Research Article 2019Background Leptospirosis is a bacterial zoonotic disease of worldwide importance, though relatively neglected in many African countries including sub Saharan Africa that is among areas with high burden of this disease. The disease is often mistaken for other febrile illnesses such as dengue, malaria, rickettsioses and enteric fever. Leptospirosis is an occupational disease likely to affect people working in environments prone to infestation with rodents which are the primary reservoir hosts of this disease. Some of the populations at risk include: sugar- cane plantation workers, wetland farmers, fishermen and abattoir workers. In this study we investigated the prevalence of antibodies against Leptospira among sugarcane plantation and factory workers, fishing communities as well as among rodents and shrews in domestic and peridomestic environments within the study areas. Methods The study was conducted in Kagera region, northwestern Tanzania and it involved sugar- cane plantation workers (cutters and weeders), sugar factory workers and the fishing com- munity at Kagera Sugar Company in Missenyi district and Musira island in Lake Victoria, Kagera, respectively. Blood was collected from consenting human adults, and from rodents and shrews (insectivores) captured live using Sherman traps. Serological detection of lepto- spiral antibodies in blood serum was carried out by the microscopic agglutination test (MAT). Results A total of 455 participants were recruited from the sugarcane plantation (n = 401) and fishing community (n = 54) while 31 rodents and shrews were captured. The overall prevalence of antibodies against Leptospira in human was 15.8%. Sugarcane cutters had higher seroprev- alence than other sugar factory workers. Prevalent antibodies against Leptospira serovars in humans were against serovars Lora (6.8%), Sokoine (5.3%), Pomona (2.4%), Hebdoma- dis (1.1%) and Kenya (0.2%). Detected leptospiral serovars in reservoir hosts were Sokoine (12.5%) and Grippotyphosa (4.2%). Serovar Sokoine was detected both in humans and small mammals. Conclusion Leptospirosis is a public health threat affecting populations at risk, such as sugarcane plan- tation workers and fishing communities. Public awareness targeting risk occupational groups is much needed for mitigation of leptospirosis in the study areas and other vulnerable populations in Tanzania and elsewhere.