real-time polymerase chain reaction-based method for rapid and specific detection of spoilage Alicyclobacillus spp. in apple juice

Aims: To develop a real‐time PCR‐based rapid detection method for spoilage Alicyclobacillus spp. in juice products. Methods and Results: The squalene‐hopene cyclase‐encoding gene was targeted for primer‐and‐probe development. Gene fragments from representative strains were cloned, and PCR primers and probe were designed by DNA sequence comparison. Selected bacteria were examined for cross‐reactivity by the new method. Cells were serially diluted in apple juice and saline, and examined by the new method to establish detection sensitivity. Using the newly developed Taqman® real‐time PCR‐based method, strains of Alicyclobacillus acidocaldarius and A. acidoterrestris were detected without cross reactivity with other common food‐borne micro‐organisms. Detection of