Natural fine-grained suspensions usually exhibit a complex rheological fingerprint – in particular a two-step yielding phenomenon – due to the presence of mineral clay particles and organic matter (often found in a flocculated state). These rheological properties may vary considerably from one location to another due to the differences in mud composition (specifically in organic matter content). In this study, the origin of this two-step yielding behaviour for natural suspensions is discussed with the help of different experimental techniques including rheology, particle sizing, rheo-optics, and video microscopy. The samples were collected from different locations in the Port of Hamburg, Germany. A rheological analysis of the samples was performed with amplitude sweep, frequency sweep, stress ramp-up and structural recovery tests. The shear-induced structural changes of mud samples was studied by using a parallel plate shearing device with a microscope. Mineral clay-organic matter flocs were studied using video microscopy to obtain the floc size, floc density and settling velocity of flocs. Higher values of rheological properties such as cross-over stress, yield stress, and moduli were observed for samples having higher organic matter content. These samples also produced the largest floc sizes. The rheo-optical analysis showed the formation of cylinder-like structures in fine-grained suspensions upon shearing action, which reflect the origin of two-step yielding behaviour in mud samples, observed in stress ramp-up and amplitude sweep tests.