Papers by Cristel Archambaud
The unforeseen intracellular lifestyle of Enterococcus faecalis in hepatocytes
Enterococcus faecalis is a bacterial species present at a sub-dominant level in the human gut mic... more Enterococcus faecalis is a bacterial species present at a sub-dominant level in the human gut microbiota. This commensal turns into an opportunistic pathogen under specific conditions involving dysbiosis and host immune deficiency. E. faecalis is also the only intestinal pathobiont identified to date as contributing to liver damage in alcoholic liver disease. We have previously observed that E. faecalis is internalized in hepatocytes. Here, the survival and fate of E. faecalis was examined in hepatocytes, the main epithelial cell type in the liver. Although referred to as an extracellular pathogen, we demonstrate that E. faecalis is able to survive and divide in hepatocytes, and form intracellular clusters in two distinct hepatocyte cell lines, in primary mouse hepatocytes, as well as in vivo. This novel process extends to kidney cells. Unravelling the intracellular lifestyle of E. faecalis, our findings contribute to the understanding of pathobiont-driven diseases.
Commensal bacteria augment Staphylococcus aureus infection by inactivation of phagocyte-derived reactive oxygen species
PLOS Pathogens
Staphylococcus aureus is a human commensal organism and opportunist pathogen, causing potentially... more Staphylococcus aureus is a human commensal organism and opportunist pathogen, causing potentially fatal disease. The presence of non-pathogenic microflora or their components, at the point of infection, dramatically increases S. aureus pathogenicity, a process termed augmentation. Augmentation is associated with macrophage interaction but by a hitherto unknown mechanism. Here, we demonstrate a breadth of cross-kingdom microorganisms can augment S. aureus disease and that pathogenesis of Enterococcus faecalis can also be augmented. Co-administration of augmenting material also forms an efficacious vaccine model for S. aureus. In vitro, augmenting material protects S. aureus directly from reactive oxygen species (ROS), which correlates with in vivo studies where augmentation restores full virulence to the ROS-susceptible, attenuated mutant katA ahpC. At the cellular level, augmentation increases bacterial survival within macrophages via amelioration of ROS, leading to proliferation an...
Gut Microbes
The capacity of bacterial pathogens to infect their hosts depends on the tight spatiotemporal reg... more The capacity of bacterial pathogens to infect their hosts depends on the tight spatiotemporal regulation of virulence genes. The Listeria monocytogenes (Lm) metal efflux pump repressor CadC is highly expressed during late infection stages, modulating lipoprotein processing and host immune response. Here we investigate the potential of CadC as broad repressor of virulence genes. We show that CadC represses the expression of the bile salt hydrolase impairing Lm resistance to bile. During late infection, in absence of CadC-dependent repression, the constitutive bile salt hydrolase expression induces the overexpression of the cholic acid efflux pump MdrT that is unfavorable to Lm virulence. We establish the CadC regulon and show that CadC represses additional virulence factors activated by σ B during colonization of the intestinal lumen. CadC is thus a general repressor that promotes Lm virulence by down-regulating, at late infection stages, genes required for survival in the gastrointestinal tract. This demonstrates for the first time how bacterial pathogens can repurpose regulators to spatiotemporally repress virulence genes and optimize their infectious capacity.
