Sisal (Agave sisalana Perrine ex Engelm) is cultivated in large areas of the Brazilian semi-arid Northeastern region. This crop allows thousands of people to obtain minimal wages. However, a disease known as bole rot, caused by Aspergillus species, is causing a decline in production. The objective of this study was to investigate the involvement of hydrolytic enzymes (cellulase, cutinase, protease and lipase) and ochratoxin A in an in vitro simulation of the interaction between A. niger Tiegh. and sisal to understand the mechanisms of fungal pathogenicity. Primers for cutinase lipase, protease, ochratoxin A, and elongation factor 1-α (EF), with this last one used as endogenous control were designed and optimized for real time quantitative PCR (qPCR) with the SYBR Green methodology. The genes potentially involved in the pathogenicity of A. niger showed higher expression in medium supplemented with sisal as compared to the minimal medium. The primers reported herein will allow detailed studies of the biological events leading to bole rot disease in sisal. These results represent the first data on genes putatively involved in the pathogenicity of A. niger to sisal.