Survival & detection of the faecal indicator bacterium Enterococcus faecalis in water stored in traditional vessels

Background and objectives: Enterococci are used as indicators of faecal pollution and are typically detected using agar-based growth media incubated under standard aerobic conditions. However, such conditions may not be fully effective in enumerating injured bacteria, due to their sensitivity to reactive oxygen species (ROS) under aerobic conditions. Investigations were carried out to assess the extent of sub-lethal damage and ROS-sensitivity on different strains of Enterococcus faecalis in water stored in traditional brass and earthern vessels by enumerating the bacteria under standard aerobic conditions and under conditions designed to neutralise the effect of ROS. Methods: Pure cultures of E. faecalis were maintained for up to 48 h in brass and earthern vessels and enumerated on various selective and non-selective media either under (i) standard aerobic conditions, (ii) aerobic conditions in a growth medium supplemented with the peroxide scavenger sodium pyruvate, (iii) anaerobic conditions using unsupplemented medium and (iv) anaerobic conditions in a growth medium supplemented with sodium pyruvate, the latter being regarded as ROS-neutralised conditions. Results: Counts of E. faecalis decreased substantially after storage for 12 h in water kept in the brass vessel but not in the earthern vessel. However, the decrease in counts depended upon the growth medium and the conditions used for enumeration, with a non-selective medium giving the highest count under ROS-neutralised conditions. While the counts obtained on various selective media were also enhanced under ROS-neutralised conditions, they remained lower than those of the non-selective medium. Interpretation and conclusions: growth conditions where reactive oxygen species are neutralised are effective in enhancing the colony count of stressed E. faecalis, irrespective of the type of medium used for enumeration.