Publications by authors named "Nicolas Wenner"

14 Publications

  • Page 1 of 1

Isolation and Characterisation of Bacteriophages with Activity against Invasive Non-Typhoidal Causing Bloodstream Infection in Malawi.

Viruses 2021 Mar 15;13(3). Epub 2021 Mar 15.

Clinical Infection, Microbiology and Immunology, Institute of Infection, Veterinary and Ecological Sciences, University of Liverpool, Liverpool L69 7ZB, UK.

In recent years, novel lineages of invasive non-typhoidal (iNTS) serovars Typhimurium and Enteritidis have been identified in patients with bloodstream infection in Sub-Saharan Africa. Here, we isolated and characterised 32 phages capable of infecting . Typhimurium and . Enteritidis, from water sources in Malawi and the UK. The phages were classified in three major phylogenetic clusters that were geographically distributed. In terms of host range, Cluster 1 phages were able to infect all bacterial hosts tested, whereas Clusters 2 and 3 had a more restricted profile. Cluster 3 contained two sub-clusters, and 3.b contained the most novel isolates. This study represents the first exploration of the potential for phages to target the lineages of that are responsible for bloodstream infections in Sub-Saharan Africa.
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http://dx.doi.org/10.3390/v13030478DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7999457PMC
March 2021

Stepwise evolution of Salmonella Typhimurium ST313 causing bloodstream infection in Africa.

Nat Microbiol 2021 Mar 21;6(3):327-338. Epub 2020 Dec 21.

Institute of Infection, Veterinary and Ecological Sciences, University of Liverpool, Liverpool, UK.

Bloodstream infections caused by nontyphoidal Salmonella are a major public health concern in Africa, causing ~49,600 deaths every year. The most common Salmonella enterica pathovariant associated with invasive nontyphoidal Salmonella disease is Salmonella Typhimurium sequence type (ST)313. It has been proposed that antimicrobial resistance and genome degradation has contributed to the success of ST313 lineages in Africa, but the evolutionary trajectory of such changes was unclear. Here, to define the evolutionary dynamics of ST313, we sub-sampled from two comprehensive collections of Salmonella isolates from African patients with bloodstream infections, spanning 1966 to 2018. The resulting 680 genome sequences led to the discovery of a pan-susceptible ST313 lineage (ST313 L3), which emerged in Malawi in 2016 and is closely related to ST313 variants that cause gastrointestinal disease in the United Kingdom and Brazil. Genomic analysis revealed degradation events in important virulence genes in ST313 L3, which had not occurred in other ST313 lineages. Despite arising only recently in the clinic, ST313 L3 is a phylogenetic intermediate between ST313 L1 and L2, with a characteristic accessory genome. Our in-depth genotypic and phenotypic characterization identifies the crucial loss-of-function genetic events that occurred during the stepwise evolution of invasive S. Typhimurium across Africa.
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http://dx.doi.org/10.1038/s41564-020-00836-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8018540PMC
March 2021

Genetic variation in the MacAB-TolC efflux pump influences pathogenesis of invasive Salmonella isolates from Africa.

PLoS Pathog 2020 08 24;16(8):e1008763. Epub 2020 Aug 24.

Department of Microbiology and Immunology, Stanford University School of Medicine, Stanford, California, United States of America.

The various sub-species of Salmonella enterica cause a range of disease in human hosts. The human-adapted Salmonella enterica serovar Typhi enters the gastrointestinal tract and invades systemic sites to cause enteric (typhoid) fever. In contrast, most non-typhoidal serovars of Salmonella are primarily restricted to gut tissues. Across Africa, invasive non-typhoidal Salmonella (iNTS) have emerged with an ability to spread beyond the gastrointestinal tract and cause systemic bloodstream infections with increased morbidity and mortality. To investigate this evolution in pathogenesis, we compared the genomes of African iNTS isolates with other Salmonella enterica serovar Typhimurium and identified several macA and macB gene variants unique to African iNTS. MacAB forms a tripartite efflux pump with TolC and is implicated in Salmonella pathogenesis. We show that macAB transcription is upregulated during macrophage infection and after antimicrobial peptide exposure, with macAB transcription being supported by the PhoP/Q two-component system. Constitutive expression of macAB improves survival of Salmonella in the presence of the antimicrobial peptide C18G. Furthermore, these macAB variants affect replication in macrophages and influence fitness during colonization of the murine gastrointestinal tract. Importantly, the infection outcome resulting from these macAB variants depends upon both the Salmonella Typhimurium genetic background and the host gene Nramp1, an important determinant of innate resistance to intracellular bacterial infection. The variations we have identified in the MacAB-TolC efflux pump in African iNTS may reflect evolution within human host populations that are compromised in their ability to clear intracellular Salmonella infections.
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http://dx.doi.org/10.1371/journal.ppat.1008763DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7446830PMC
August 2020

Decoding the stoichiometric composition and organisation of bacterial metabolosomes.

