Optimization of inoculum production of Stemphylium botryosum for large-scale resistance screening of lentils

Abstract

BackgroundStemphylium blight incited by Stemphylium botryosum poses a significant threat to lentil crops worldwide, inducing severe defoliation and causing substantial yield losses in susceptible varieties under favorable conditions. While some moderate levels of resistance have been identified within lentil germplasm, a low number of resistant cultivars are available to farmers. Adding to the common constraints of resistance breeding, a notable challenge is generating a sufficient number of spores for large-scale screenings, which are essential for pinpointing additional sources of resistance for integration into breeding programs. Therefore, there is a pressing need to improve existing screening methods and tailor them for large-scale material selection. In pursuit of this objective, a protocol for the efficient production of fungal material has been adapted.ResultsOptimization of fungal material production was successfully achieved by comparing the use of fungal mycelia and spores. Spore production was found to be optimal when produced on solid V8-PDA(hi) medium, while liquid Richard’s medium was identified as superior for mycelium yield. Furthermore, a refined screening method was developed by evaluating the resistance of six lentil accessions to stemphylium blight. This assessment included the use of either fungal mycelia (at densities ranging from 1 to 5 g L− 1) or spores (with densities ranging from 5 × 104 to 2 × 105 conidia mL− 1) under three different relative humidity levels (from 50 to 100%). Both humidity levels and inoculum dose significantly influenced the final disease rating (DR) and the relative Area Under the Disease Progress Curve (rAUDPC). Differences among genotypes in final symptom severity (DR) became more pronounced after inoculation with inoculum densities of 5 g L− 1 of mycelium or of 105 and 2 × 105 conidia mL− 1 of spore under 100% relative humidity. Given the challenges associated with the large-scale production of S. botryosum spores, inoculations with 5 g L− 1 of mycelium is highly recommended as a practical alternative for conducting mass-scale screenings.ConclusionsThe findings from this study underscore the critical importance of maintaining high level of humidity during inoculation and disease progression development for accurately assessing resistance to stemphylium blight. The optimization of mycelial production for suspension inoculation emerges as a more reliable and efficient approach for conducting large-scale screening to assess germplasm resistance against stemphylium blight in lentil crops.