Considering the well-documented impacts of land-use change on water resources and the rapid land-use conversions occurring throughout Africa, in this study, we conducted a spatiotemporal analysis of surface water quality and its relation with the land use and land cover (LULC) pattern in Mokopane, Limpopo province of South Africa. Various physico-chemical parameters were analyzed for surface water samples collected from five sampling locations from 2016 to 2020. Time-series analysis of key surface water quality parameters was performed to identify the essential hydrological processes governing water quality. The analyzed water quality data were also used to calculate the heavy metal pollution index (HPI), heavy metal evaluation index (HEI) and weighted water quality index (WQI). Also, the spatial trend of water quality is compared with LULC changes from 2015 to 2020. Results revealed that the concentration of most of the physico-chemical parameters in the water samples was beyond the World Health Organization (WHO) adopted permissible limit, except for a few parameters in some locations. Based on the calculated values of HPI and HEI, water quality samples were categorized as low to moderately polluted water bodies, whereas all water samples fell under the poor category (>100) and beyond based on the calculated WQI. Looking precisely at the water quality’s temporal trend, it is found that most of the sampling shows a deteriorating trend from 2016 to 2019. However, the year 2020 shows a slightly improving trend on water quality, which can be justified by lowering human activities during the lockdown period imposed by COVID-19. Land use has a significant relationship with surface water quality, and it was evident that built-up land had a more significant negative impact on water quality than the other land use classes. Both natural processes (rock weathering) and anthropogenic activities (wastewater discharge, industrial activities etc.) were found to be playing a vital role in water quality evolution. This study suggests that continuous assessment and monitoring of the spatial and temporal variability of water quality in Limpopo is important to control pollution and health safety in the future.