Nat Commun 2020 04 24;11(1):1976. Epub 2020 Apr 24.

Institute of Integrative Biology, University of Liverpool, Crown Street, Liverpool, L69 7ZB, UK.

Some enteric bacteria including Salmonella have evolved the propanediol-utilising microcompartment (Pdu MCP), a specialised proteinaceous organelle that is essential for 1,2-propanediol degradation and enteric pathogenesis. Pdu MCPs are a family of bacterial microcompartments that are self-assembled from hundreds of proteins within the bacterial cytosol. Here, we seek a comprehensive understanding of the stoichiometric composition and organisation of Pdu MCPs. We obtain accurate stoichiometry of shell proteins and internal enzymes of the natural Pdu MCP by QconCAT-driven quantitative mass spectrometry. Genetic deletion of the major shell protein and absolute quantification reveal the stoichiometric and structural remodelling of metabolically functional Pdu MCPs. Decoding the precise protein stoichiometry allows us to develop an organisational model of the Pdu metabolosome. The structural insights into the Pdu MCP are critical for both delineating the general principles underlying bacterial organelle formation, structural robustness and function, and repurposing natural microcompartments using synthetic biology for biotechnological applications.
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http://dx.doi.org/10.1038/s41467-020-15888-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7181861PMC
April 2020

A window into lysogeny: revealing temperate phage biology with transcriptomics.

Microb Genom 2020 02 5;6(2). Epub 2020 Feb 5.

Institute of Integrative Biology, University of Liverpool, Liverpool, UK.

Prophages are integrated phage elements that are a pervasive feature of bacterial genomes. The fitness of bacteria is enhanced by prophages that confer beneficial functions such as virulence, stress tolerance or phage resistance, and these functions are encoded by 'accessory' or 'moron' loci. Whilst the majority of phage-encoded genes are repressed during lysogeny, accessory loci are often highly expressed. However, it is challenging to identify novel prophage accessory loci from DNA sequence data alone. Here, we use bacterial RNA-seq data to examine the transcriptional landscapes of five prophages. We show that transcriptomic data can be used to heuristically enrich for prophage features that are highly expressed within bacterial cells and represent functionally important accessory loci. Using this approach, we identify a novel antisense RNA species in prophage BTP1, STnc6030, which mediates superinfection exclusion of phage BTP1. Bacterial transcriptomic datasets are a powerful tool to explore the molecular biology of temperate phages.
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http://dx.doi.org/10.1099/mgen.0.000330DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7067206PMC
February 2020

The use of chicken and insect infection models to assess the virulence of African Salmonella Typhimurium ST313.

PLoS Negl Trop Dis 2019 07 26;13(7):e0007540. Epub 2019 Jul 26.

Department of Functional and Comparative Genomics, Institute of Integrative Biology, University of Liverpool, Liverpool, United Kingdom.

Over recent decades, Salmonella infection research has predominantly relied on murine infection models. However, in many cases the infection phenotypes of Salmonella pathovars in mice do not recapitulate human disease. For example, Salmonella Typhimurium ST313 is associated with enhanced invasive infection of immunocompromised people in Africa, but infection of mice and other animal models with ST313 have not consistently reproduced this invasive phenotype. The introduction of alternative infection models could help to improve the quality and reproducibility of pathogenesis research by facilitating larger-scale experiments. To investigate the virulence of S. Typhimurium ST313 in comparison with ST19, a combination of avian and insect disease models were used. We performed experimental infections in five lines of inbred and one line of outbred chickens, as well as in the alternative chick embryo and Galleria mellonella wax moth larvae models. This extensive set of experiments identified broadly similar patterns of disease caused by the African and global pathovariants of Salmonella Typhimurium in the chicken, the chicken embryo and insect models. A comprehensive analysis of all the chicken infection experiments revealed that the African ST313 isolate D23580 had a subtle phenotype of reduced levels of organ colonisation in inbred chickens, relative to ST19 strain 4/74. ST313 isolate D23580 also caused reduced mortality in chicken embryos and insect larvae, when compared with ST19 4/74. We conclude that these three infection models do not reproduce the characteristics of the systemic disease caused by S. Typhimurium ST313 in humans.
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http://dx.doi.org/10.1371/journal.pntd.0007540DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6685681PMC
July 2019

The diversity, evolution and ecology of Salmonella in venomous snakes.

