Department of Food Science and Nutrition, Faculty of the Environment, University of the Aegean, Mitropoliti Ioakeim 2, Myrina 81400, Lemnos Island, Greece.
Veterinary Research Institute of Athens, Greek Agricultural Organization "Demeter", Neapoleos 25, Aghia Paraskeui 15310, Greece.
Int J Food Microbiol. 2014 Feb 3;171:1-7. doi: 10.1016/j.ijfoodmicro.2013.11.013. Epub 2013 Nov 19.
This study aimed to evaluate the possible impact of acid adaptation of Listeria monocytogenes cells on their attachment to stainless steel (SS) during long-term incubation under either low or moderate temperature conditions and on the subsequent recalcitrance of attached cells to lethal acid treatments. Initially, nonadapted or acid-adapted stationary phase L. monocytogenes cells were used to inoculate (ca. 10⁸ CFU/ml) brain-heart infusion (BHI) broth in test tubes containing vertically placed SS coupons. Incubation was carried out at either 5 or 30 °C for up to 15 days, under static conditions. On the 5th, 10th and 15th days of incubation, attached cells were subjected to lethal acid treatments by exposing them, for either 6 or 60 min, to pH 2, adjusted with either hydrochloric or lactic acid. Following the acid treatments, remaining viable cells were detached (through strong vortexing with glass beads) and enumerated by agar plating, and also indirectly quantified by conductance measurements via their metabolic activity. Results obtained from both quantification techniques, employed here in parallel, revealed that although the numbers of attached cells for nonadapted and acid-adapted ones were similar, the latter were found to present significantly (p<0.05) increased recalcitrance to all the acid treatments for both incubation temperatures and all sampling days. In addition and regardless of acid adaptation, when long (60 min) acid treatments were applied, conductance measurements revealed that the weak organic lactic acid exhibited significantly (p<0.05) stronger antilisterial activity compared to the strong inorganic hydrochloric acid (at the same pH value of 2). To conclude, present results show that acid adaptation of L. monocytogenes cells during their planktonic growth is conserved even after 15 days of incubation under both low and moderate temperature conditions, and results in the increased recalcitrance of their sessile population to otherwise lethal acid treatments. This "stress hardening" should be severely taken into account when acidic decontamination interventions are used to kill attached to equipment surfaces cells of this important pathogenic bacterium.
本研究旨在评估李斯特菌细胞在低温或中温条件下长期孵育过程中酸适应对其附着在不锈钢（SS）上的可能影响，以及随后附着细胞对致死酸处理的顽固性。最初，使用非适应或酸适应的静止期李斯特菌细胞接种（约 10⁸ CFU/ml）脑心浸液（BHI）肉汤于含有垂直放置的 SS 优惠券的试管中。在静态条件下，于 5 或 30°C 下孵育长达 15 天。在孵育的第 5、10 和 15 天，通过将其暴露于 pH 2 的盐酸或乳酸调整的酸处理 6 或 60 分钟，使附着的细胞经受致死酸处理。酸处理后，通过强玻璃珠涡旋将剩余的存活细胞分离（detached）并通过琼脂平板计数，并且通过其代谢活性的电导率测量间接定量。此处平行采用的两种定量技术获得的结果表明，尽管未适应和适应酸的附着细胞数量相似，但发现后者在两种孵育温度和所有采样日的所有酸处理中均表现出明显（p<0.05）更高的顽固性。此外，无论酸适应如何，当应用长（60 分钟）酸处理时，电导率测量表明，与强无机盐酸（在相同的 pH 值 2 下）相比，弱有机乳酸表现出明显（p<0.05）更强的抗李斯特菌活性。总之，目前的结果表明，李斯特菌细胞在浮游生长过程中的酸适应即使在低温和中温条件下孵育 15 天后也得以保留，并导致其固定群体对其他致死酸处理的顽固性增加。在使用酸性去污干预措施杀死附着在设备表面上的这种重要致病菌细胞时，应严重考虑这种“应激硬化”。
