Kupang City is an endemic area for mosquito vector-based diseases. The incidence of mosquito vector-based diseases has increased since 2017 with 32 cases per 100,000 population increasing in 2019 to 156 per 100,000 population and as of April 2020 there were 610 cases reported. The environment is one of the determinants of the incidence of mosquito vector-based diseases, both in the form of the physical, biological and social environment. Climate is classified in the physical environment that influences the pattern of mosquito vector-based disease. Certain climatic conditions can increase the risk of disease transmission. This study was carried out with the aim of knowing the effect of the variability of climatic factors in the form of air temperature, humidity and wind speed on the incidence of mosquito vector-based diseases in Kupang City in 2020. Analysis of the effect was carried out using secondary data, namely monthly mosquito vector-based disease data obtained from Eleven health centers in Kupang City and climate factors for the same period were obtained from the BMKG Climatology Station of Kupang City. Test the effect using multiple linear regression analysis partially (t test) and simultaneously (F test). The results of the t-test showed that air temperature and wind speed had no significant effect (significance = 0.714 and 0.889 > 0.05) while air humidity had a positive effect on mosquito vector-based diseases (significance = 0.001 < 0.05). Simultaneously, the F test showed that humidity had an effect on mosquito vector-based diseases (significance = 0.000 < 0.05).