PLoS Negl Trop Dis 2019 06 4;13(6):e0007169. Epub 2019 Jun 4.

Department of Functional and Comparative Genomics, Institute of Integrative Biology, University of Liverpool, Liverpool, United Kingdom.

Background: Reptile-associated Salmonella bacteria are a major, but often neglected cause of both gastrointestinal and bloodstream infection in humans globally. The diversity of Salmonella enterica has not yet been determined in venomous snakes, however other ectothermic animals have been reported to carry a broad range of Salmonella bacteria. We investigated the prevalence and diversity of Salmonella in a collection of venomous snakes and non-venomous reptiles.

Methodology/principle Findings: We used a combination of selective enrichment techniques to establish a unique dataset of reptilian isolates to study Salmonella enterica species-level evolution and ecology and used whole-genome sequencing to investigate the relatedness of phylogenetic groups. We observed that 91% of venomous snakes carried Salmonella, and found that a diverse range of serovars (n = 58) were carried by reptiles. The Salmonella serovars belonged to four of the six Salmonella enterica subspecies: diarizonae, enterica, houtanae and salamae. Subspecies enterica isolates were distributed among two distinct phylogenetic clusters, previously described as clade A (52%) and clade B (48%). We identified metabolic differences between S. diarizonae, S. enterica clade A and clade B involving growth on lactose, tartaric acid, dulcitol, myo-inositol and allantoin.

Significance: We present the first whole genome-based comparative study of the Salmonella bacteria that colonise venomous and non-venomous reptiles and shed new light on Salmonella evolution. Venomous snakes examined in this study carried a broad range of Salmonella, including serovars which have been associated with disease in humans such as S. Enteritidis. The findings raise the possibility that venomous snakes could be a reservoir for Salmonella serovars associated with human salmonellosis.
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http://dx.doi.org/10.1371/journal.pntd.0007169DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6548357PMC
June 2019

Adding function to the genome of African Salmonella Typhimurium ST313 strain D23580.

PLoS Biol 2019 01 15;17(1):e3000059. Epub 2019 Jan 15.

Institute of Integrative Biology, University of Liverpool, Liverpool, United Kingdom.

Salmonella Typhimurium sequence type (ST) 313 causes invasive nontyphoidal Salmonella (iNTS) disease in sub-Saharan Africa, targeting susceptible HIV+, malarial, or malnourished individuals. An in-depth genomic comparison between the ST313 isolate D23580 and the well-characterized ST19 isolate 4/74 that causes gastroenteritis across the globe revealed extensive synteny. To understand how the 856 nucleotide variations generated phenotypic differences, we devised a large-scale experimental approach that involved the global gene expression analysis of strains D23580 and 4/74 grown in 16 infection-relevant growth conditions. Comparison of transcriptional patterns identified virulence and metabolic genes that were differentially expressed between D23580 versus 4/74, many of which were validated by proteomics. We also uncovered the S. Typhimurium D23580 and 4/74 genes that showed expression differences during infection of murine macrophages. Our comparative transcriptomic data are presented in a new enhanced version of the Salmonella expression compendium, SalComD23580: http://bioinf.gen.tcd.ie/cgi-bin/salcom_v2.pl. We discovered that the ablation of melibiose utilization was caused by three independent SNP mutations in D23580 that are shared across ST313 lineage 2, suggesting that the ability to catabolize this carbon source has been negatively selected during ST313 evolution. The data revealed a novel, to our knowledge, plasmid maintenance system involving a plasmid-encoded CysS cysteinyl-tRNA synthetase, highlighting the power of large-scale comparative multicondition analyses to pinpoint key phenotypic differences between bacterial pathovariants.
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http://dx.doi.org/10.1371/journal.pbio.3000059DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6333337PMC
January 2019

Role of a single noncoding nucleotide in the evolution of an epidemic African clade of .

Proc Natl Acad Sci U S A 2018 03 27;115(11):E2614-E2623. Epub 2018 Feb 27.

