BACKGROUND: Aphis gossypii is an important pest of cotton that has developed resistance to many chemicals used for its control. Any lack of understanding of its genetic structure, resistance status and host plant specialisation hampers effective management. RESULTS: Eight microsatellite markers were genotyped for a collection of Australian A. gossypii field isolates from 55 plant species from major Australian cotton-producing regions. The aphid’s pirimicarb resistance status linked to the ACE1 (acetylcholinesterase) S431F mutationwas determined by PCR-RFLP.Overall, the genetic diversity was low and there were only 13 multilocus genotype (MLG) groups found in a total of 936 aphids, suggesting asexual reproduction. Three MLGs (Aust-01, Aust-02 and Aust-04) represented 78% of all aphids tested. MLGs Aust-01 (41%) and Aust-02 (18%) were linked to the ACE1 S431F mutation and found on cotton and a range of hosts. Aust-04 (19%) hosted mainly on cotton (but also Asteraceae and Malvaceae) was predominantly susceptible to pirimicarb. Given their abundance and widespread occurrence, these three clones were considered to be superclones. CONCLUSION: The study demonstrated that any strategy to control A. gossypii andmanage pirimicarb resistance should target A. gossypii strains of all MLG types residing on any plant species and not just cotton.