Antifungal class I [beta]-1,3-glucanases are believed to be part of the constitutive and induced defenses of plants against fungal infection. Unexpectedly, mutants deficient in these enzymes generated by antisense transformation showed markedly reduced lesion size, lesion number, and virus yield in the local-lesion response of Havana 425 tobacco to tobacco mosaic virus (TMV) and of Nicotiana sylvestris to tobacco necrosis virus. These mutants also showed decreased severity of mosaic disease symptoms, delayed spread of symptoms, and reduced yield of virus in the susceptible response of N. sylvestris to TMV. The symptoms of disease in the responses of both plant species were positively correlated with [beta]-1,3-glucanase content in a series of independent transformants. Taken together, these results provide direct evidence that [beta]-1,3-glucanases function in viral pathogenesis. Callose, a substrate for [beta]-1,3-glucanase, acts as a physical barrier to the spread of virus. Callose deposition in and surrounding TMV-induced lesions was increased in the [beta]-1,3-glucanase-deficient, local-lesion Havana 425 host, suggesting as a working hypothesis that decreased susceptibility to virus resulted from increased deposition of callose in response to infection. Our results suggest novel means, based on antisense transformation with host genes, for protecting plants against viral infection. These observations also raise the intriguing possibility that viruses can use a defense response of the host against fungal infection[mdash]production of [beta]-1,3-glucanases[mdash]to promote their own replication and spread.