This study focuses on the diversity of circular single-stranded DNA (ssDNA) viruses associated with Poaceae growing in the fynbos ecosystem and its neighbouring agroecosystems in the Cape floristic region of Southern Africa. We have developed a sequenceindependent approach that combined two methods, including a first step of rolling circle amplification (RCA) followed by a second step of whole genome amplification (WGA). We have first tested and validated this approach using controls from our laboratory, including 4 Geminiviridae (sugarcane/Sugarcane streak Egypt virus; Datura stramonium/Tomato leaf curl virus, Datura stramonium/Tomato yellow leaf curl virus and Euphorbia caputmedusae/ Euphorbia caput-medusae stunt virus) and 2 double-stranded DNA Caulimoviridae (banana/Banana streak Obino l'Ewai virus and Arabidopsis thaliana/Cauliflower mosaic virus). The RCA/WGA approach was successful for unambiguously detecting the 4 Geminiviridae tested but failed to detect both Caulimoviridae. In addition to the 4 expected Geminiviridae, 2 Mastrevirus (located within the sugarcane plant), two Circovirus (banana and turnip) and three mycoviruses (banana, turnip and Euphorbia caput-medusae) were detected by performing Blast searches. This approach was then used for detecting the presence of circular ssDNA viruses within 23 South African Poaceae. Again, several ssDNA viruses were detected, including Mastrevirus, Begomovirus, Circovirus, mycoviruses and unknown ssDNA viruses. This study confirms the presence of a wealth of ssDNA viral sequences within plants that blur the boundaries between previously well-defined groups and stresses the need to better understand the evolutionary history of circular ssDNA viruses and to revisit current taxonomic classification schemes (Rosario et al., 2012). (Texte intégral)