3.1. Detection of the known type II FHB resistance QTL/gene inYM18, ZM9, YM16, and NM13
To find out whether four parents and YM 20 possess the known major type II FHB resistance QTL/genes, they were surveyed with the gene-specific marker of Fhb1, and flanking markers of Fhb2, using WSB, SM3 and AN 8455 as the references. Target band polymorphism of the Fhb1 and Fhb2 markers was revealed between WSB, SM3, YM18, ZM9, YM16, NM13, YM20 and AN 8455, indicating that YM18, NM13 possess Fhb1, while ZM9, YM16 and AN8455 do not possess Fhb1. Four parents and YM 20 all do not possess Fhb2 (Fig. 2).
3.2. Validation of KASP markers
The screening results of the effects of Fhb1, QFhb.yaas-2DL and QFhb.yaas-3BL were presented in Table S1. The results showed that the FHB resistance of cultivars/lines carrying QFhb.yaas-2DL or QFhb.yaas-3BL were significantly (P<0.05) higher than those carrying none of the genes; but the effects of QFhb.yaas-2DL or QFhb.yaas-3BL were significantly (P<0.05) lower than those of Fhb1 (Table S1, Fig. 3). The FHB severity of accessions with QFhb.yaas-3BL alone was the highest among all QTL/gene combination, and those harboring Fhb1, QFhb.yaas-2DL and QFhb.yaas-3BL had the lowest average FHB severity (Table S1, Fig. 3).
The validation of KASP markers in the 77 cultivars/lines (6 from Hubei, and 71 from Jiangsu) showed that YM18 carried Fhb1 alone, NM13 carried Fhb1 plus QFhb.yaas-3BL, ZM9 and YM20 carried QFhb.yaas-2DL and QFhb.yaas-3BL, SM3 carried Fhb1 plus QFhb.yaas-2DL and QFhb.yaas-3BL, whereas AN 8455 did not carry any of the above resistance QTL (Table S1). Nineteen accessions from Jiangsu carried Fhb1, while 3 accessions from Hubei and 49 accessions from Jiangsu carried QFhb.yaas-2DL, 3 accessions from Hubei and 37 cultivars/lines from Jiangsu carried QFhb.yaas-3BL. All accessions from Hubei did not carry Fhb1. The genotype result indicated that the KASP markers for Fhb1, QFhb.yaas-2DL and QFhb.yaas-3BL were consistent with previous report (Su et al. 2018; Zhu et al 2018; Zhu et al 2020; Zhu et al 2021)
3.3. Development of lines by agronomic trait evaluation and FHB marker-assisted selection
Based on the results of self-crossing and performance of several traits such as PH, HT, FT, NSP, and seed infection similar or superior to YM20 had been selected and grown continuously since F2 generation (Fig. 1). 32, 43 and 13 plots of three F5 populations were conducted for genotyping and investigating agronomic traits and yield, respectively. The genotypic results, FHB resistance status, agronomic characters and yield of three F5 populations were depicted in Fig. 4 and Table S2–S3.
3.4. FHB resistance with different QTL combinations in three RIL populations
The FHB resistance condition and QTL constitution of the selected F5 lines were presented in Table S3. For YM18/ZM9 and YM18/YM16 populations, as expected, lines harboring all three QTL had the highest FHB resistance with an average PSS of 6.43% and 7.59%, respectively, which were similar to SM 3 (Table 2, Table S1, S3). The average PSS of the lines carrying Fhb1 plus QFhb.yaas-2DL and Fhb1 plus QFhb.yaas-3BL were both significantly lower (P <0.05) than those carrying QFhb.yaas-2DL plus QFhb.yaas-3BL in these two populations, respectively. Lines carrying QFhb.yaas-2DL alone had the higher FHB severity (P <0.05) than those carrying QFhb.yaas-3BL alone in these two populations, respectively. Lines harboring none of the resistance QTL had the highest FHB severity (P<0.05) in these two populations, respectively. For YM18/ZM9 population, lines carrying Fhb1 alone had significantly higher FHB resistance (P <0.05) than those carrying QFhb.yaas-2DL alone or QFhb.yaas-3BL alone. For YM18/ YM 16 population, lines carrying Fhb1 alone had similar FHB resistance to those carrying QFhb.yaas-2DL alone plus QFhb.yaas-3BL, and slightly more resistance than lines carrying QFhb.yaas-2DL alone. In YM18/NM13 population, the average PSS of the lines carrying Fhb1 alone was 29.36%, and lines carrying Fhb1 plus QFhb.yaas-3BL had significantly lower FHB severity with an average PSS of 20.89% (P <0.05). None of the combinations had similar FHB resistance to SM3 in this population (P <0.05) (Table 2).
