Simple SummaryTo investigate seroprevalence and identify the risk factors associated with foot-and-mouth disease (FMD) in Pakistan’s northern border region, 385 serum samples were compiled from 75 flocks/herds in seven sub-regions in this region, including sheep (n= 97), goats (n= 142), cattle (n= 60), and buffaloes (n= 86) and tested using an FMDV 3ABC-Mab-bELISA for detection of antibody to FMD virus non-structural protein. The results showed that an overall apparent seroprevalence in this region was 67.0% of the total analyzed samples, the seroprevalence in sheep, goats, cattle, and buffaloes were 51.5%, 71.8%, 58.3%, and 82.5%, respectively. From the different risk factors investigated, age, sex, species of animal, seasons, flock/herd size, farming methods, outbreak location, and nomadic animal movement in search of water and pasture from region to region were found to be significantly associated (p< 0.05) with the seroprevalence of FMD. FMD remains a significant disease of ruminants in Pakistan’s northern border region. Sero-epidemiology and identification of associated risk factors of FMD is essential to bind the consequence of FMD in the region.AbstractThe present cross-sectional survey was carried out to investigate the distribution and risk factors of FMD in Pakistan’s northern border regions. About 385 serum samples were compiled from small ruminants (239) and large ruminants (146) and tested using 3ABC-Mab-bELISA. An overall apparent seroprevalence of 67.0% was documented. The highest seroprevalence of 81.1% was reported in the Swat, followed by 76.6% in Mohmand, 72.7% in Gilgit, 65.6% in Shangla, 63.4% in Bajaur, 46.6% in Chitral and lowest 46.5% in Khyber region. Statistically significant variations in seroprevalence of 51.5%, 71.8%, 58.3%, and 74.4% were recorded in sheep, goats, cattle, and buffaloes, respectively. From the different risk factors investigated, age, sex, species of animal, seasons, flock/herd size, farming methods, outbreak location, and nomadic animal movement were found to be significantly associated (p< 0.05) with the seroprevalence of FMD. It was concluded that proper epidemiological study, risk-based FMD surveillance in small ruminants, vaccination strategy, control measures for transboundary animal movement, collaborations, and awareness programs need to be practiced in the study regions to investigate the newly circulating virus strains in large and small ruminants and associated factors for the wide seroprevalence to plan proper control policies to bound the consequence of FMD in the region.