Grapevines from 13 vineyards in Pakistan were surveyed for the prevalence of several pathogens. Using RT-qPCR, 257 samples were tested for 19 viruses, phytoplasmas and Xylella fastidiosa. Prevalent viruses were: grapevine virus A (GVA, 47.8%), grapevine leafroll-associated virus 2 (GLRaV-2, 37.3%), grapevine rupestris stem pitting-associated virus (GRSPaV, 36.1%), and grapevine fleck virus (GFkV, 35%). Other viruses detected were: grapevine leafroll-associated virus 1 (GLRaV-1, 2.3%), grapevine leafroll-associated virus 2RG (GLRaV-2RG, 5%), grapevine leafroll-associated virus 3 (GLRaV-3, 7%), grapevine leafroll-associated virus 4 (GLRaV-4) and its strains (5, 6, and Pr, 16.6%), grapevine leafroll-associated virus 7 (GLRaV-7, 4.2%), grapevine fanleaf virus (GFLV, 11.6%), grapevine virus B (GVB, 4.2%), grapevine virus D (GVD, 0.7%), grapevine virus E (GVE, 1.1%), and grapevine Pinot gris virus (GPGV, 1.9%). Mixed infections were detected in 75.9% of samples. Pathogens tested for, but not detected include GLRaV-4 strains 9 and Car, grapevine red blotch virus (GRBV), tomato ringspot virus (ToRSV), tobacco ringspot virus (TRSV), Arabis mosaic virus (ArMV), grapevine virus F (GVF), phytoplasmas and X. fastidiosa. Additionally, 16 samples were analyzed by high-throughput sequencing (HTS) to confirm RT-qPCR results. In this paper we present an extensive survey for grapevine pathogens and thus the first report of GVA, GVB, GVD, GVE, GRSPaV, GFkV, GLRaV-1, GLRaV-2, GLRaV-2RG, GLRaV-3, GLRaV-4, GLRaV-4 strains 5, 6, and Pr, and GLRaV-7 in Pakistan.