We have analysed DNA from African cassava mosaic virus (ACMV) infected Nicotlana benthamlana by twodimensional agarose gel electrophoresls and detected ACMV-speclflc DNAs by blot-hybridisation. ACMV DNA forms Including the previously characterised singlestranded, open-circular, linear and supercoiled DNAs along with five previously uncharacterised heterogeneous DNAs (H1 - H5) were resolved. The heterogeneous DNAs were characterised by their chromatographlc properties on BND-cellulose and their ability to hybridise to strand-specific and doublestranded probes. The data suggest a rolling circle mechanism of DNA replication, based on the sizes and strand specificity of the heterogeneous single-stranded DNA forms and their electrophoretic properties in relation to genome length single-stranded DNAs. Second-strand synthesis on a single-stranded virussense template is evident from the position of heterogeneous subgenomic complementary-sense DNA (H3) associated with genome-length virus-sense template (VT) DNA. The position of heterogeneous virus-sense DNA (H5), ranging in size from one to two genome lengths, Is consistent with its association with genome-length complementary-sense template (CT) DNA, reflecting virus-sense strand displacement during replication from a double-stranded intermediate. The absence of subgenomic complementary-sense DNA associated with the displaced virus-sense strand suggests that replication proceeds via an obligate single-stranded intermediate. The other species of heterogeneous DNAs comprised concatemeric singlestranded virus-sense DNA (H4), and double-stranded or partially single-stranded DNA (H1 and H2).