Scientific Reports
Rigottier-Gois & pascale serror enterococci are subdominant members of the human gastrointestinal... more Rigottier-Gois & pascale serror enterococci are subdominant members of the human gastrointestinal microbiota. Enterococcus faecalis is generally harmless for healthy individuals, but it can cause a diverse range of infections in immunodeficient or elderly patients with severe underlying diseases. In this study, we analysed the levels of intestinal translocation of indigenous enterococci in C57BL/6, CF-1 and CX3CR1 −/− mice upon clindamycin antibiotic-induced dysbiosis. We found that C57BL/6 was the most permissive model for enterococcal translocation and that initiation of E. faecalis translocation coincided with a threshold of enterococcal colonisation in the gut lumen, which once reached, triggered E. faecalis dissemination to deeper organs. We showed that the extent to which E. faecalis clinical strain VE14821 competed with indigenous enterococci differed between the C57BL/6 and CX3CR1 −/− models. Finally, using a simplified gnotobiotic model, we observed E. faecalis crossing an intact intestinal tract using intestinal epithelial cells as one route to reach the lamina propria. Our study opens new perspectives for assessing the effect of various immunodeficiencies and for investigating mechanisms underlying enterococcal translocation. Enterococci are typical intestinal pathobionts. Although subdominant in the core intestinal microbiota and relatively harmless for healthy humans, under certain circumstances enterococci can cause infections such as bacteraemia, peritonitis, endocarditis, and urinary tract, wound, and device-related infections 1. The successful survival of enterococci in the hospital environment can be attributed to multidrug resistance and pathogenic traits. E. faecalis and E. faecium are the Enterococcus species most commonly associated with infection and vancomycin resistance. Enterococci are now ranked as the third most common type of nosocomial pathogen, with E. faecalis accounting for 60 to 80% of enterococcal infections 2,3. Despite increasing epidemiological evidence that intestinal domination by vancomycin-resistant enterococci (VRE) precedes human bloodstream infections 4-6 , it remains to be established whether E. faecalis blooming upon antibiotic treatment leads to translocation and subsequent human bloodstream infection. Pioneering work using mouse models highlighted the effect of antibiotic treatments on enterococcal colonisation and translocation 7,8. Enterococcal translocation has since been reported after antibiotic-induced dysbiosis, coinfection, severe inflammatory conditions, severe burn injuries and acute irradiation, and after disruption of intestinal mucosal barrier function in various mouse genetic backgrounds 8-16. Miyazaki et al. 10 showed that mice after treatment with antimicrobial agents and cyclophosphamide displayed increased susceptibility to VRE bacteraemia, leading to death in one-third of the mice following oral VRE inoculation. Amongst the 13 mouse lines tested under these conditions, C57BL/6 mice were identified as the strain most susceptible to VRE infection after oral inoculation. However, whether higher susceptibility of C57BL/6 mice correlates with a higher translocation efficiency remains to be established. Enterococcal translocation has also been investigated in some studies combining deficiency of mucosal immunity and intestinal dysbiosis 17,18. CX3CR1 −/− mice have been shown to display increased levels of bacterial translocation to the mesenteric lymph nodes, with Enterococcus being the predominant genus 18 .
Scientific reports, Jan 29, 2018
Enterococcus faecalis, an organism generally not pathogenic for healthy humans, has the potential... more Enterococcus faecalis, an organism generally not pathogenic for healthy humans, has the potential to cause disease in susceptible hosts. While it seems to be equipped to interact with and circumvent host immune defense, most of the molecular and cellular mechanisms underlying the enterococcal infectious process remain elusive. Here, we investigated the role of the Enterococcal Leucine Rich protein A (ElrA), an internalin-like protein of E. faecalis also known as a virulence factor. ElrA was previously shown to prevent adhesion to macrophages. We show that ElrA does not inhibit the basic phagocytic process, but is able to prevent sensing and migration of macrophages toward E. faecalis. Presence or absence of FHL2, a eukaryotic partner of ElrA, does not affect the ElrA-dependent mechanism preventing macrophage migration. However, we highlight a partial contribution of FHL2 in ElrA-mediated virulence in vivo. Our results indicate that ElrA plays at least a dual role of which anti-phago...
