represent mean ± SEM for three experimental replicates. Statistical significance was analyzed using ordinary one-way ANOVA/Dunnette’s multiple comparison followed by Bonferroni post hoc test. Different letters indicate the statistical significance compared with buffer inoculation as mock control. Statistical analysis was performed using GraphPad Prism 9.

Figure 3

Re-evaluation of selected Solanum pimpinellifolium lines for their resistance to ToBRFV using an ELISA test to assess virus titers on leaf tissue samples collected from systemic leaves. Based on statistical analysis, plants from four lines were considered resistant (PI 390713, PI 390714, PI 390716, and PI 390717), whereas plants from two other lines (PI 390712 and PI 390718) were susceptible to ToBRFV. Buffer, healthy tomato, and ToBRFV-infected tomato were included in the same ELISA test as controls for comparison. Data (absorbance readings measured at OD_{405 nm}) represent mean ± SEM for three experimental replicates. Statistical significance was analyzed using ordinary one-way ANOVA/Dunnette’s multiple comparison followed by Bonferroni post hoc test. Different letters indicate the statistical significance compared with buffer inoculation as mock control. Statistical analysis was performed using GraphPad Prism 9.

Figure 4

Comparative evaluation on selected tomato lines for their resistance to ToBRFV using RT-qPCR to assess the virus titers on leaf tissue samples collected from systemic leaves. The Ct values from 3 plants (3 biological replicates) of the ToBRFV-resistant line (332-1, 332-2, and 332-3: PI 390717), in comparison to that of the ToBRFV-susceptible line (333: PI 390718), LA2830 (Tm-1 and Tm-2²), as well as ToBRFV-infected ‘Moneymaker’, and its mock inoculation control. Data (Ct values) represent mean ± SD for three experimental replicates. Statistical significance was analyzed using ordinary one-way ANOVA/Dunnette’s multiple comparison followed by Bonferroni post hoc test. Different letters indicate the statistical significance compared with buffer inoculation as mock control. Statistical analysis was performed using GraphPad Prism 9.

Figure 5

Assessing F₁ plants derived from selected Solanum pimpinellifolium lines for their resistance to ToBRFV as evaluated for their absorbance values (virus titers) using an ELISA test. F₁ plants from four crosses: 1-F₁: 327-1 [PI 390712 (S)] × 326 [PI 390711 (R)]; 2-F₁: 328-1 [PI 390713 (R)] × 326-1 [PI390711 (R)]; 3-F₁: 333-1 [PI 390718 (S)] × 329-1 [PI 390714 (R)]; and 4-F₁: 333-1 [PI 390718 (S)] × 332-1 [PI 390717 (R)] were evaluated for their resistance to ToBRFV, in comparison to the mock inoculation control and a positive control (ToBRFV+). Data (absorbance readings measured at OD_{405 nm}) represent mean ± SEM for three experimental replicates. Statistical significance was analyzed using ordinary one-way ANOVA/Dunnette’s multiple comparison followed by Bonferroni post hoc test. Different letters indicate the statistical significance compared with buffer inoculation as mock control. Statistical analysis was performed using GraphPad Prism 9.