peer reviewed ; In the context of increasing occurrences of toxic cyanobacterial blooms worldwide, their monitoring in Belgium is currently performed by regional environmental agencies (in two of three regions) using different protocols and is restricted to some selected recreational ponds and lakes. Therefore, a global assessment based on the comparison of existing datasets is not possible. For this study, 79 water samples from a monitoring of five lakes in Wallonia and occasional blooms in Flanders and Brussels, including a canal, were analyzed. A Liquid Chromatography with tandem mass spectrometry (LC-MS/MS) method allowed to detect and quantify eight microcystin congeners. The mcyE gene was detected using PCR, while dominant cyanobacterial species were identified using 16S RNA amplification and direct sequencing. The cyanobacterial diversity for two water samples was characterized with amplicon sequencing. Microcystins were detected above limit of quantification (LOQ) in 68 water samples, and the World Health Organization (WHO) recommended guideline value for microcystins in recreational water (24 µg L−1) was surpassed in 18 samples. The microcystin concentrations ranged from 0.11 µg L−1 to 2798.81 µg L−1 total microcystin. For 45 samples, the dominance of the genera Microcystis sp., Dolichospermum sp., Aphanizomenon sp., Cyanobium/Synechococcus sp., Planktothrix sp., Romeria sp., Cyanodictyon sp., and Phormidium sp. was shown. Moreover, the mcyE gene was detected in 75.71% of all the water samples.