Rôle de la sérine/thréonine phosphatase Stp dans le processus infectieux de Listeria monocytogenes
Http Www Theses Fr, 2006
Lactobacillus paracasei subsp. paracasei, as an agent for inhibiting listeria monocytogenes in vivo infection
Quantitative Proteome Analyses Identify PrfA-Responsive Proteins and Phosphoproteins in Listeria monocytogenes
Journal of Proteome Research, 2014
Protein phosphorylation is a major mechanism of signal transduction in bacteria. Here, we analyze... more Protein phosphorylation is a major mechanism of signal transduction in bacteria. Here, we analyzed the proteome and phosphoproteome of a wild-type strain of the food-borne pathogen Listeria monocytogenes that was grown in either chemically defined medium or rich medium containing glucose. We then compared these results with those obtained from an isogenic prfA* mutant that produced a constitutively active form of PrfA, the main transcriptional activator of virulence genes. In the prfA* mutant grown in rich medium, we identified 256 peptides that were phosphorylated on serine (S), threonine (T), or tyrosine (Y) residues, with a S/T/Y ratio of 155:75:12. Strikingly, we detected five novel phosphosites on the virulence protein ActA. This protein was known to be phosphorylated by a cellular kinase in the infected host, but phosphorylation by a listerial kinase had not previously been reported. Unexpectedly, SILAC experiments with the prfA* mutant grown in chemically defined medium revealed that, in addition to previously described PrfA-regulated proteins, several other proteins were significantly overproduced, among them were several proteins involved in purine biosynthesis. This work provides new information for our understanding of the correlation among protein phosphorylation, virulence mechanisms, and carbon metabolism.
F1000 - Post-publication peer review of the biomedical literature, 2000
The Bacteroides are a numerically dominant genus of the human intestinal microbiota. These organi... more The Bacteroides are a numerically dominant genus of the human intestinal microbiota. These organisms harbor a rare bacterial pathway for incorporation of exogenous fucose into capsular polysaccharides and glycoproteins. The infrequency of glycoprotein synthesis by bacteria prompted a more detailed analysis of this process. Here, we demonstrate that Bacteroides fragilis has a general O-glycosylation system. The proteins targeted for glycosylation include those predicted to be involved in protein folding, protein-protein interactions, peptide degradation, as well as surface lipoproteins. Protein glycosylation is central to the physiology of B. fragilis and is necessary for the organism to competitively colonize the mammalian intestine. We provide evidence that general Oglycosylation systems are conserved among intestinal Bacteroides species and likely contribute to the predominance of Bacteroides in the human intestine.
Supplemental Data Loss of the LAT Adaptor Converts Antigen-Responsive T Cells into Pathogenic Effectors that Function Independently of the T Cell Receptor
Faculty of 1000 evaluation for Listeria monocytogenes ActA-mediated escape from autophagic recognition
F1000 - Post-publication peer review of the biomedical literature, 2000
F1000 - Post-publication peer review of the biomedical literature, 2000
Our knowledge on species and function composition of the human gut microbiome is rapidly increasi... more Our knowledge on species and function composition of the human gut microbiome is rapidly increasing, but it is still based on very few cohorts and little is known about their variation across the world. Combining 22 newly sequenced fecal metagenomes of individuals from 4 countries with previously published datasets, we identified three robust clusters (enterotypes hereafter) that are not nation or continent-specific. We confirmed the enterotypes also in two published, larger cohorts suggesting that intestinal microbiota variation is generally stratified, not continuous. This further indicates the existence of a limited number of well-balanced host-microbial symbiotic states that might respond differently to diet and drug intake. The enterotypes are mostly driven by species composition, but abundant molecular functions are not necessarily provided by abundant species, highlighting the importance of a functional analysis for a community understanding. While individual host properties such as body mass index, age, or gender cannot explain the observed enterotypes, data-driven marker genes or functional modules can be identified for each
Proceedings of the National Academy of Sciences, 2013
Research on gut microbiota and its influence on normal physiological processes and various diseas... more Research on gut microbiota and its influence on normal physiological processes and various diseases, including inflammatory bowel diseases, obesity, or infectious diseases, is rapidly
Proceedings of the National Academy of Sciences, 2012
