Members of group III histidine kinases from different filamentous fungi were previously shown to mediate osmoregulation and resistance to dicarboximide, phenylpyrrole and, aromatic hydrocarbon fungicides. In this study, we report the disruption of the gene encoding group III histidine kinase, AlHK1, in the economically important plant pathogen Alternaria longipes. The AlHK1 gene disruption had pleiotropic effects on this fungus. Besides the expected osmosensitivity and fungicides resistance, AlHK1 participated in the spore production process. In addition, the Delta AlHK1 strains had stronger aggressive ability to infect their host plant than that of their parental strain, the wild-type strain C-00, suggested that AlHK1 was involved in the pathogenicity of A. longipes and performed in this function by a negative manner. This is the first report to our knowledge.