AbstractThe highly invasive Campuloclinium macrocephalum (Asteraceae; ‘pompom weed’), native to South America, was introduced to South Africa where it has flourished and now threatens the native diversity of the grasslands it has invaded. The Argentine C. macrocephalum population is known to comprise at least three ploidy levels, and exhibit aneuploidy. Ploidy levels had not yet been assessed across South African populations of pompom weed and could influence the effectiveness of a biological control (biocontrol) programme. Therefore, we used flow cytometry to estimate ploidy levels of individuals from invaded regions in South Africa and Argentina. Diploids, triploids, tetraploids and aneuploids were detected in the sampled Argentine individuals. In South Africa, only triploid and tetraploid cytotypes were identified, with possible aneuploid variation. Mapping the distribution of these ploidy levels in South Africa revealed that the tetraploids are more widespread across all five sampled provinces, whereas the triploids occur only in Gauteng (the likely origin of the invasion) and neighbouring North West provinces and one sampled area in KwaZulu‐Natal province. Distinct phenotypic variation is evident between Argentine individuals of different ploidy levels. Polyploids have now been confirmed to occur in both the native and introduced ranges. Because triploids and tetraploids are present in both native and introduced ranges, we predict that polyploidy is not likely to be a constraint in effective biocontrol in South Africa as the natural enemies have probably co‐evolved with polyploids in the native range. However, other factors may impact the effectiveness of the biocontrol agents, such as feeding, oviposition (potentially affecting their reproduction), establishment, and spread, therefore, warranting further research.