Understanding the relaxation mechanisms of photoexcited charge carriers in two-dimensional materials is indispensable from the fundamental point of view and for future optoelectronic applications. Through the photoconductivity and electronic transport experiments, we probe the mechanisms behind the persistent photoconductivity (PPC) in monolayer molybdenum disulfide (MoS2). The temperature (T) and power-dependent photoresponse studies reveal that the relaxation of excited charge carriers is strongly affected by the random fluctuations of local potentials. The relaxation time (τ) increases from τ ≃ 12 s at T = 16.5 K to τ ≃ 1235 s at T = 297 K, indicating PPC is a high T phenomenon in monolayer MoS2. The transport measurements demonstrate that the defect states with the density ≃4.43 × 1014 eV−1 cm–2 in a low gate voltage regime, originating from the sulfur vacancies, are responsible for these fluctuations. With a rise in temperature, the defect states undergo a transition from localization to extended states at T ≥ 100 K and thereby form the percolation network, which profoundly influences the relaxation mechanism. Our meticulous experiments and quantitative analysis provide newer insight into the origin of PPC in monolayer MoS2.