Two-Component-System Histidine Kinases Involved in Growth of Listeria monocytogenes EGD-e at Low Temperatures

Abstract

Two-component systems (TCSs) aid bacteria in adapting to a wide variety of stress conditions. While the role of TCS response regulators in the cold tolerance of the psychrotrophic foodborne pathogen Listeria monocytogenes has been demonstrated previously, no comprehensive studies showing the role of TCS histidine kinases of L. monocytogenes at low temperature have been performed. We compared the expression levels of each histidine kinase-encoding gene of L. monocytogenes EGD-e in logarithmic growth phase at 3°C and 37°C, as well as the expression levels 30 min, 3 h, and 7 h after cold shock at 5°C and preceding cold shock (at 37°C). We constructed a deletion mutation in each TCS histidine kinase gene, monitored the growth of the EGD-e wild-type and mutant strains at 3°C and 37°C, and measured the minimum growth temperature of each strain. Two genes, yycG and lisK, proved significant in regard to induced relative expression levels under cold conditions and cold-sensitive mutant phenotypes. Moreover, the ⌬resE mutant showed a lower growth rate than that of wild-type EGD-e at 3°C. Eleven other genes showed upregulated gene expression but revealed no cold-sensitive phenotypes. The results show that the histidine kinases encoded by yycG and lisK are important for the growth and adaptation of L. monocytogenes EGD-e at low temperature. Listeria monocytogenes is a low-GϩC, Gram-positive, nonspore-forming coccobacillus ubiquitously found in the environment (1, 2). It has been isolated from foods and food-processing premises, where it has been confirmed to be able to persist for several years (3-7). L. monocytogenes is the causative agent of listeriosis, with a reported annual incidence of approximately 0.3 case per 100,000 population in the United States and 0.2 to 0.8 case per 100,000 population in Europe (8-10). Although relatively rare, listeriosis is nevertheless a severe foodborne disease with a mortality of up to 20 to 30% (1,8,11,12). L. monocytogenes surmounts a wide range of stress conditions and is able to grow at temperatures as low as Ϫ0.4°C (13). This sets critical challenges for the modern food industry, since L. monocytogenes is consequently able to grow in ready-to-eat foods with a long shelf-life and in products stored at refrigeration temperatures. Thus, it is crucial to study and elucidate the factors enhancing the cold tolerance of L. monocytogenes. Two-component regulatory signaling systems (TCSs) are among the major systems that aid bacteria in adapting to many arduous stress factors encountered in nature and during food processing. A typical TCS is constituted of a transmembrane sensor histidine kinase (HK) and a cognate cytoplasmic response regulator (RR) (14-17). The signaling pathway of the TCS is based on protein phosphorylation. In the simplest form of signal transduction, the sensor domain of the HK detects a specific stimulus or stress factor in cells or in their environment. This leads to the autophosphorylation of a specific His residue in the HK dimerization domain. The cognate RR then ca...

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