Specific primers for detection of Alicyclobacillus acidoterrestris by RT-PCR.

The reverse transcription-polymerase chain reaction (RT-PCR), after a short enrichment culture, was used to detect Alicyclobacillus acidoterrestris in acidic beverages. Two specific primers were selected from the region of V2 and V4 on 16S rRNA gene. With this primer set, 294-bp fragments from A. acidoterrestris could be amplified. The detection limit of the RT-PCR with the FHLP filters was about 10-1 fg of pure total RNA per reaction. Juice samples inoculated with 10(4) cfu of A. acidoterrestris per ml were RT-PCR positive without enrichment. However, after 15 h of enrichment, the samples inoculated with 2-3 cfu ml-1 were positive. This RT-PCR culture assay would enable rapid and specific detection of strains of A. acidoterrestris in acidic beverages.

Isolation and identification of Alicyclobacillus acidoterrestris from acidic beverages.

Thermophilic bacteria isolated from spoiled acidic beverages were identified on the basis of DNA-DNA homology and 16S ribosomal DNA sequence similarity to Alicyclobacillus acidoterrestris. This result suggested that the isolates were a new type of spoilage bacterium in acidic beverages in Japan.