Food spoilage is a major issue leading to waste, lost revenue, and consumer dissatisfaction. Beverages are known to be spoiled by Alicyclobacillus spp., which are extremophilic sporeformers that cannot be killed by traditional pasteurization methods. The spoilage metabolite guaiacol is only produced by a subset of Alicyclobacillus spp. under specific beverage storage conditions, making precise assessment of spoilage risk a priority for beverage and ingredient producers. In Chapter 1, whole genome sequencing and an ANIb cutoff of 95% was used to establish three new species of Alicyclobacillus. One of these, Alicyclobacillus suci, was shown to be a guaiacol producer highly related to Alicyclobacillus acidoterrestris. In Chapter 2, we aimed to uncover genomic determinants of guaiacol production across the genus. Using a genome-wide association study, several of the genes significantly enriched in guaiacol-producing Alicyclobacillus spp. were found to be associated with oxidative stress response. When looking at guaiacol production using a peroxidase colorimetric assay, we found that strains of A. suci produced significantly (p<0.01) more guaiacol on average in media than did strains of A. acidoterrestris. Additionally, A. suci and A. fastidiosus genomes each had duplicate copies of vdcC, while only a single copy of vdcC was found in the genomes of A. acidoterrestris, A.acidiphilus, and A. herbarius. The levels of guaiacol produced by A. acidoterrestris and A. suci were investigated further in Chapter 3, but this time in apple juice under industry-relevant storage conditions. A. suci and A. acidoterrestris both produced more guaiacol at 25°C than at 30°C or 40°C, even though growth rate was slower at the lower temperatures. After 14 days, the highest levels of guaiacol in the pH trials were produced at pH 4.0 (A. acidoterrestris) and pH 3.0 (A. suci). Chapter 4 explored ways of assessing Alicyclobacillus contamination in beverages by comparing a newly-developed RT-PCR-based method, the GENE-UP® PRO ACB assay, to the IFU Method No. 12 Enumeration and Enrichment methods. This study showed consistent detection of Alicyclobacillus spp. using either the IFU Enrichment protocol or the tested RT-PCR assay, which both outperformed the IFU Enumeration protocol. This study validated a faster, novel assay which can be utilized by producers to make better decisions about spoilage risk.