Institute of Integrative Biology, University of Liverpool, L69 7ZB Liverpool, United Kingdom;

serovar Typhimurium ST313 is a relatively newly emerged sequence type that is causing a devastating epidemic of bloodstream infections across sub-Saharan Africa. Analysis of hundreds of genomes has revealed that ST313 is closely related to the ST19 group of Typhimurium that cause gastroenteritis across the world. The core genomes of ST313 and ST19 vary by only ∼1,000 SNPs. We hypothesized that the phenotypic differences that distinguish African from ST19 are caused by certain SNPs that directly modulate the transcription of virulence genes. Here we identified 3,597 transcriptional start sites of the ST313 strain D23580, and searched for a gene-expression signature linked to pathogenesis of We identified a SNP in the promoter of the gene that caused high expression of the PgtE virulence factor in African Typhimurium, increased the degradation of the factor B component of human complement, contributed to serum resistance, and modulated virulence in the chicken infection model. We propose that high levels of PgtE expression by African Typhimurium ST313 promote bacterial survival and dissemination during human infection. Our finding of a functional role for an extragenic SNP shows that approaches used to deduce the evolution of virulence in bacterial pathogens should include a focus on noncoding regions of the genome.
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http://dx.doi.org/10.1073/pnas.1714718115DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5856525PMC
March 2018

Characterization of the Prophage Repertoire of African Typhimurium ST313 Reveals High Levels of Spontaneous Induction of Novel Phage BTP1.

Front Microbiol 2017 23;8:235. Epub 2017 Feb 23.

Institute of Integrative Biology, University of Liverpool Liverpool, UK.

In the past 30 years, bloodstream infections have become a significant health problem in sub-Saharan Africa and are responsible for the deaths of an estimated 390,000 people each year. The disease is predominantly caused by a recently described sequence type of Typhimurium: ST313, which has a distinctive set of prophage sequences. We have thoroughly characterized the ST313-associated prophages both genetically and experimentally. ST313 representative strain D23580 contains five full-length prophages: BTP1, Gifsy-2, ST64B, Gifsy-1, and BTP5. We show that common Typhimurium prophages Gifsy-2, Gifsy-1, and ST64B are inactivated in ST313 by mutations. Prophage BTP1 was found to be a functional novel phage, and the first isolate of the proposed new species "Salmonella virus BTP1", belonging to the genus. Surprisingly, ∼10 BTP1 virus particles ml were detected in the supernatant of non-induced, stationary-phase cultures of strain D23580, representing the highest spontaneously induced phage titer so far reported for a bacterial prophage. High spontaneous induction is shown to be an intrinsic property of prophage BTP1, and indicates the phage-mediated lysis of around 0.2% of the lysogenic population. The fact that BTP1 is highly conserved in ST313 poses interesting questions about the potential fitness costs and benefits of novel prophages in epidemic Typhimurium ST313.
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http://dx.doi.org/10.3389/fmicb.2017.00235DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5322425PMC
February 2017

Functional analysis of pyochelin-/enantiopyochelin-related genes from a pathogenicity island of Pseudomonas aeruginosa strain PA14.

Biometals 2014 Jun 30;27(3):559-73. Epub 2014 Mar 30.

Département de Microbiologie Fondamentale, Université de Lausanne, Bâtiment Biophore, Quartier UNIL-Sorge, 1015, Lausanne, Switzerland.

Genomic islands are foreign DNA blocks inserted in so-called regions of genomic plasticity (RGP). Depending on their gene content, they are classified as pathogenicity, symbiosis, metabolic, fitness or resistance islands, although a detailed functional analysis is often lacking. Here we focused on a 34-kb pathogenicity island of Pseudomonas aeruginosa PA14 (PA14GI-6), which is inserted at RGP5 and carries genes related to those for pyochelin/enantiopyochelin biosynthesis. These enantiomeric siderophores of P. aeruginosa and certain strains of Pseudomonas protegens are assembled by a thiotemplate mechanism from salicylate and two molecules of cysteine. The biochemical function of several proteins encoded by PA14GI-6 was investigated by a series of complementation analyses using mutants affected in potential homologs. We found that PA14_54940 codes for a bifunctional salicylate synthase/salicyl-AMP ligase (for generation and activation of salicylate), that PA14_54930 specifies a dihydroaeruginoic acid (Dha) synthetase (for coupling salicylate with a cysteine-derived thiazoline ring), that PA14_54910 produces a type II thioesterase (for quality control), and that PA14_54880 encodes a serine O-acetyltransferase (for increased cysteine availability). The structure of the PA14GI-6-specified metabolite was determined by mass spectrometry, thin-layer chromatography, and HPLC as (R)-Dha, an iron chelator with antibacterial, antifungal and antitumor activity. The conservation of this genomic island in many clinical and environmental P. aeruginosa isolates of different geographical origin suggests that the ability for Dha production may confer a selective advantage to its host.
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http://dx.doi.org/10.1007/s10534-014-9729-4DOI Listing
June 2014

NrsZ: a novel, processed, nitrogen-dependent, small non-coding RNA that regulates Pseudomonas aeruginosa PAO1 virulence.

