A rhodamine-based organic–inorganic hybrid solid fluorescent chemosensor by covalently immobilized R6G derivative on core–shell structured mesoporous silica microspheres with perpendicularly aligned mesopore channels has been prepared. The fluorescent responses of the prepared chemosensor to Hg2+ have been investigated, and the results demonstrated that the proposed hybrid solid fluorescent chemosensor featured a high affinity Hg2+-specific fluorescence response in water by considering the highly dense modification of the rhodamine probe. The detection limit for Hg2+ is 0.1 nM (S/N = 3, which is well below the guideline value given by the World Health Organization) under optimized conditions. Excellent wide linear range (1.0–100.0 nM) and good repeatability (relative standard deviation of 3.2%) were obtained for Hg2+. The proposed chemosensor also exhibited excellent selectivity for Hg2+ over competing environmentally relevant metal ions, and can be used in a wide pH span and regenerated readily. These values, particularly the high sensitivity and excellent selectivity in contrast to the values reported previously in the area of fluorescent Hg2+ detection, demonstrated the analytical performance of the proposed chemosensor could be used for efficient determination of Hg2+ in natural water samples. Toward the purpose of practical application, the proposed chemosensor was further used to determine Hg2+ in real environmental water samples.