Date palm (Phoenix dactylifera L.) is considered the main crop in deserts and arid areas such as Saudi Arabia. Fifteen species belonging to 7 fungal genera were isolated from date palm rhizosphere soil at Riyadh, Saudi Arabia. Twenty-one isolates of Aspergillus spp. were used in producing silver nanoparticles (SNPs), five of A. flavus, nine of A. niger and seven of A. terreus. Synthesis of SNPs by these fungi is emerging as an important branch of nanotechnology due to it'secofriendly, safe and cost-effective nature. SNPs have been characterized by UV-Visible Spectrophotometer and Transmission Electron Microscope (TEM). In order to increase the yield of biosynthesized SNPs of desired shape and size, it is necessary to control the cultural and physical parameters during the synthesis. We reported the optimum synthesis of SNPs on a liquid medium at 1.5mM of silver nitrate, pH 9 and 26°C after 96 hours. Antifungal activity of SNPs colloids has indicated that the highest inhibition zone was detected with SNPs. In the case of SNPs synthesized by A. terreus PNU37, the highest Inhibition percentage (IP %) 67.6% at the concentration 150 ppm of SNPs. Results have also indicated that the SNPs synthesized by A. flavus PNU05 at a concentration of 150 ppm/100 ml culture medium gave the highest reduction of B1 determined by HPLC, where the percentages of reduction (PR%) was 56.45%.ISSR analysis revealed a high level of genetic diversity in the Aspergillus spp. population and useful for genetic characterization. ISSR markers were not suitable to discriminate between producing and non-producing SNPs isolates. There was no clear-cut relationship between the ISSR markers (genotype of isolates), antifungal and anti-aflatoxigenic properties.