Environ Microbiol 2014 Apr;16(4):1053-68

The opportunistic pathogen Pseudomonas aeruginosa PAO1 has a remarkable capacity to adapt to various environments and to survive with limited nutrients. Here, we report the discovery and characterization of a novel small non-coding RNA: NrsZ (nitrogen-regulated sRNA). We show that under nitrogen limitation, NrsZ is induced by the NtrB/C two component system, an important regulator of nitrogen assimilation and P. aeruginosa's swarming motility, in concert with the alternative sigma factor RpoN. Furthermore, we demonstrate that NrsZ modulates P. aeruginosa motility by controlling the production of rhamnolipid surfactants, virulence factors notably needed for swarming motility. This regulation takes place through the post-transcriptional control of rhlA, a gene essential for rhamnolipids synthesis. Interestingly, we also observed that NrsZ is processed in three similar short modules, and that the first short module encompassing the first 60 nucleotides is sufficient for NrsZ regulatory functions.
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http://dx.doi.org/10.1111/1462-2920.12272DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4253122PMC
April 2014

Novel targets of the CbrAB/Crc carbon catabolite control system revealed by transcript abundance in Pseudomonas aeruginosa.

PLoS One 2012 24;7(10):e44637. Epub 2012 Oct 24.

Max F. Perutz Laboratories, Department of Microbiology, Immunobiology and Genetics, University of Vienna, Dr. Bohrgasse 9, Vienna, Austria.

The opportunistic human pathogen Pseudomonas aeruginosa is able to utilize a wide range of carbon and nitrogen compounds, allowing it to grow in vastly different environments. The uptake and catabolism of growth substrates are organized hierarchically by a mechanism termed catabolite repression control (Crc) whereby the Crc protein establishes translational repression of target mRNAs at CA (catabolite activity) motifs present in target mRNAs near ribosome binding sites. Poor carbon sources lead to activation of the CbrAB two-component system, which induces transcription of the small RNA (sRNA) CrcZ. This sRNA relieves Crc-mediated repression of target mRNAs. In this study, we have identified novel targets of the CbrAB/Crc system in P. aeruginosa using transcriptome analysis in combination with a search for CA motifs. We characterized four target genes involved in the uptake and utilization of less preferred carbon sources: estA (secreted esterase), acsA (acetyl-CoA synthetase), bkdR (regulator of branched-chain amino acid catabolism) and aroP2 (aromatic amino acid uptake protein). Evidence for regulation by CbrAB, CrcZ and Crc was obtained in vivo using appropriate reporter fusions, in which mutation of the CA motif resulted in loss of catabolite repression. CbrB and CrcZ were important for growth of P. aeruginosa in cystic fibrosis (CF) sputum medium, suggesting that the CbrAB/Crc system may act as an important regulator during chronic infection of the CF lung.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0044637PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3480352PMC
April 2013

Iron acquisition with the natural siderophore enantiomers pyochelin and enantio-pyochelin in Pseudomonas species.

Biometals 2011 Jun 25;24(3):513-22. Epub 2010 Dec 25.

Département de Microbiologie Fondamentale, Université de Lausanne, Bâtiment Biophore, Lausanne, Switzerland.

The bacterial siderophore pyochelin is composed of salicylate and two cysteine-derived heterocycles, the second of which is modified by reduction and N-methylation during biosynthesis. In Pseudomonas aeruginosa, the first cysteine residue is converted to its D-isoform during thiazoline ring formation, whereas the second cysteine remains in its L-configuration. Stereochemistry is opposite in the Pseudomonas fluorescens siderophore enantio-pyochelin, in which the first ring originates from L-cysteine and the second ring from D-cysteine. Both siderophores promote growth of the producer organism during iron limitation and induce the expression of their biosynthesis genes by activating the transcriptional AraC-type regulator PchR. However, neither siderophore is functional as an iron carrier or as a transcriptional inducer in the other species, demonstrating that both processes are highly stereospecific. Stereospecificity of pyochelin/enantio-pyochelin-mediated iron uptake is ensured at two levels: (i) by the outer membrane siderophore receptors and (ii) by the cytosolic PchR regulators.
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http://dx.doi.org/10.1007/s10534-010-9399-9DOI Listing
June 2011