Bacterial Stress Response Group, Microbiology, Ryan Institute, School of Biological & Chemical Sciences, University of Galway, Galway H91 TK33, Ireland.
Teagasc Food Research Centre, Moorepark, Fermoy, Cork, Ireland.
Int J Food Microbiol. 2023 Aug 16;399:110238. doi: 10.1016/j.ijfoodmicro.2023.110238. Epub 2023 May 2.
Listeria monocytogenes is a foodborne pathogen that is characterized by its ability to withstand mild stresses (i.e. cold, acid, salt) often encountered in food products or food processing environments. In the previous phenotypic and genotypic characterization of a collection of L. monocytogenes strains, we have identified one strain 1381, originally obtained from EURL-lm, as acid sensitive (reduced survival at pH 2.3) and extremely acid intolerant (no growth at pH 4.9, which supports the growth of most strains). In this study, we investigated the cause of acid intolerance in strain 1381 by isolating and sequencing reversion mutants that were capable of growth at low pH (pH 4.8) to a similar extent as another strain (1380) from the same MLST clonal complex (CC2). Whole genome sequencing showed that a truncation in mntH, which encodes a homologue of an NRAMP (Natural Resistance-Associated Macrophage Protein) type Mn transporter, is responsible for the acid intolerance phenotype observed in strain 1381. However, the mntH truncation alone was not sufficient to explain the acid sensitivity of strain 1381 at lethal pH values as strain 1381R1 (a mntH revertant) exhibited similar acid survival to its parental strain at pH 2.3. Further growth experiments demonstrated that Mn (but not Fe, Zn, Cu, Ca, or Mg) supplementation fully rescues the growth of strain 1381 under low pH conditions, suggesting that a Mn limitation is the likely cause of growth arrest in the mntH background. Consistent with the important role of Mn in the acid stress response was the finding that mntH and mntB (both encoding Mn transporters) had higher transcription levels following exposure to mild acid stress (pH 5). Taken together, these results provide evidence that MntH-mediated Mn uptake is essential for the growth of L. monocytogenes under low pH conditions. Moreover, since strain 1381 was recommended for conducting food challenge studies by the European Union Reference Laboratory, the use of this strain in evaluating the growth of L. monocytogenes in low pH environments where Mn is scarce should be reconsidered. Furthermore, since it is unknown when strain 1381 acquired the mntH frameshift mutation, the ability of the strains used for challenge studies to grow under food-related stresses needs to be routinely validated.
李斯特菌单核细胞增生李斯特菌是一种食源性病原体，其特点是能够耐受食品产品或食品加工环境中经常遇到的温和压力（即低温、酸性、盐分）。在对李斯特菌单核细胞增生李斯特菌菌株的先前表型和基因型特征分析中，我们发现一株最初从 EURL-lm 获得的 1381 株菌对酸敏感（在 pH 2.3 下存活能力降低）且对酸极不耐受（在 pH 4.9 下无法生长，而大多数菌株在此 pH 下能够生长）。在这项研究中，我们通过分离和测序能够在低 pH（pH 4.8）下生长到与另一株菌（1380）相似程度的回复突变体，研究了 1381 株菌不耐酸的原因。该菌来自同一 MLST 克隆群（CC2）。全基因组测序表明，mntH 发生截断，该基因编码一种 NRAMP（天然抗性相关巨噬细胞蛋白）型 Mn 转运体同源物，导致 1381 株菌出现观察到的不耐酸表型。然而，mntH 截断本身不足以解释 1381 株菌在致死 pH 值下的酸敏感性，因为 1381R1（mntH 回复突变体）在 pH 2.3 下的酸存活能力与亲本菌株相似。进一步的生长实验表明，Mn（而不是 Fe、Zn、Cu、Ca 或 Mg）补充完全挽救了 mntH 背景下菌株 1381 在低 pH 条件下的生长，这表明 Mn 限制可能是导致生长停滞的原因。mntH 和 mntB（均编码 Mn 转运体）在暴露于温和酸性应激（pH 5）后转录水平升高，这与 Mn 在酸应激反应中的重要作用一致。总之，这些结果表明 MntH 介导的 Mn 摄取对于李斯特菌单核细胞增生李斯特菌在低 pH 条件下的生长是必不可少的。此外，由于欧盟参考实验室推荐使用 1381 株菌进行食品挑战研究，因此应该重新考虑在缺乏 Mn 的低 pH 环境中使用该菌株来评估李斯特菌单核细胞增生李斯特菌的生长情况。此外，由于目前尚不清楚 1381 株菌何时获得 mntH 移码突变，因此需要定期验证用于挑战研究的菌株在与食物相关的应激条件下的生长能力。
