The regulation of genes of unknown function implicated in nitrosative stress tolerance in Escherichia coli K-12

This study was designed to determine the regulatory network that controls expression from two $Escherichia$ $coli$ K-12 promoters, $pyeaR$ and $pogt$, during anaerobic growth. These promoters were identified from transcriptomic studies as being positively regulated by NarL independently of FNR, the master regulator of anaerobic respiration. Biochemical and genetic analyses presented in this study confirmed that expression from both the $yeaR$ and $ogt$ promoters is dependent upon NarL, which binds to a single site in the $yeaR$ promoter and two sites in the $ogt$ promoter. The nucleoid-associated protein, Fis, repressed transcription from both promoters, especially in rich medium, by binding to sites that overlap the NarL site, excluding the essential activator. Both promoters were more active in the absence of functional FNR. However, mutational analysis revealed that FNR does not bind to the $yeaR$ promoter region, so this effect is indirect. How the absence of functional FNR might affect NarL-dependent nitrite signalling was investigated. The Ogt protein is known function as an O$^6$-alkyguanine methyltransferase. However, the functions of the gene products of $yeaR-yoaG$ and another operon implicated in nitrosative stress management, $hcp-hcr$, were unknown. Strains carrying a chromosomal $yeaR-yoaG$ deletion were not more sensitive to nitric oxide or hydroxylamine compared with the parental strain, suggesting that the products of this operon are not essential for dealing with these toxic nitrogen species. Conversely, a strain deleted in $hcp-hcr$ was shown to be slightly more sensitive to both nitric oxide and hydroxylamine, implicating Hcp and Hcr in nitrosative stress management.EThOS - Electronic Theses Online ServiceGBUnited Kingdo

Achieving minimum standards for infection prevention and control in Sierra Leone: Urgent need for a quantum leap in progress in the COVID-19 era!

From MDPI via Jisc Publications RouterHistory: accepted 2022-04-20, pub-electronic 2022-05-06Publication status: PublishedIntroduction: Good Infection prevention and control (IPC) is vital for tackling antimicrobial resistance and limiting health care-associated infections. We compared IPC performance before (2019) and during the COVID-19 (2021) era at the national IPC unit and all regional (4) and district hospitals (8) in Sierra Leone. Methods: Cross-sectional assessments using standardized World Health Organizations IPC checklists. IPC performance scores were graded as inadequate = 0−25%, basic = 25.1−50%, intermediate = 50.1−75%, and advanced = 75.1−100%. Results: Overall performance improved from ‘basic’ to ‘intermediate’ at the national IPC unit (41% in 2019 to 58% in 2021) and at regional hospitals (37% in 2019 to 54% in 2021) but remained ‘basic’ at district hospitals (37% in 2019 to 50% in 2021). Priority gaps at the national IPC unit included lack of: a dedicated IPC budget, monitoring the effectiveness of IPC trainings and health care-associated infection surveillance. Gaps at hospitals included no assessment of hospital staffing needs, inadequate infrastructure for IPC and lack of a well-defined monitoring plan with clear goals, targets and activities. Conclusion: Although there is encouraging progress in IPC performance, it is slower than desired in light of the COVID-19 pandemic. There is urgent need to mobilize political will, leadership and resources and make a quantum leap forward.19pubpub

Evolution of microscopic heterogeneity and dynamics in choline chloride-based deep eutectic solvents

Deep eutectic solvents (DESs) are an emerging class of non-aqueous solvents that are potentially scalable, easy to prepare and functionalize for many applications ranging from biomass processing to energy storage technologies. Predictive understanding of the fundamental correlations between local structure and macroscopic properties is needed to exploit the large design space and tunability of DESs for specific applications. Here, we employ a range of computational and experimental techniques that span length-scales from molecular to macroscopic and timescales from picoseconds to seconds to study the evolution of structure and dynamics in model DESs, namely Glyceline and Ethaline, starting from the parent compounds. We show that systematic addition of choline chloride leads to microscopic heterogeneities that alter the primary structural relaxation in glycerol and ethyleneglycol and result in new dynamic modes that are strongly correlated to the macroscopic properties of the DES formed

The regulation of genes of unknown function implicated in nitrosative stress tolerance in Escherichia coli K-12