3.5. The effects of FHB QTL on yield-related traits and yield
The performances of agronomic traits and yield of the selected lines were assessed and exhibited in Table S2–S3. For YM18/ZM9 and YM18/YM16 populations, lines carrying Fhb1 alone had the lowest NKS (P <0.05). Lines carrying QFhb.yaas-2DL alone, QFhb.yaas-3BL alone, and QFhb.yaas-3BL plus QFhb.yaas-2DL had similar NKS, while higher than those carrying none of the resistance QTL, Fhb1 plus QFhb.yaas-3BL, Fhb1 plus QFhb.yaas-2DL and all three resistance QTL. For YM18/ZM9 populations, no significant difference in NSP were observed among the lines carrying different QTL combinations except those carrying none of the resistance QTL and QFhb.yaas-2DL alone. Lines carrying QFhb.yaas-2DL alone had the highest NSP (P <0.05), while those carrying none of the resistance QTL had the lowest NSP (P <0.05). For YM18/YM16 population, lines carrying QFhb.yaas-2DL alone, QFhb.yaas-3BL alone and Fhb1 plus QFhb.yaas-2DL had the highest NSP (P <0.05). Lines carrying Fhb1 plus QFhb.yaas-3BL, QFhb.yaas-3BL plus QFhb.yaas-2DL and all three resistance QTL had similar NSP, while higher than those carrying Fhb1 alone. Lines carrying none of the resistance QTL had the lowest NSP (P <0.05) compared with others. For YM18/ZM9 population, lines carrying QFhb.yaas-3BL alone had the highest TKW (P <0.05) compared to others. Lines carrying Fhb1 plus QFhb.yaas-3BL, QFhb.yaas-3BL plus QFhb.yaas-2DL and all three resistance QTL had slightly higher TKW than those carrying none of the resistance QTL, Fhb1 alone, QFhb.yaas-2DL alone and Fhb1 plus QFhb.yaas-2DL. For YM18/YM16 population, lines carrying QFhb.yaas-2DL alone, QFhb.yaas-3BL alone, Fhb1 plus QFhb.yaas-3BL, QFhb.yaas-3BL plus QFhb.yaas-2DL and all three resistance QTL had similar TKW, and all with the highest TKW (P <0.05) compared to others. Lines carrying Fhb1 plus QFhb.yaas-2DL had slightly higher TKW than those carrying none of the resistance QTL and Fhb1 alone. For YM18/ZM9 and YM18/YM16 populations, lines carrying none of the resistance QTL and Fhb1 alone had the lowest yield (P <0.05) compared with others. No significant difference in yield were observed among other lines with different QTL combinations (Table 3–4). For YM18/NM13 population, no significant difference in yield and its related traits was observed among the lines carrying different QTL combinations (Table 5).
3.6. Development of lines with similar FHB resistance to SM3
In F5 populations, six lines, YM18/ZM9-5, YM18/ZM9-12, YM18/YM16-15, YM18/YM16-18, YM18/YM16-22 and YM18/YM16-24 carrying three resistance QTL, had similar FHB resistance to SM3 (Fig. 5–6). YM18/ZM9-12, YM18/YM16-18 and YM18/YM16-22 showed consistently superior yield (2586.22g/plot–2639.58g/plot) to YM20 (2397.07g/plot–2450.49g/plot) while growing in two environments for evaluating the type II FHB resistance and yield in 2021. The result showed that the three lines had superior yield to YM20 and similar FHB resistance to SM3 in both YZ and GY (Table 6).