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Canine vector-borne diseases (CVBD) are an important and emerging health concern for humans and animals worldwide. The purpose of the presented study was to assess, from 2016 to 2019, the seroprevalence of CVBD agents and clarify the epidemiology of tick-borne disease in stray dogs living in the Campania Region, Southern Italy. For this purpose, blood samples were collected from January 2016 to December 2019 from 1023 dogs in authorized kennels located in the five municipalities of the Campania Region. SNAP^® 4DX^® from IDEXX^® Laboratories was used for detection of <i>Ehrlichia</i> spp., <i>Anaplasma</i> spp., <i>Borrelia burgdorferi sensu lato (s.l.),</i> and <i>Dirofilaria immitis</i> antibodies. The overall seroprevalence of CVBD in stray dogs was 19.6% (95% Confidence Intervals (CI): 17.2–22.8%; 201/1023). The most common pathogen was <i>Ehrlichia</i> spp., with a percentage of positivity of 16.03%, followed by <i>Anaplasma</i> spp. with 7.8%. <i>B. burgdorferi s.l.</i> and <i>D. immitis</i> were detected in only 0.2% of dogs; co-infection was detected in 4.5% of stray dogs tested. No link was detected between the gender, age, location, and CVBD seropositivity, except for <i>Ehrlichia</i> spp. for which location (Avellino Province; <i>p</i> = 0.007) and gender (male, <i>p</i> = 0.002) were risk factors for seropositivity. Our results demonstrated that animals are exposed to at least one of the four etiological agents (<i>Ehrlichia</i> spp., <i>Anaplasma</i> spp. <i>Borrelia burgdorferi s.l.</i>, and <i>Dirofilaria immitis</i>) transmitted by vectors. Finally, this study highlighted the utility of serological monitoring in stray dogs, housed in kennels, given the threat posed by CVBD to animals and the zoonotic implications of these etiological agents and their vectors on human health.