This study was designed to determine the regulatory network that controls expression from two $Escherichia$ $coli$ K-12 promoters, $pyeaR$ and $pogt$, during anaerobic growth. These promoters were identified from transcriptomic studies as being positively regulated by NarL independently of FNR, the master regulator of anaerobic respiration. Biochemical and genetic analyses presented in this study confirmed that expression from both the $yeaR$ and $ogt$ promoters is dependent upon NarL, which binds to a single site in the $yeaR$ promoter and two sites in the $ogt$ promoter. The nucleoid-associated protein, Fis, repressed transcription from both promoters, especially in rich medium, by binding to sites that overlap the NarL site, excluding the essential activator. Both promoters were more active in the absence of functional FNR. However, mutational analysis revealed that FNR does not bind to the $yeaR$ promoter region, so this effect is indirect. How the absence of functional FNR might affect NarL-dependent nitrite signalling was investigated. The Ogt protein is known function as an O$^6$-alkyguanine methyltransferase. However, the functions of the gene products of $yeaR-yoaG$ and another operon implicated in nitrosative stress management, $hcp-hcr$, were unknown. Strains carrying a chromosomal $yeaR-yoaG$ deletion were not more sensitive to nitric oxide or hydroxylamine compared with the parental strain, suggesting that the products of this operon are not essential for dealing with these toxic nitrogen species. Conversely, a strain deleted in $hcp-hcr$ was shown to be slightly more sensitive to both nitric oxide and hydroxylamine, implicating Hcp and Hcr in nitrosative stress management

Competition between NarL-dependent activation and Fis-dependent repression controls expression from the Escherichia coli yeaR and ogt promoters

International audienceThe Escherichia coli NarL protein is a global gene regulatory factor that activates transcription at many target promoters in response to nitrate and nitrite ions. Although most NarL-dependent promoters are also co-dependent on a second transcription factor, FNR, two targets, the yeaR and ogt promoters are activated by NarL alone with no involvement of FNR. Biochemical and genetic studies presented here show that activation of the yeaR promoter is dependent on the binding of NarL to a single target centred at position 43.5, whilst activation at the ogt promoter requires NarL binding to tandem DNA targets centred at position -45.5 and -78.5. NarL dependent activation at both the yeaR and ogt promoters is decreased in rich medium and this depends on Fis, a nucleoid-associated protein. DNAse I footprinting studies identified Fis binding sites that overlap the yeaR promoter NarL site at position -43.5, and the ogt promoter NarL site at position -78.5, and suggest that Fis represses both promoters by displacing NarL. The ogt gene encodes an O6-alkylguanine DNA alkyltransferase and, hence, this is the first report of expression of a DNA repair function being controlled by nitrate ions

Organization of the Electron Transfer Chain to Oxygen in the Obligate Human Pathogen Neisseria gonorrhoeae: Roles for Cytochromes c4 and c5, but Not Cytochrome c2, in Oxygen Reduction▿ †

Although Neisseria gonorrhoeae is a prolific source of eight c-type cytochromes, little is known about how its electron transfer pathways to oxygen are organized. In this study, the roles in the respiratory chain to oxygen of cytochromes c2, c4, and c5, encoded by the genes cccA, cycA, and cycB, respectively, have been investigated. Single mutations in genes for either cytochrome c4 or c5 resulted in an increased sensitivity to growth inhibition by excess oxygen and small decreases in the respiratory capacity of the parent, which were complemented by the chromosomal integration of an ectopic, isopropyl-β-d-thiogalactopyranoside (IPTG)-inducible copy of the cycA or cycB gene. In contrast, a cccA mutant reduced oxygen slightly more rapidly than the parent, suggesting that cccA is expressed but cytochrome c2 is not involved in electron transfer to cytochrome oxidase. The deletion of cccA increased the sensitivity of the cycB mutant to excess oxygen but decreased the sensitivity of the cycA mutant. Despite many attempts, a double mutant defective in both cytochromes c4 and c5 could not be isolated. However, a strain with the ectopically encoded, IPTG-inducible cycB gene with deletions in both cycA and cycB was constructed: the growth and survival of this strain were dependent upon the addition of IPTG, so gonococcal survival is dependent upon the synthesis of either cytochrome c4 or c5. These results define the gonococcal electron transfer chain to oxygen in which cytochromes c4 and c5, but not cytochrome c2, provide alternative pathways for electron transfer from the cytochrome bc1 complex to the terminal oxidase cytochrome cbb3

The NsrR Regulon of Escherichia coli K-12 Includes Genes Encoding the Hybrid Cluster Protein and the Periplasmic, Respiratory Nitrite Reductase▿

