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Background Dengue fever is an important infectious disease in Guangzhou, China; previous studies on the effects of weather factors on the incidence of dengue fever did not consider the linearity of the associations. Methods This study evaluated the effects of daily mean temperature, relative humidity and rainfall on the incidence of dengue fever. A generalized additive model with splines smoothing function was performed to examine the effects of daily mean, minimum and maximum temperatures, relative humidity and rainfall on incidence of dengue fever during 2006–2014. Results Our analysis detected a non-linear effect of mean, minimum and maximum temperatures and relative humidity on dengue fever with the thresholds at 28 °C, 23 °C and 32 °C for daily mean, minimum and maximum temperatures, 76% for relative humidity, respectively. Below the thresholds, there was a significant positive effect, the excess risk in dengue fever for each 1 °C in the mean temperature at lag7–14 days was 10.21%, (95% CI: 6.62% to 13.92%), 7.10% (95% CI: 4.99%, 9.26%) for 1 °C increase in daily minimum temperature in lag 11 days, and 2.27% (95% CI: 0.84%, 3.72%) for 1 °C increase in daily maximum temperature in lag 10 days; and each 1% increase in relative humidity of lag7–14 days was associated with 1.95% (95% CI: 1.21% to 2.69%) in risk of dengue fever. Conclusions Future prevention and control measures and epidemiology studies on dengue fever should consider these weather factors based on their exposure-response relationship. [ABSTRACT FROM AUTHOR]