The commercial potential of high pressure and thermal processing (HPTP) was investigated against Alicyclobacillus acidoterrestris spores in commercial acidic apple juice beverage and in acidified and neutral potassium buffers. With starting spore counts prior to treatments being 6.5 and 7.2 log10 respectively for strains AJA 66 (D90°C 15.4 min) and ATCC 49025 (D90°C 8.5 min), HPTP at 600 MPa at 80 °C for 3 min provided an optimal treatment with spore viability reduced below the detection limit for both strains. HPTP at 80 °C for 1 min and HPTP at 70 °C for 3 min achieved 4.1–4.5 log10 CFU/mL reduction. HPTP at 70 °C for 1 min reduced the number of viable spores by 2.0–2.5-log10 CFU/mL. Flow cytometry revealed the presence of membrane compromised spores among culturable spores following HPTP and heat alone treatments at different temperatures. Electron microscopy clearly showed the efficiency of HPTP with crushed or hollow spores predominating after treatments. No correlation between HPTP susceptibility and genetic diversity was observed for two genotypes of A. acidoterrestris spores. The treatment combination provides a promising option for industrial utility since it requires lower heat and processing time.