Foot-and-mouth disease virus (FMDV) circulates as multiple serotypes and strains in many endemic regions. In particular the three Southern African Territories (SAT) serotypes are maintained effectively in their wildlife reservoir, the African buffalo, and individuals may harbour multiple SAT-serotypes for extended periods in the pharyngeal region. However the exact site and mechanism for persistence remain unclear. FMD in buffaloes offers a unique opportunity to study FMDV-persistence, as transmission from carrier ruminants has only convincingly been demonstrated for this species. Following co-infection of naïve African buffaloes with three SAT-serotypes isolated from field buffaloes; palatine tonsil swabs were the sample of choice for recovering infectious FMDV up to 400 days post infection (dpi). Post-mortem examination identified infectious virus for up to 185 dpi and viral genome up to 400 dpi in lymphoid tissue of the head and neck, mainly focussed in germinal centres. Interestingly viral persistence in vivo was not homogenous and the SAT-1 isolate persisted for longer than SAT-2 and SAT-3. Co-infection and passage of these SAT isolates in goat and buffalo cell lines demonstrated a direct correlation between persistence and cell killing capacity. These data suggest FMDV persistence occurs in the germinal centres of lymphoid tissue but the duration of persistence is related to virus replication and cell killing capacity.Importance Foot-and-mouth disease virus (FMDV) causes a highly contagious acute vesicular disease in domestic livestock and wildlife species. African buffalo (Syncerus caffer) are the primary carrier host of FMDV in African savannah ecosystems, where the disease is endemic. We have shown the virus persists for up to 400 days in buffaloes and there is competition between viruses during mixed infections. There was similar competition in cell culture; virus that killed quickly persisted more efficiently in passaged cell cultures. These results may provide a mechanism for the dominance of particular viruses in an ecosystem.