Mechanisms of anaerobic nitric oxide detoxification by Salmonella enterica serovar Typhimurium

Salmonella is the cause of millions of food- and water-borne infections worldwide. Systemic infection and gastroenteritis are the main diseases and often prove fatal to immunocompromised patients. Key to Salmonella’s pathogenicity is the survival of several components of the innate immune system encountered during infection. Reactive oxygen and nitrogen species (ROS and RNS) are an integral part of this antibacterial defence of the immune system. Exposure to ROS and RNS occurs within phagocytic immune cells such as macrophages, where such generation of radicals is used to combat pathogens. NO is a radical belonging to the group of RNS that damages bacterial DNA and proteins. Detoxification of NO is essential during infection to allow Salmonella to survive and replicate within macrophages. Three enzymes are currently known to help Salmonella to detoxify NO, but their deletion, however, does not eliminate Salmonella’s survival. Therefore, it is predicted that further mechanisms for NO detoxification exist. In this study, the core NO regulon has been identified: Expression of nine genes is significantly increased during endogenous and exogenous NO exposure of S. Typhimurium. Their functions range from carbon starvation, cytochrome oxidation, iron-sulphur repair and NO reduction to putative proteins with unknown function, some of which contain domains for tellurite resistance. Single and combination deletion strains have shown that these genes are important to decrease anaerobic NO sensitivity of S. Typhimurium and for intracellular survival in murine macrophages. Furthermore, we have shown for the first time that the core NO regulon also provides protection against tellurite. Tellurite is toxic and requires detoxification when encountered. Reducing tellurite to yield the elemental tellurium results in the release of ROS, which then need to be detoxified further. Deletion strains sensitive to tellurite have also shown increased sensitivity to NO. Concurrently, tellurite resistance genes also facilitate the defence against NO

The production and detoxification of a potent cytotoxin, nitric oxide, by pathogenic enteric bacteria

The nitrogen cycle is based on several redox reactions that are mainly accomplished by prokaryotic organisms, some archaea and a few eukaryotes, which use these reactions for assimilatory, dissimilatory or respiratory purposes. One group is the Enterobacteriaceae family of Gammaproteobacteria, which have their natural habitats in soil, marine environments or the intestines of humans and other warm-blooded animals. Some of the genera are pathogenic and usually associated with intestinal infections. Our body possesses several physical and chemical defence mechanisms to prevent pathogenic enteric bacteria from invading the gastrointestinal tract. One response of the innate immune system is to activate macrophages, which produce the potent cytotoxin nitric oxide (NO). However, some pathogens have evolved the ability to detoxify NO to less toxic compounds, such as the neuropharmacological agent and greenhouse gas nitrous oxide (N2O), which enables them to overcome the host's attack. The same mechanisms may be used by bacteria producing NO endogenously as a by-product of anaerobic nitrate respiration. In the present review, we provide a brief introduction into the NO detoxification mechanisms of two members of the Enterobacteriaceae family: Escherichia coli and Salmonella enterica serovar Typhimurium. These are discussed as comparative non-pathogenic and pathogenic model systems in order to investigate the importance of detoxifying NO and producing N2O for the pathogenicity of enteric bacteria

Resolving the contributions of the membrane-bound and periplasmic nitrate reductase systems to nitric oxide and nitrous oxide production in Salmonella enterica serovar Typhimurium

The production of cytotoxic nitric oxide (NO) and conversion into the neuropharmacological agent and potent greenhouse gas nitrous oxide (N2O) is linked with anoxic nitrate catabolism by Salmonella enterica serovar Typhimurium. Salmonella can synthesize two types of nitrate reductase: a membrane-bound form (Nar) and a periplasmic form (Nap). Nitrate catabolism was studied under nitrate-rich and nitrate-limited conditions in chemostat cultures following transition from oxic to anoxic conditions. Intracellular NO production was reported qualitatively by assessing transcription of the NO-regulated genes encoding flavohaemoglobin (Hmp), flavorubredoxin (NorV) and hybrid cluster protein (Hcp). A more quantitative analysis of the extent of NO formation was gained by measuring production of N2O, the end-product of anoxic NO-detoxification. Under nitrate-rich conditions, the nar, nap, hmp, norV and hcp genes were all induced following transition from the oxic to anoxic state, and 20% of nitrate consumed in steady-state was released as N2O when nitrite had accumulated to millimolar levels. The kinetics of nitrate consumption, nitrite accumulation and N2O production were similar to those of wild-type in nitrate-sufficient cultures of a nap mutant. In contrast, in a narG mutant, the steady-state rate of N2O production was ~30-fold lower than that of the wild-type. Under nitrate-limited conditions, nap, but not nar, was up-regulated following transition from oxic to anoxic metabolism and very little N2O production was observed. Thus a combination of nitrate-sufficiency, nitrite accumulation and an active Nar-type nitrate reductase leads to NO and thence N2O production, and this can account for up to 20% of the nitrate catabolized