Key message <p>Two novel QTLs conferring Fusarium stalk rot resistance were identified in the Kemater flint landrace population, thereby broadening the genetic base for FSR resistance breeding. Given the polygenic inheritance of Fusarium stalk rot resistance, genomic selection could be explored as a potential strategy for improving resistance to stalk rot in maize.</p> Abstract <p>Fusarium stalk rot (FSR) is one of the most prevalent and destructive soil-borne diseases of maize, caused by <i>Fusarium graminearum</i> (teleomorph <i>Gibberella zeae</i>), which leads to substantial yield losses worldwide. This study investigated the genetic basis of FSR resistance in the Austrian “Kemater Landmais Gelb” (KE) population under field conditions. A total of 180 KE testcrosses were generated by crossing 180 randomly chosen KE doubled-haploid lines with a dent tester line. These testcrosses were evaluated following artificial inoculation with <i>F. graminearum</i> using needle injection, and disease severity was assessed based on internode proportion, a robust phenotyping scale for FSR. Field trials were conducted using an alpha-lattice design with two replications across two locations over two years. Moderate-to-high broad-sense heritability estimates (<i>H</i>² = 0.48–0.82) were realized for FSR severity, plant height, and days to silking. Low–moderate phenotypic (<i>r</i> =  − 0.17 to -0.22) and genotypic correlations (<i>r</i>_g = − 0.22 to − 0.31) were found between FSR severity and agronomic traits. Strong linkage disequilibrium (LD) was observed between the QTL, ZmSYNBREED_65065_837, associated with days to silking and all QTLs for plant height (<i>r</i> <InlineEquation ID="IEq1"> <EquationSource Format="TEX">\(\sim 0.85\)</EquationSource> <EquationSource Format="MATHML"><math> <mrow> <mo>∼</mo> <mn>0.85</mn> </mrow> </math></EquationSource> </InlineEquation>). This high LD could explain the observed correlations between these traits. Genome-wide association analysis in the KE testcross population identified two QTLs that jointly explained 21% of the genotypic variance in FSR severity. Several putative candidate genes located within these QTL regions were associated with pathogen recognition, signal transduction, and defense responses to FSR infection. Once the stability and functionality of these QTLs and candidate genes are established, these loci will provide valuable insights into the molecular mechanisms underlying FSR resistance and serve as useful targets for genomic selection in maize breeding programs.</p>