Dissolved Carbon Dioxide: The Lifespan of Staphylococcus aureus and Enterococcus faecalis in Bottled Carbonated Mineral Water

SIMPLE SUMMARY: The consumption of bottled mineral water has been rising in the last decades. During the water bottling process, the risk of bacterial contamination with pathogen bacteria is a well-known problem. In this study, the influence of different carbon dioxide concentrations, in mineral water bottles, on the growth of Staphylococcus aureus and Enterococcus faecalis was investigated. Inside an Austrian mineral water bottling factory, a sum of 160 glass bottles with different carbon dioxide concentrations (3.0 g/L, 5.5 g/L, and 7.0 g/L) were capped after artificial contamination, stored at room temperature (20 °C), kept away from light, and dried in the laboratory. Water samples from freshly opened bottles were filtrated and cultivated on agar media to determine the number of colony-forming units over a period of 30 days. After the 23rd day, no more cultivable bacteria were detected in the samples from the freshly opened bottles. This observation leads to the conclusion that carbonated mineral water is an inappropriate habitat for Staphylococcus aureus and Enterococcus faecalis. ABSTRACT: During the process of mineral water production, many possible contamination settings can influence the quality of bottled water. Microbial contamination can originate from different sources, for example, the ambient air, the bottles, the caps, and from the bottling machine itself. The aim of this study was to investigate the influence of three different carbon dioxide (CO(2)) concentrations (3.0 g/L, 5.5 g/L, and 7.0 g/L; 20 bottles each) in bottled mineral water on the bacterial growth of Staphylococcus aureus (S. aureus) and Enterococcus faecalis (Ent. faecalis). The examined mineral water was artificially contaminated before capping the bottles inside the factory. After a specific number of days, water samples were taken from freshly opened bottles and after filtration (100 mL), filters were placed on Columbia Agar with 5% Sheep blood to cultivate S. aureus and Slanetz and Bartley Agar to cultivate Ent. faecalis. The respective colony-forming units (CFU) were counted after incubation times ranging from 24 to 120 h. Colony-forming units of S. aureus were not detectable after the 16th and 27th day, whereas Ent. faecalis was not cultivable after the 5th and 13th day when stored inside the bottles. The investigation of the bottles that were stored open for a certain amount of time with CO(2) bubbling out showed only single colonies for S. aureus after the 5th day and no CFUs for Ent. faecalis after the 17th day. A reduction in the two investigated bacterial strains during storage in carbonated mineral water bottles means that a proper standardized disinfection and cleaning procedure, according to valid hygiene standards of industrial bottling machines, cannot be replaced by carbonation.