High-Throughput Virus Screening in Crosses of South American and African Cassava Germplasm Reveals Broad-Spectrum Resistance against Viruses Causing Cassava Brown Streak Disease and Cassava Mosaic Virus Disease

Abstract

Screening cassava lines for resistance against viruses causing cassava brown streak disease (CBSD) is cumbersome because of the unpredictable and erratic virus infections in the slow plant infection processes that are frequently not associated with distinct leaf symptoms and because of the reliance on the assessment of root necrosis as an indicator of plant resistance/tolerance. The selection of resistant candidates thus extends over several growing cycles and is still associated with uncertainties about the stage of virus infection. To reduce the time for selection of resistant crosses and the uncertainties associated with field screening, we have developed a fast-forward virus screening workflow to assess cassava seedlings from crosses of cassava brown streak- and cassava mosaic virus-resistant parents. After passing through an intensive and precise virus infection routine, cassava seedlings that carried resistance against cassava brown streak and mosaic viruses were identified. Taking the results of 195 seedlings from 18 crossing families together, it became evident that resistance against the viruses causing CBSD is a dominant trait. The protocol developed for virus resistance screening in cassava can be readily adopted. It shifts resistance evaluation from the field to the nursery and replaces the erroneous and lengthy virus infection and screening process with a method of precision and speed.

Developing broad-spectrum resistance in cassava against viruses causing the cassava mosaic and the cassava brown streak diseases

“…At the same time, the lines are being grown at three 4 Screening for resistance against viruses causing CBSD X CMD in the field Several strategies can be followed to accelerate virus-resistance screening under field conditions. The high throughput screening protocol (Sheat et al, 2022) we developed solved the main uncertainties associated with U/CBSV resistance assessment in cassava from uncontrolled and erratic infection processes in the field. When CBSD x CMD crosses were tested (3.2), the screening started with CBSV infections of seedlings because we assumed that the parents, including COL 40, PER 221, and PER 353, would either be highly resistant or highly sensitive to CBSV.…”

“…Seedlings from crosses comprising U/CBSV resistant parents ( Table 3 ) were subjected to a stringent virus-resistance screening ( Sheat et al., 2022 ). Since the resistance status of both seedling parents was known, we modified our resistance discovery workflow ( Sheat et al., 2019 ) and adopted a more rapid virus screening process that would identify/confirm virus resistance/susceptibility in seedlings within a few weeks if this resistance phenotype was evident in F1 seedlings.…”

“…Seedlings that showed virus symptoms and tested positive with qRT-PCR ( Sheat et al., 2019 ) were eliminated from further testing. Seedlings that tested negative were grafted with scions of cassava plants that were mixed-infected with UCBSV and EACMV-UG (GenBank accessions UCBSV, MW961202; EACMV-UG OL44492, OL444943), as described in ( Sheat et al., 2022 ). All plants that passed cycle two and tested negative by qRT-PCR and qPCR for U/CBSV and EACMV entered the confirmation cycle, in which all steps of the workflow were repeated with a higher number of plants.…”

“…All experiments, including molecular testing, are described in detail ( Sheat et al., 2019 ). A graphical overview of this workflow and further descriptions can be found in ( Sheat et al., 2022 ).…”

“…The first generation of prototypes with CBSD x CMD immunity was selected from crosses with South American and African germplasm using our high-precision virus screening ( Sheat et al., 2022 ). Indeed, the limited number of resistant plants does not represent a diversity sufficient for breeding populations, but does provide the resistances for further breeding.…”