Influence of pH on bile sensitivity amongst various strains of Listeria monocytogenes under aerobic and anaerobic conditions

Abstract

Listeria monocytogenes is a dangerous bacterium that causes the food-borne disease listeriosis and accounts for nearly 20% of food-borne deaths. This organism can survive the body's natural defences within the digestive tract, including acidic conditions and bile. Although the bile response has been analysed, limited information is available concerning the ability of L. monocytogenes to resist bile under anaerobic conditions, especially at acidic pH, which mimics conditions within the duodenum. Additionally, it is not known how the bile response varies between serotypes. In this study, the survival of strains representing six serotypes was analysed under aerobic and anaerobic conditions following exposure to bile. Exposure to bile salts at acidic pH increased toxicity of bile, resulting in a significant reduction in survival for all strains tested. However, following this initial reduction, no significant reduction was observed for an additional 2 h except for strain 10403S (P = 0.002). Anaerobic cultivation increased bile resistance, but a significant increase was only observed in virulent strains when exposed to bile at pH 5.5. Exposure to pH 3.0 prior to bile decreased viability amongst avirulent strains in bile in acidic conditions; oxygen availability did not influence viability. Together, the data suggested that being able to sense and respond to oxygen availability may influence the expression of stress response mechanisms, and this response may correspond to disease outcome. Further research is needed on additional strains to determine how L. monocytogenes senses and responds to oxygen and how this varies between invasive and non-invasive strains.

Fig. 1.

Viability of L. monocytogenes in porcine bile extract at pH 7.5 and 5.5. L. monocytogenes F2365, ScottA, 2011L-2636, 2011L-2676, 10403S, 15313, HCC23, HCC7, LO28 and EGDe were grown to mid exponential phase prior to treatment for 3 h in the following conditions: TSB pH 7.5 with either 0 or 1 % bile, or TSB pH 5.5 with either 0 or 1 % bile. Percent viability was calculated at 1 (black), 2 (grey) and 3 h (white) post-exposure to bile in relation to TSB-only controls at the respective pH. Results represent mean ± sd of three independent experiments. *P < 0.05; **P < 0.001.

Fig. 2.

Influence of pH on toxicity of GDCA and TDCA. L. monocytogenes F2365, ScottA, 2011L-2636, 2011L-2676, 10403S, 15313, HCC23, HCC7, LO28 and EGDe were cultured in TSB at pH 7.5 (black), 6.5 (grey), 5.5 (hatched) or 4.5 (white). Cultures were grown for 16 h in media supplemented with 1 % porcine bile extract, 5 mM GDCA or 5 mM TDCA. Viability was assessed by plate counts and per cent survival was determined in relation to TSB-only controls at the respective pH. Results represent mean ± sd of three independent experiments. *P < 0.05; **P < 0.001.

Fig. 3.

Influence of anaerobic cultivation on bile resistance of L. monocytogenes. L. monocytogenes F2365, ScottA, 2011L-2636, 2011L-2676, 10403S, 15313, HCC23, HCC7, LO28 and EGDe were cultured in media supplemented with either 0 or 1 % bile under either aerobic (black) or anaerobic (grey) conditions. Percent survivals were determined in relation to media-only controls at the respective pH. Results represent mean ± sd of three independent experiments. *P < 0.05; **P < 0.001.

Fig. 4.

Viability of L. monocytogenes in bile when pre-exposed to acidic conditions. L. monocytogenes F2365, ScottA, 2011L-2636, 2011L-2676, 10403S, 15313, HCC23, HCC7, LO28 and EGDe were grown to late exponential phase, exposed to TSB at pH 7.5 or 3.0 for 1 h and then cultured in media at pH 7.5 or 5.5 supplemented with either 0 or 1 % bile for 16 h. Per cent survivals were determined in relation to media-only controls at the respective pH. Aerobic cultures exposed to pH 7.5 media, followed by exposure to bile (black); aerobic cultures exposed to pH 3.0 media, followed by exposure to bile (grey); anaerobic cultures exposed to pH 7.5 media, followed by exposure to bile (hatched); anaerobic cultures exposed to pH 3.0 media, followed by exposure to bile (white). Results represent mean ± sd of three independent experiments. *P < 0.05; **P < 0.001; ***P < 0.0001.