A laboratory study was conducted to evaluate the influence of cocultivation of toxigenic Fusarium (F.) and Alternaria (A.) fungi with respect to growth and mycotoxin production. Fusarium culmorum Fc13, Fusarium graminearum Fg23 and two Alternaria tenuissima isolates (At18 and At220) were simultaneously or consecutively co-incubated on wheat kernels in an in vitro test system. Fungal biomass was quantified by determining ergosterol content. Three Fusarium toxins (DON, NIV and ZON) and three Alternaria toxins (AOH, AME and ALT) were analysed by a newly developed HPLC/MS/MS method. In simultaneous cocultures, the fungal biomass was enhanced up to 460% compared with individual cultures; Alternaria toxins were considerably depressed down to <5%. Combining At18 and At220 with Fg23 inhibited the toxin production of both fungal partners. In contrast, Fc13 increased its DON and ZON production in competitive interaction with both A. strains. The interfungal competitive effects aid the understanding of the processes of competition of both fungi in natural environments and the involvement of mycotoxins as antifungal factors. Cocultivation significantly affects fungal growth and mycotoxin production of phytopathogenic Alternaria and Fusarium strains. The impact of mycotoxins on the interfungal competition is highlighted.