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Lassa fever is a viral hemorrhagic fever caused by the Lassa virus LASV, which was first isolated in the rodent <i>Mastomys natalensis</i> in 1974 in Kenema, Sierra Leone. As little is known about the abundance and the presence of LASV in rodents living in the Bo area, we carried out a small mammal longitudinal population survey. A standardized trapping session was performed in various habitats and seasons in six villages over two years (2014–2016) and samples collected were tested for arenavirus IgG and LASV. A Bayesian phylogenetic analysis was performed on sequences identified by PCR. A total of 1490 small mammals were collected, and 16 rodent species were identified, with <i>M. natalensis</i> (355, 24%) found to be the most prevalent species. Forty-one (2.8%) samples were IgG positive, and 31 of these were trapped in homes and 10 in surrounding vegetation. Twenty-nine of 41 seropositive rodents were <i>M. natalensis.</i> We detected four LASV by PCR in two villages, all found in <i>M. natalensis</i>. Phylogenetic analysis showed that the sequences were distributed within the Sierra Leonean clade within lineage IV, distinguishing a Bo sub-clade older than a Kenema sub-clade. Compared to other settings, we found a low abundance of <i>M. natalensis</i> and a low circulation of LASV in rodents in villages around Bo district.