Successful pathogens must be able to protect themselves against reactive nitrogen species generated either as part of host defense mechanisms or as products of their own metabolism. The regulatory protein NsrR (a member of the Rrf2 family of transcription factors) plays key roles in this stress response. Microarray analysis revealed that NsrR represses nine operons encoding 20 genes in Escherichia coli MG1655, including the hmpA, ytfE, and ygbA genes that were previously shown to be regulated by NsrR. Novel NsrR targets revealed by this study include hcp-hcr (which were predicted in a recent bioinformatic study to be NsrR regulated) and the well-studied nrfA promoter that directs the expression of the periplasmic respiratory nitrite reductase. Conversely, transcription from the ydbC promoter is strongly activated by NsrR. Regulation of the nrf operon by NsrR is consistent with the ability of the periplasmic nitrite reductase to reduce nitric oxide and hence protect against reactive nitrogen species. Gel retardation assays were used to show that both FNR and NarL bind to the hcp promoter. The expression of hcp and the contiguous gene hcr is not induced by hydroxylamine. As hmpA and ytfE encode a nitric oxide reductase and a mechanism to repair iron-sulfur centers damaged by nitric oxide, the demonstration that hcp-hcr, hmpA, and ytfE are the three transcripts most tightly regulated by NsrR highlights the possibility that the hybrid cluster protein, HCP, might also be part of a defense mechanism against reactive nitrogen stress

Sequencing a piece of history:complete genome sequence of the original Escherichia coli strain

In 1885, Theodor Escherich first described the Bacillus coli commune, which was subsequently renamed Escherichia coli. We report the complete genome sequence of this original strain (NCTC 86). The 5 144 392 bp circular chromosome encodes the genes for 4805 proteins, which include antigens, virulence factors, antimicrobial-resistance factors and secretion systems, of a commensal organism from the pre-antibiotic era. It is located in the E. coli A subgroup and is closely related to E. coli K-12 MG1655. E. coli strain NCTC 86 and the non-pathogenic K-12, C, B and HS strains share a common backbone that is largely co-linear. The exception is a large 2 803 932 bp inversion that spans the replication terminus from gmhB to clpB. Comparison with E. coli K-12 reveals 41 regions of difference (577 351 bp) distributed across the chromosome. For example, and contrary to current dogma, E. coli NCTC 86 includes a nine gene sil locus that encodes a silver-resistance efflux pump acquired before the current widespread use of silver nanoparticles as an antibacterial agent, possibly resulting from the widespread use of silver utensils and currency in Germany in the 1800s. In summary, phylogenetic comparisons with other E. coli strains confirmed that the original strain isolated by Escherich is most closely related to the non-pathogenic commensal strains. It is more distant from the root than the pathogenic organisms E. coli 042 and O157 : H7; therefore, it is not an ancestral state for the species

Achieving Minimum Standards for Infection Prevention and Control in Sierra Leone: Urgent Need for a Quantum Leap in Progress in the COVID-19 Era!

From Europe PMC via Jisc Publications RouterHistory: ppub 2022-05-01, epub 2022-05-06Publication status: PublishedIntroductionGood Infection prevention and control (IPC) is vital for tackling antimicrobial resistance and limiting health care-associated infections. We compared IPC performance before (2019) and during the COVID-19 (2021) era at the national IPC unit and all regional (4) and district hospitals (8) in Sierra Leone.MethodsCross-sectional assessments using standardized World Health Organizations IPC checklists. IPC performance scores were graded as inadequate = 0-25%, basic = 25.1-50%, intermediate = 50.1-75%, and advanced = 75.1-100%.ResultsOverall performance improved from 'basic' to 'intermediate' at the national IPC unit (41% in 2019 to 58% in 2021) and at regional hospitals (37% in 2019 to 54% in 2021) but remained 'basic' at district hospitals (37% in 2019 to 50% in 2021). Priority gaps at the national IPC unit included lack of: a dedicated IPC budget, monitoring the effectiveness of IPC trainings and health care-associated infection surveillance. Gaps at hospitals included no assessment of hospital staffing needs, inadequate infrastructure for IPC and lack of a well-defined monitoring plan with clear goals, targets and activities.ConclusionAlthough there is encouraging progress in IPC performance, it is slower than desired in light of the COVID-19 pandemic. There is urgent need to mobilize political will, leadership and resources and make a quantum leap forward