This study was conducted to determine the spatiotemporal distribution of foot-and-mouth disease (FMD) virus (FMDV) serotypes and evaluate the awareness of livestock keepers about FMD in Tanzania. An observational prospective study involving serological analysis, FMDV antigen detection and questionnaire survey was carried out in the lake zone of Tanzania. Seroprevalence of antibodies to the nonstructural protein 3ABC of FMDV and serotype-specific antigen detection were investigated by using SVANOVIR^® FMDV 3ABC-Ab ELISA and indirect-sandwich ELISA (sELISA), respectively, whilst a structured questionnaire was used to evaluate the awareness of livestock keepers about FMD. During the period of 2010–2011, both serum and tissue (foot-and-mouth epithelia) samples were collected from cattle suspected of FMD in 13 districts of the four regions of the lake zone. A total of 107 (80.5%) out of 133 tested serum samples were seropositive to nonstructural protein 3ABC, with at least one sample being positive from all 10 districts screened. Fifteen (53.6%) out of 28 tissue epithelial samples collected from FMD cases in eight districts during the course of this study were positive to serotype O FMDV antigen. Of these eight districts, serotype O FMDV antigens were detected from seven districts and no other serotypes were recovered from animal samples screened. Questionnaire surveys in six districts indicated that livestock keepers in the lake zone were aware of the clinical manifestations (26/29 = 90.0%) and economic impact (23/29 = 79.0%) of FMD in the region. The questionnaire data showed that FMD outbreaks often occurred after rainy seasons (22/29 = 75.9%), with the highest peaks predominantly occurring just after the long rains in May and June, and at the end of the short rains in November and December of each year. The spatial distribution of the FMD cases suggested that serotype O virus exposure was the only widespread cause of the 2010–2011 outbreaks in the lake zone.