Ethanol Adaptation Strategies in Salmonella enterica Serovar Enteritidis Revealed by Global Proteomic and Mutagenic Analyses

Abstract

Salmonella enterica subsp. enterica serovar Enteritidis is able to adapt to sublethal concentrations of ethanol, which subsequently induce tolerance of this pathogen to normally lethal ethanol challenges. This work aims to elucidate the underlying ethanol adaptation mechanisms of S. Enteritidis by proteomic and mutagenic analyses. The global proteomic response of S. Enteritidis to ethanol adaptation (5% ethanol for 1 h) was determined by isobaric tags for relative and absolute quantification (iTRAQ), and it was found that a total of 138 proteins were differentially expressed in ethanol-adapted cells compared to nonadapted cells. A total of 56 upregulated proteins were principally associated with purine metabolism and as transporters for glycine betaine, phosphate, d-alanine, thiamine, and heme, whereas 82 downregulated proteins were mainly involved in enterobactin biosynthesis and uptake, the ribosome, flagellar assembly, and virulence. Moreover, mutagenic analysis further revealed the functions of two highly upregulated proteins belonging to purine metabolism (HiuH, 5-hydroxyisourate hydrolase) and glycine betaine transport (ProX, glycine betaine-binding periplasmic protein) pathways. Deletion of either hiuH or proX resulted in the development of a stronger ethanol tolerance response, suggesting negative regulatory roles in ethanol adaptation. Collectively, this work suggests that S. Enteritidis employs multiple strategies to coordinate ethanol adaptation. IMPORTANCE Stress adaptation in foodborne pathogens has been recognized as a food safety concern since it may compromise currently employed microbial intervention strategies. While adaptation to sublethal levels of ethanol is able to induce ethanol tolerance in foodborne pathogens, the molecular mechanism underlying this phenomenon is poorly characterized. Hence, global proteomic analysis and mutagenic analysis were conducted in the current work to understand the strategies employed by Salmonella enterica subsp. enterica serovar Enteritidis to respond to ethanol adaptation. It was revealed that coordinated regulation of multiple pathways involving metabolism, ABC transporters, regulators, enterobactin biosynthesis and uptake, the ribosome, flagellar assembly, and virulence was responsible for the development of ethanol adaptation response in this pathogen. Such knowledge will undoubtedly contribute to the development and implementation of more-effective food safety interventions.

Heat Adaptation Induced Cross Protection Against Ethanol Stress in Tetragenococcus halophilus: Physiological Characteristics and Proteomic Analysis

“…45-, and 1.83-fold increase in survival) of cells, especially for ethanol stress (7.24-fold increase in survival under 10% ethanol for 2.5 h) (Figure 1). Similar results preformed in A. acidoterrestris and Salmonella enterica were also reported by the researchers that an increase in survival under thermal stress (65 • C, 5 min) from 14.72 to 62.09% in A. acidoterrestris or an improvement of ethanol tolerance (26.39fold increase in survival under 15% ethanol, 1 h) for S. enterica was observed through cross protection (He et al, 2019;Zhao et al, 2021). These results indicated that heat preadaptation may be considered as a potential strategy to engineer the robustness of strains used during food fermentation.…”

“…Genetic information processing was essential to the growth and proliferation of lactic acid bacteria. For instance, the process of translation and DNA replication were indispensable for microbes to transmit genomic information (He et al, 2019;Yao et al, 2019). He et al (2019) reported that the upregulation of ribosome-related proteins contributed to improving the stress resistance of S. enterica under high ethanol conditions.…”

“…For instance, the process of translation and DNA replication were indispensable for microbes to transmit genomic information (He et al, 2019;Yao et al, 2019). He et al (2019) reported that the upregulation of ribosome-related proteins contributed to improving the stress resistance of S. enterica under high ethanol conditions. In this manuscript, the expressions of 7 ribosomal proteins (RplX, RpmD, RpmI, RpmF, RpmB, RpmG, and RpsU) were remarkably increased in response to heat preadaptation (Figure 6).…”

Microbial Food Safety in China: Past, Present, and Future

“…Scientific and technical advancements throughout the food chain (e.g., from agriculture and food manufacturing to preparation in the home) are critical to ensure microbial food safety. In the past two decades, microbial food safety research in China was initiated on many important aspects, such as rapid detection methods and devices, food control measures, antibiotic resistance mechanisms, stress response strategies, biofilm formation mechanisms, sublethal injury mechanisms, and microbial risk assessment (Shi and Zhu, 2009;He et al, 2019;Xie et al, 2019;Ling et al, 2020;Shen et al, 2020;Sheng et al, 2020). Research progress in the aforementioned aspects has contributed a lot to the improvement of microbial food safety in China, and these areas should thus be further explored as the focuses in the future.…”

Global transcriptomic analysis of ethanol tolerance response in Salmonella Enteritidis

“…By means of the 2-DE technique, Yeh (2012) found that a total of 16 proteins were differentially expressed after exposure of C. sakazakii to 5% ethanol for 60 min. Moreover, the same sublethal ethanol treatment resulted in the differential expression of 138 proteins belonging to metabolism, enterobactin biosynthesis, virulence, and other pathways in S. Enteritidis ( He et al, 2019 ). To our best knowledge, however, there is no available literature on elucidating the ETR mechanisms in pathogenic bacteria at the transcriptome level.…”