Isolation and Identification of Guaiacol-Producing Alicyclobacillus Fastidiosus Strains from Orchards in Poland

Abstract

The genus Alicyclobacillus comprises a group of Grampositive, endospore-forming, and thermo-acidophilic bacteria. Some of the Alicyclobacillus species are categorized as spoilage bacteria due to their ability to produce off-flavor compounds (e.g. guaiacol and halophenols) that adversely affect the taste and aroma of beverages. In our study, Alicyclobacillus species isolated from Polish orchard soils and fruits were subjected to 16S rDNA sequencing and rpoB gene RFLP analysis. The results showed that the isolated strains belong to the species A. acidoterrestris and A. fastidiosus. Additionally, a fragment of the vdc operon, which is responsible for guaiacol production, was amplified and subjected to RFLP analysis. The resulting RFLP patterns allowed classifying five strains as A. acidoterrestris type II, while the patterns of another three strains showed no similarity to the patterns of any reference strains used for comparison. The reference strain A. fastidiosus DSM 17978 did not give any PCR product in this reaction, nor showed guaiacol production. All the three isolated strains of A. fastidiosus were subjected to a series of tests, including determination of the biochemical profile using API 50CH tests and determination of the ability to produce guaiacol at 20°C, 25°C, and 45°C. All A. fastidiosus strains exhibited similar morphological and biochemical properties as the strain described in the literature. However, our strains, unlike the strain described previously, were able to produce guaiacol at all tested temperatures, and could therefore be included in the group of spoilage species. To date, this study is the first to report the isolation of A. fastidiosus from Poland and is the second such report in the world.