Man's oldest fungicide has probably long functioned in this role in plants, as a natural component of induced antifungal defence. Elemental sulphur (S0) is the only inorganic phytoalexin and the only phytoalexin produced by so many different taxa. S0 (detected by GC-MS as 32S8) is produced in representative species of Sterculiaceae (cocoa), Solanaceae (tomato, tobacco), Malvaceae (cotton), and Leguminosae (French bean) in response to xylem-invading fungal and bacterial pathogens. Production was more rapid and intensive in disease-resistant genotypes. Gene expression for S0 production may be xylem-specific as S0 was not present in leaves of six species undergoing hypersensitivity to Pseudomonas syringae. Anomalously, high constitutive S0 levels occurred in leaves of Arabidopsis and Brassica oleracea. S0 was highly toxic (ED50 1–3 μg ml−1) to many fungal pathogens representing ascomycetes, basidiomycetes, and deuteromycetes, but not to an oomycete, Phytophthora, or to bacteria. Levels in cocoa and tomato xylem and Arabidopsis leaves were potentially inhibitory, but in other interactions were below theoretically toxic concentrations. However, S0 accumulation is highly localized, suggesting that the element is produced in sufficient amounts, at the right time and place to be effective. SEM-EDX revealed S in tomato and cocoa xylem walls, xylem parenchyma, and vascular gels and tyloses, all sites appropriate to counter vascular pathogenic Verticillium dahliae. Transient increases in sulphate, glutathione and cysteine occurred in tomato xylem. The sulphate may reflect the over-expression of sulphate transporters, but the thiols might be possible precursors. Analysis of differential gene expression should reveal what may be a novel biosynthetic pathway of S0 formation in eukaryotes.