Two protected indigenous freshwater crayfish species, the noble crayfish Astacus astacus (Linnaeus, 1758) and the stone crayfish Austropotamobius torrentium (Schrank, 1803), inhabit waterbodies of the Plitvice Lakes National Park.. Freshwater crayfish are keystone species in freshwater ecosystems, but their numbers aredeclining. One of the main threats for indigenous crayfish species is the pathogen Aphanomyces astaci (Schikora, 1906) (Oomycetes), the causative of agent crayfish plague disease. Presence of this pathogen has already been recorded in the Plitvice Lakes but the previous research was done on a small sample. Thus, the main goal of this research was to monitor the presence of the pathogen A. astaci on a large number of individuals and at multiple locations within the National Park. Research was conducted at four locations. Astacus astacus was sampled at Burgeti (68 individuals) and Prštavci (10), and A. torrentium at streams Prijeboj (22) and Sartuk (15). New, non-invasive method for A. astaci detection was applied. The method is based on collecting swabs of mixed epibiotic microbial communities present on the crayfish carapax. The DNA was isolated and the presence of the pathogen's DNA was detected by PCR. Pathogen was detected only in nine noble crayfish specimens (12%, 9/78) from Burgeti and Prštavci. Positive samples were classified into semi-quantitative categories of pathogen load by quantitative PCR. Microsatellite genotyping was applied to identyfy A. astaci Group A strain (As genotype) which is consistent with the results of the previou sresearch. Obtained results are applicable in the development of effective management plans aiming to protect vulnerable native species in NP.