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Zymoseptoria tritici is the causal fungus of Septoria tritici blotch (STB), a globally important foliar disease of wheat. Infections by more than one strain of a pathogen, or co‐infections, are known to be the most prevalent form of infection in the field. The impact that co‐infection of Z. tritici has on disease severity for the plant host and its effect on plant defence are not fully understood. Here, isolates of Z. tritici were extracted from STB‐presenting leaves from UK sites. Disease assays comparing single‐isolate inocula to mixed inocula were carried out on three cultivars of wheat, Flame, Stigg and Longbow. Leaves infected with mixed inocula were found to show low disease symptoms, including lesion and pycnidial coverage, compared with those infected with single isolates. Growing mixed isolates together in liquid media did not reveal evidence of competition or decreased growth compared with single‐isolate inocula. Wheat infected with a mixture of at least two Z. tritici isolates (of differing aggressiveness) could induce Pr‐1 (Pathogenesis‐related 1) at a significantly higher level in distal uninfected leaves compared with distal leaves of plants infected with a single aggressive isolate. Expression of Pr‐1 in plants with mixed inocula was intermediate to plants infected with an aggressive isolate (lower expression) and plants infected with a less aggressive isolate (higher expression). This suggests that wheat undergoes systemic "sensitization" by the least aggressive Z. tritici isolate during co‐infection. [ABSTRACT FROM AUTHOR]