Listeria monocytogenes causes the severe foodborne illness listeriosis and survives in food-associated environments due to its high stress tolerance. A data assembly and analysis protocol for microbial growth experiments was compiled to elucidate the strain variability of L. monocytogenes stress tolerance. The protocol includes 1) measurement of growth ability under stress; 2) selection of a suitable method for growth parameter calculation; 3) comparison of growth patterns between strains; and 4) biological interpretation of the discovered differences. In step 1, L. monocytogenes strains (n = 388) of various serovars and origins grown on media with 9.0% NaCl were measured using Bioscreen C Microbiology Reader. Technical variability of the growth measurements was assessed and eliminated. In step 2, the growth parameters determined by Gompertz, modified-Gompertz, logistic, and Richards models and model-free splines were compared, illustrating differences in the suitability of these methods to describe the experimental data. In step 3, hierarchical clustering was used to describe the NaCl tolerance of L. monocytogenes measured by strain-specific variation in growth ability; tolerant strains had higher growth rates and maximum optical densities and shorter lag phases than susceptible strains. The spline parameter area under the curve best classified "poor", "average", and "good" growers. In step 4, the tested L. monocytogenes lineage I strains (serovars 4b and 1/2b) proved to be significantly more tolerant towards NaCl 9.0% than lineage II strains (serovars 1/2a, 1/2c, and 3a). Our protocol provides systematic tools to gain comparable data for investigating strain-specific variation of bacterial growth under stress.

IMPORTANCE The pathogen Listeria monocytogenes causes the foodborne disease listeriosis, which can be fatal in immunocompromised individuals. L. monocytogenes tolerates several environmental stressors and can persist in food-processing environments and grow in foodstuffs despite traditional control measures such as high salt content. Nonetheless, L. monocytogenes strains differ in their ability to withstand stressors. Elucidating the intra-species strain variability of L. monocytogenes stress tolerance is crucial for the identification of particularly tolerant strains. To enhance reliable identification of variability in bacterial stress tolerance phenotypes, we compiled a large-scale protocol for the entire data assembly and analysis of microbial growth experiments, providing a systematic approach and check-list for experiments on strain-specific growth ability. Our study illustrated the diversity and strain-specific variation of L. monocytogenes stress tolerance with an unprecedented scope and discovered biologically relevant serovar- and lineage-dependent phenotypes of NaCl tolerance.

单核细胞增生性李斯特菌引起严重的食源性疾病李斯特菌病，并且由于其高度的压力耐受性，可在与食物相关的环境中生存。编制了用于微生物生长实验的数据汇编和分析协议，以阐明单核细胞增生李斯特菌胁迫耐受性的菌株变异性。该协议包括：1）在压力下测量生长能力；2）选择合适的生长参数计算方法；3）菌株间生长方式的比较；4）对发现差异的生物学解释。在步骤1中，单核细胞增生李斯特菌使用Bioscreen C Microbiology Reader测量了在含9.0％NaCl的培养基上生长的各种血清型的菌株（n = 388）。评估并消除了生长测量的技术差异。在第2步中，比较了由Gompertz，改良Gompertz，logistic和Richards模型和无模型样条确定的生长参数，从而说明了这些方法在描述实验数据方面的适用性差异。在步骤3中，使用层次聚类描述单核细胞增生李斯特菌的NaCl耐受性通过菌株特定的生长能力变化来测量；与易感菌株相比，耐性菌株具有更高的生长速率，最大光密度和更短的滞后阶段。曲线下的样条曲线参数区域最好分类为“差”，“平均”和“好”种植者。在步骤4中，证明测试的单核细胞增生李斯特氏菌谱系I菌株（血清型4b和1 / 2b）比谱系II菌株（血清型1 / 2a，1 / 2c和3a）对NaCl 9.0％的耐受性强得多。我们的协议提供系统的工具，以获取可比的数据，以研究应激条件下细菌生长的菌株特异性变异。

重要事项单核细胞增生李斯特氏菌病原体引起食源性李斯特菌病，对免疫功能低下的人可能致命。尽管传统的控制措施（例如高盐含量），单核细胞增生李斯特菌可以耐受多种环境胁迫，并且可以在食品加工环境中持续存在并在食品中生长。尽管如此，单核细胞增生李斯特氏菌菌株承受压力的能力不同。阐明单核细胞增生李斯特菌种内菌株的变异性应力耐受性对于鉴定特别耐受的菌株至关重要。为了增强对细菌胁迫耐受性表型变异性的可靠鉴定，我们为整个数据汇编和微生物生长实验分析汇编了大规模协议，为菌株特异性生长能力的实验提供了系统的方法和清单。我们的研究说明了单核细胞增生李斯特氏菌胁迫耐受性的多样性和特定菌株的变化具有前所未有的范围，并发现了NaCl耐受的生物学相关血清型和谱系依赖性表型。