Viruses can adversely affect vines' vitality, longevity, quality, and quantity in single or mixed infections. In this work, to detect single and mixed viral infections of grapevine in two central viticulture regions of Azerbaijan, a total of 166 samples showing virus-like symptoms including yellows, reddening, fan leaf, mosaic, leaf rolling, and shortening of internodes were collected during the growing seasons of 2022-2023 from different vineyards and subjected to ELISA and RT-PCR analyses. Plant samples were initially assayed using the one-step assay AgriStrip based on lateral flow immunochromatography and DAS-ELISA with specific antisera for Grapevine fanleaf virus (GFLV), Grapevine leafroll-associated virus 1 (GLRaV-1), Grapevine leafroll-associated virus 2 (GLRaV-2), Grapevine leafroll-associated virus 3 (GLRaV-3), and Grapevine leafroll-associated virus 4 (GLRaV-4). Following serological tests, 65% (108 out of 166) of the examined samples exhibited a positive result for one or more of the specified viruses. Total RNA was extracted, and synthesized cDNA was used to amplify a part of the coat protein (CP) gene in RT-PCR using specific primers. Amplicons of the expected size of 1560 bp, 401 bp, 600 bp, 942 bp, 500 bp, and 272 bp were amplified for the mentioned viruses, respectively (but not from healthy samples). The result of this study can be used in the management strategies of these viruses in grapevine-producing areas of Azerbaijan.