This research article published by Research square, 2021Background: Rift Valley fever virus (RVFV) is a zoonotic arbovirus of public health impact infecting livestock, wildlife, and humans mainly in Africa and other parts of the world. Despite its public health importance, mechanisms of RVFV maintenance during inter-epidemic (IEPS) periods and potentially spread to new areas remain unclear.We aimed to comparatively examine exposure to RVFV and RVFV infection among humans, goats and mosquitoes in an agro-pastoral community in Lower Moshi area of Moshi rural district. Results:Results show that the male gender was related to RVFV seropositivity (χ2 = 5.351; p=0.030). Being 50 years and above was related to seropositivity (χ2=14.430; p=0.006) whereas bed net use, larger numbers of persons living in the same house (>7 persons) and RVFV seropositivity in goats were related to higher seropositivity to RVFV among humans (χ2=6.003; p=0.021, χ2=23.213; p=0.000 and 27.053; p=0.000), respectively.RVFV antibody concentrations were only marginally higher in humans without statistically significant difference [t (112) =0.526; p=0.60)]. By the use of RT-qPCR, goats exhibited the highest RVFV infection rate of 4.1%, followed by humans (2.6%), Aedes spp(2.3%), and Culex spp(1.5%). Conclusions: In the absence of RVFV infection data in areas nearby the study site, our findings suggest Lower Moshi area as a potential hotspot for RVF, posing the danger of being a source of RVFV spread to other areas. Goats had the highest infection rate, suggesting goats as important hosts in the virus maintenance during IEPs. We recommend the design and implementation of strategies that will warrant effective active surveillance of RVF through the identification of RVF hotspots for targeted control of RVF.