Fused deposition modelling (FDM) is the most widely used additive manufacturing process in customised and low-volume production industries due to its safe, fast, effective operation, freedom of customisation, and cost-effectiveness. Many different thermoplastic polymer materials are used in FDM. Acrylonitrile butadiene styrene (ABS) is one of the most commonly used plastics owing to its low cost, high strength and temperature resistance. The fabricated FDM ABS parts commonly work under thermo-mechanical loads in actual practice. For producing FDM ABS components that show high fatigue performance, the 3D printing parameters must be effectively optimized. Hence, this study evaluated the bending fatigue performance for FDM ABS beams under different thermo-mechanical loading conditions with varying printing parameters, including building orientations, nozzle size, and layer thickness. The combination of three building orientations (0°, ±45°, and 90°), three nozzle sizes (0.4, 0.6, and 0.8 mm) and three-layer thicknesses (0.05, 0.1, and 0.15 mm) were tested at different environmental temperatures ranging from 50 to 70 °C. The study attempted to find the optimal combination of the printing parameters to achieve the best fatigue behaviour of the FDM ABS specimen. The experiential results showed that the specimen with 0° building orientation, 0.8 mm filament width, and 0.15 mm layer thickness vibrated for the longest time before the fracture at each temperature. Both a larger nozzle size and thicker layer height can increase the fatigue life. It was concluded that printing defects significantly decreased the fatigue life of the 3D-printed ABS beam.