Leishmania spp. are kinetoplastid protozoa that cause zoonotic disease in humans and animals. No large-scale cross-sectional survey has been performed to study canine leishmaniasis and its spatial distribution in Israel. We aimed to survey the prevalence and geographical distribution of Leishmania infection in dogs in Israel. Paired blood and sera were collected from domestic dogs at private clinics and animal shelters throughout Israel. Leishmania infection was determined by ELISA serology and blood qPCR targeting the kinetoplast minicircle (kDNA) and the ribosomal ITS1 region. Altogether, 1,392 paired samples were tested from dogs from all parts of Israel. Thirty nine of 1,392 (2.8%) dogs were seropositive to Leishmania. Leishmania spp. infection was detected in 35/1392 (2.5%) and 10/1392 (0.7%) dogs by kDNA and ITS1 qPCR, respectively. All ITS1 PCR-positive dogs were confirmed to be infected by Leishmania infantum after DNA sequencing. Seventy four of 1,392 (5.3 %) dogs were either seropositive or PCR-positive.ITS1 PCR-positive dogs were 20 times more likely to be seropositive than negative dogs (p < 0.001) while kDNA PCR-positive dogs were 5.1 times more likely (p = 0.003). Dogs with clinical signs suggestive of leishmaniasis had significantly higher prevalence of seropositivity (7.5%) compared to dogs without signs (2.2%; p= 0.001).This survey of Leishmania infection in dogs covering the entire region of Israel revealed widespread infection in the northern, central and mid-southern regions of Israel with dominance of sub-clinical infection. The risk of Leishmania infection warrants protection against vector sand fly bites and disease screening among dogs throughout Israel.