In recent decades, the incidence of Lyme borreliosis (LB) in Europe seems to have increased, underpinning a growing public health concern. LB surveillance systems across the continent are heterogeneous, and the spatial and temporal patterns of LB reports have been little documented. In this study, we explored the spatio-temporal patterns of LB cases reported in France from 2016 to 2019, to describe high-risk clusters and generate hypotheses on their occurrence. The space-time K-function and the Kulldorf's scan statistic were implemented separately for each year to evaluate space-time interaction between reported cases and searching clusters. The results show that the main spatial clusters, of radius size up to 97 km, were reported in central and northeastern France each year. In 2017-2019, spatial clusters were also identified in more southern areas (near the Alps and the Mediterranean coast). Spatio-temporal clustering occurred between May and August, over one-month to three-month windows in 2016-2017 and in 2018-2019. A strong spatio-temporal interaction was identified in 2018 within 16 km and seven days, suggesting a potential local and intense pathogen transmission process. Ongoing improved surveillance and accounting for animal hosts, vectors, meteorological factors and human behaviors are keys to further elucidate LB spatio-temporal patterns.