This paper presents the development of specific primers for the detection of Fusarium oxysporum f. sp. vasinfectum (FOV) race 4. Primers were designed from the sequences of AFLP markers unique to race 4 and screened against a collection of California FOV isolates. Characterization of disease development caused by FOV race 4 was tested at different inoculum densities, ranging from 10^1 to 10^6 spores per gram of soil. Results for two Upland cotton varieties, Phytogen 72 and Ultima EF, and three Pima cotton varieties, Delta Pine 340, Delta Pine 744, and Phytogen 800, suggest an inoculum threshold around 10^4 spores per gram of soil for disease development.