<p>Time-reversal and space-inversion symmetries are fundamental properties of crystals and play a role in underlying phenomena such as magnetism, topology and non-trivial spin textures. Transition metal dichalcogenides (TMDs) represent an excellent tunable model system to explore the interplay between these symmetries as they can be engineered on demand by tuning the number of layers and via all-optical bandgap modulation. In this work, we modulate and study time-reversal symmetry in mono- and bilayer TMDs with all-optical methods using third-harmonic Faraday rotation. We excite the samples using elliptically polarized light, achieve spin-selective bandgap modulation and consequent breaking of time-reversal symmetry. The reduced symmetry modifies the nonlinear susceptibility tensor, causing a rotation of the emitted third-harmonic polarization. With this method, we probe broken time-reversal symmetry in both non-centrosymmetric (monolayer) and centrosymmetric (bilayer) WS₂ crystals. Furthermore, we discuss how the detected third-harmonic rotation angle directly links to spin-valley locking in monolayer TMDs and spin-valley-layer locking in bilayer TMDs. Our results show a powerful approach to study broken time-reversal symmetry in crystals regardless of space-inversion symmetry, and shed light on the spin, valley and layer coupling of atomically thin semiconductors.</p>