PLoS Genetics, 2014
The human bacterial pathogen Listeria monocytogenes is emerging as a model organism to study RNA-... more The human bacterial pathogen Listeria monocytogenes is emerging as a model organism to study RNA-mediated regulation in pathogenic bacteria. A class of non-coding RNAs called CRISPRs (clustered regularly interspaced short palindromic repeats) has been described to confer bacterial resistance against invading bacteriophages and conjugative plasmids. CRISPR function relies on the activity of CRISPR associated (cas) genes that encode a large family of proteins with nuclease or helicase activities and DNA and RNA binding domains. Here, we characterized a CRISPR element (RliB) that is expressed and processed in the L. monocytogenes strain EGD-e, which is completely devoid of cas genes. Structural probing revealed that RliB has an unexpected secondary structure comprising basepair interactions between the repeats and the adjacent spacers in place of canonical hairpins formed by the palindromic repeats. Moreover, in contrast to other CRISPR-Cas systems identified in Listeria, RliB-CRISPR is ubiquitously present among Listeria genomes at the same genomic locus and is never associated with the cas genes. We showed that RliB-CRISPR is a substrate for the endogenously encoded polynucleotide phosphorylase (PNPase) enzyme. The spacers of the different Listeria RliB-CRISPRs share many sequences with temperate and virulent phages. Furthermore, we show that a cas-less RliB-CRISPR lowers the acquisition frequency of a plasmid carrying the matching protospacer, provided that trans encoded cas genes of a second CRISPR-Cas system are present in the genome. Importantly, we show that PNPase is required for RliB-CRISPR mediated DNA interference. Altogether, our data reveal a yet undescribed CRISPR system whose both processing and activity depend on PNPase, highlighting a new and unexpected function for PNPase in ''CRISPRology''.
The excludon: a new concept in bacterial antisense RNA-mediated gene regulation
Nature Reviews Microbiology, 2012
In recent years, non-coding RNAs have emerged as key regulators of gene expression. Among these R... more In recent years, non-coding RNAs have emerged as key regulators of gene expression. Among these RNAs, the antisense RNAs (asRNAs) are particularly abundant, but in most cases the function and mechanism of action for a particular asRNA remains elusive. Here, we highlight a recently discovered paradigm termed the excludon, which defines a genomic locus encoding an unusually long asRNA that spans divergent genes or operons with related or opposing functions. Because these asRNAs can inhibit the expression of one operon while functioning as an mRNA for the adjacent operon, they act as fine-tuning regulatory switches in bacteria.
Nature Cell Biology, 2005
E-cadherin mediates the formation of adherens junctions between epithelial cells 1. It serves as ... more E-cadherin mediates the formation of adherens junctions between epithelial cells 1. It serves as a receptor for Listeria monocytogenes, a bacterial pathogen that enters epithelial cells 2. The L. monocytogenes surface protein, InlA, interacts with the extracellular domain of E-cadherin 3-5. In adherens junctions, this ectodomain is involved in homophilic interactions whereas the cytoplasmic domain binds β-catenin, which then recruits α-catenin. α-catenin binds to actin directly, or indirectly, thus linking E-cadherin to the actin cytoskeleton 6,7. Entry of L. monocytogenes into cells and adherens junction formation are dynamic events that involve actin and membrane rearrangements. To understand these processes better, we searched for new ligands of α-catenin. Using a two-hybrid screen, we identified a new partner of α-catenin: ARHGAP10. This protein colocalized with α-catenin at cell-cell junctions and was recruited at L. monocytogenes entry sites. In ARHGAP10-knockdown cells, L. monocytogenes entry and α-catenin recruitment at cell-cell contacts were impaired. The GAP domain of ARHGAP10 has GAP activity for RhoA and Cdc42. Its overexpression disrupted actin cables, enhanced α-catenin and cortical actin levels at cell-cell junctions and inhibited L. monocytogenes entry. Altogether, our results show that ARHGAP10 is a new component of cell-cell junctions that controls α-catenin recruitment and has a key role during L. monocytogenes uptake. α-catenin links the E-cadherin-β-catenin complex to the actin cytoskeleton and recruits proteins that regulate actin polymerization at developing junctions 6. We searched for new partners of α-catenin and investigated whether they were involved in the InlA-E-cadherin-mediated entry of L. monocytogenes, and in the composition or regulation of adherens junctions. α-catenin (906 amino acids) was used as the bait in a yeast twohybrid screen. Twenty-seven preys corresponded to the amino-terminal domain sequence of β-catenin, the best characterized partner of α-catenin.
