Fecal matter is considered to be one of the primary sources of water pollution. Understanding the aggregation behavior of the fecal pigments (FPs) could play a critical role in their detection and analysis. This work shows that in aqueous media, the fluorescence of FPs indicates the presence of multiple emitting species, which have been assigned to monomers, lower-order H-aggregates (dimers), and higher-order H-aggregates. Steady-state absorbance, fluorescence and time-resolved fluorescence decay studies conclude that the emission of FPs in aqueous medium indicates H-type of aggregation, even up to nanomolar and sub-nanomolar concentrations. Four sets of independent experiments involving the variation of (i) concentration of FPs, (ii) temperature, (iii) pH, and (iv) ethanol/water composition as solvent media suggest the presence of monomer (540 nm), dimer (516 nm), and higher-order aggregates (500 nm) of FPs in aqueous solutions. The dimeric FP species appear to be present in the entire concentration range of 1 pM to 1 μM. Fluorescence lifetimes of H-aggregates are relatively longer as compared to the corresponding monomers. Hydrogen bonding appears to play an important role in forming H-aggregates in the aqueous phase of FPs as observed in the IR spectra of the FPs in dichloromethane. Density functional theory (DFT) calculations using the B3LYP functional and the LANL2DZ basis set show the contributions of π-π stacking and hydrogen-bonding interactions toward the formation of H-aggregated dimer of FPs in aqueous media.