RESUMO
Some species of Alicyclobacillus spoil beverages by producing guaiacol. Current culture-based methods detect the presence of Alicyclobacillus spp. and a subsequent peroxidase assay determines if the isolate can produce guaiacol. However, these methods are time-consuming and can yield false negatives due to differences in growth optima among species. The purpose of this study was to compare a RT-PCR-based method, the GENE-UP® PRO ACB assay, to the IFU Method No. 12 Enumeration and Enrichment methods. Ten species of Alicyclobacillus were detected using the tested RT-PCR assay, while A. dauci and A. kakegewensis were not detected using either IFU protocol. Low concentrations (1-10, 10-100, and 100-1,000 CFU/10 mL) of A. acidoterrestris, A. suci, and A. acidocaldarius were tested in five matrices. The proportion of positive samples identified using the tested RT-PCR assay (62/84) or the IFU Enrichment protocol (62/84) did not differ significantly from the proportion of inoculated samples (63/84). However, the IFU Enumeration method (32/84) detected statistically fewer positives. Additionally, methods identifying guaiacol production were compared. The proportion of correctly identified guaiacol producers using the tested RT-PCR assay (51/63) was not significantly different than those identified using the 3 h Cosmo Bio assay (54/63). Finally, four commercial samples of orange juice and sucrose solution were tested. Alicyclobacillus spp. were identified in all four samples using the IFU Enrichment method and in two samples using the tested RT-PCR assay. However, Alicyclobacillus was not detected in any sample using the IFU Enumeration method. Overall, 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. Both the 3 h guaiacol bioassay and the tested RT-PCR assays consistently differentiated guaiacol-producing and nonproducing strains.
Assuntos
Alicyclobacillus , Guaiacol , Bebidas , Sucos de Frutas e VegetaisRESUMO
Six thermo-acidophilic, spore-forming strains were isolated from a variety of juice products and were characterized genetically and phenotypically. According to 16S rRNA and rpoB gene phylogenetic analyses and average nucleotide identity comparisons against the species demarcation cutoff at <95â%, these six strains were determined to represent three novel species of Alicyclobacillus. The isolates were designated FSL-W10-0018T, FSL-W10-0037, FSL-W10-0048, VF-FSL-W10-0049T, FSL-W10-0057 and FSL-W10-0059T. All six isolates were Gram-positive, motile, rod shaped, contained menaquinone 7 as the major respiratory quinone and had ω-cyclohexane C17â:â0 as a major fatty acid. They were all able to grow aerobically in a range of acidic and moderate thermal conditions. Only isolates FSL-W10-0048 and VF-FSL-W10-0049T were able to produce guaiacol. The following names are proposed for the three new species: Alicyclobacillus mali sp. nov. (type strain FSL-W10-0018T =DSM 112016T=NCIMB 15266T); Alicyclobacillus suci sp. nov (VF-FSL-W10-0049T=DSM 112017T=NCIMB 15265T); and Alicyclobacillus fructus sp. nov. (FSL-W10-0059T=DSM 112018T=NCIMB 15264T).