Molecular Systems Biology, 2012
Listeria monocytogenes is a human, food-borne pathogen. Genomic comparisons between L. monocytoge... more Listeria monocytogenes is a human, food-borne pathogen. Genomic comparisons between L. monocytogenes and Listeria innocua, a closely related non-pathogenic species, were pivotal in the identification of protein-coding genes essential for virulence. However, no comprehensive comparison has focused on the non-coding genome. We used strand-specific cDNA sequencing to produce genome-wide transcription start site maps for both organisms, and developed a publicly available integrative browser to visualize and analyze both transcriptomes in different growth conditions and genetic backgrounds. Our data revealed conservation across most transcripts, but significant divergence between the species in a subset of non-coding RNAs. In L. monocytogenes, we identified 113 small RNAs (33 novel) and 70 antisense RNAs (53 novel), significantly increasing the repertoire of ncRNAs in this species. Remarkably, we identified a class of long antisense transcripts (lasRNAs) that overlap one gene while also serving as the 5 0 UTR of the adjacent divergent gene. Experimental evidence suggests that lasRNAs transcription inhibits expression of one operon while activating the expression of another. Such a lasRNA/operon structure, that we named 'excludon', might represent a novel form of regulation in bacteria.
Molecular Microbiology, 2005
Listeria monocytogenes is a pathogen that causes listeriosis, a severe food-borne infection. This... more Listeria monocytogenes is a pathogen that causes listeriosis, a severe food-borne infection. This bacterium, in order to survive and grow in the multiple conditions encountered in the host and the environment, has evolved a large number of regulatory elements, in particular many signal transduction systems based on reversible phosphorylation. The genome sequence has revealed genes for 16 putative two-component systems, four putative tyrosine phosphatases, three putative serine-threonine kinases and two putative serine-threonine phosphatases. We found that one of the latter genes, stp , encodes a functional Mn 2 +-dependent serine-threonine phosphatase similar to PPM eukaryotic phosphatases (Mg 2 +-or Mn 2 +-dependent protein phosphatase) and is required for growth of L. monocytogenes in a murine model of infection. We identified as the first target for Stp, the elongation factor EF-Tu. Post-translational phosphorylation of EF-Tu had been shown to prevent its binding to amino-acylated transfer RNA as well as to kirromycin, an antibiotic known to inhibit EF-Tu function. Accordingly, an stp deletion mutant is less sensitive to kirromycin. These results suggest an important role for Stp in regulating EF-Tu and controlling bacterial survival in the infected host.
mBio, 2014
For nearly 3 decades, listeriologists and immunologists have used mainly three strains of the sam... more For nearly 3 decades, listeriologists and immunologists have used mainly three strains of the same serovar (1/2a) to analyze the virulence of the bacterial pathogen Listeria monocytogenes. The genomes of two of these strains, EGD-e and 10403S, were released in 2001 and 2008, respectively. Here we report the genome sequence of the third reference strain, EGD, and extensive genomic and phenotypic comparisons of the three strains. Strikingly, EGD-e is genetically highly distinct from EGD (29,016 single nucleotide polymorphisms [SNPs]) and 10403S (30,296 SNPs), and is more related to serovar 1/2c than 1/2a strains. We also found that while EGD and 10403S strains are genetically very close (317 SNPs), EGD has a point mutation in the transcriptional regulator PrfA (PrfA*), leading to constitutive expression of several major virulence genes. We generated an EGD-e PrfA* mutant and showed that EGD behaves like this strain in vitro, with slower growth in broth and higher invasiveness in human...
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Papers by Cristel Archambaud