Francis Repoila

French National Institute for Agriculture, Food, and Environment (INRAE) | INRAE

Enterococcus cecorum is an emerging pathogen responsible for osteomyelitis, spondylitis, and femoral head necrosis causing animal suffering and mortality and requiring antimicrobial use in poultry. Paradoxically, E. cecorum is a common inhabitant of the intestinal microbiota of adult chickens. Despite evidence suggesting the existence of clones wit...

Enterococcus cecorum is an emerging pathogen responsible for osteomyelitis, spondylitis, and femoral head necrosis causing animal suffering, mortality, and requiring antimicrobial use in poultry. Paradoxically, E. cecorum is a common inhabitant of the intestinal microbiota of adult chickens. Despite evidence suggesting the existence of clones with...

Enterococcus faecalis is a natural inhabitant of the human gastrointestinal tract. This bacterial species is subdominant in a healthy physiological state of the gut microbiota (eubiosis) in adults, but can become dominant and cause infections when the intestinal homeostasis is disrupted (dysbiosis). The relatively high concentrations of bile acids...

The family Flavobacteriaceae (phylum Bacteroidetes) is a major component of soil, marine and freshwater ecosystems. In this understudied family, Flavobacterium psychrophilum is a freshwater pathogen that infects salmonid fish worldwide, with critical environmental and economic impact. Here, we report an extensive transcriptome analysis that establi...

Transcription initiation and RNA processing govern gene expression and enable bacterial adaptation by reshaping the RNA landscape. The aim of this study was to simultaneously observe these two fundamental processes in a transcriptome responding to an environmental signal. A controlled σE system in E. coli was coupled to our previously-described tag...

In eukaryotes, RNA species originating from pervasive transcription are regulators of various cellular processes, from the expression of individual genes to the control of cellular development and oncogenesis. In prokaryotes, the function of pervasive transcription and its output on cell physiology is still unknown. Most bacteria possess terminatio...

RT-PCR analysis of antisense transcription of flhOP operon in the BSB1 and NCIB 3610 WT and RM strains. Total RNA was extracted as described in Materials and Methods and cDNA synthesis was performed using 50 ng of total RNA as template and mixture of specific oligonucleotides. Reactions were performed with (+) and without (−) reverse transcriptase...

Ectopic expression of the flhO-flhP operon does not restore biofim formation in B. subtilis RM cells. Colony and pellicle biofilm formation by B. subtilis NCIB 3610 WT and RM cells. The colony column shows individual colonies grown on MSgg agar medium for 72h at 30°C. The scale bar is 5mm. The pellicle column shows microtitre wells (diameter 1.5 cm...

Extension of S1475 RNA in the 1012 RM strain. Genomic region corresponding to the slrA gene shows the extension of S1475 RNA (can be visualized on http://genome.jouy.inra.fr/cgi-bin/seb/index.py). Sections show annotated genome (top) and expression profiles of WT (black) and RM (red) on the (+) and (–) strands (mid and bottom sections). (TIF)

Effect of Rho inactivation on sporulation efficiency of the PY79 strain mainly involves KinB. (A) Rho inactivation partially restores sporulation in the kinA mutant and has no effect in the kinB mutant. Sporulation efficiency of the PY79 WT, PY79 RM strains and their respective kinA and kinB mutants. Cells were inoculated in DS medium at OD600 0.05...

The tagged Rho-SPA protein remains functional. (A and B) Quantitative swarming motility assay of the NCIB 3610 (blue line), NCIB 3610 RM (red line) and NCIB 3610 rho-spa (dotted red line) strains. Bacterial cultures were grown to an OD600 of 0.5.concentrated and spotted on the plate as described (Materials and Methods). Plates were incubated at 37°...

Proteomic raw data. (XLSX)

Transcription levels in the 5'-part, central-part and 3'-part of the kinB gene across 275 RNA samples. (XLSX)

Oligonucleotides used for strains construction. (DOCX)

Inactivation of the abrB does not restore biofilm formation by B. subtilis RM and double RM sinR mutant strains. Pellicle biofilm formation by B. subtilis NCIB 3610 abrB, NCIB 3610 sinR, abrB strains and their respective RM derivatives. Relevant genotypes are indicated on the top of each column. The images show microtitre wells (diameter 1.5 cm) in...

Modification of the phosphorelay in B. subtilis RM cells. Schematic representation of the multicomponent Spo0A phosphorelay. Only the key elements relevant to this study are shown. Phosphoryl groups are transferred from sensor protein kinases (KinA-E) to Spo0F, Spo0B, and ultimately to Spo0A. KbaA protein stimulates KinB activity. SivA and SivB pro...

Over-production of Rho-SPA protein in the B. subtilis WT (pRho-SPA) cells. (A) B. subtilis BSB1 WT cells containing rho-SPA translational fusion in the native chromosomal locus (lines 1 and 3) or cloned at pDG148 plasmid (lines 2 and 4; Materials and Methods) were grown in LB medium at 37°C to OD600 ∼ 0.5 and analyzed for Rho-SPA protein as describ...

Differential expression analysis of B. subtilis 1012 and NCIB 3610 RM vs. WT cells. (XLSX)

asRNAs specific for the SigD regulon. (XLSX)

Deletion of the flgM gene alleviates motility defect of NCIB 3610 RM strain. (A) Bacterial cultures were grown to an OD600 of 0.5, concentrated and spotted on the plate as described in Materials and Methods and [149]. Each icon represents top-grown image of centrally inoculated Petri plate (diameter 9 cm) containing LB and 0.3% of agar after 5 hour...

Functional enrichment analysis of differentially expressed genes. (XLSX)

Strains and plasmids used in this study. (DOCX)

Bacterial regulatory RNAs have been defined as diverse classes of cis and trans elements that may intervene at each step of gene expression, from RNA and protein synthesis to degradation. Here, we report on a few examples from Gram-positive bacteria that extend the definition of regulatory RNAs to include 5′ and 3′ UTRs that also act as cis and tra...

We discovered a chromosomal locus containing two toxin-antitoxin modules (TAs) with an antisense transcriptional organization in the E. faecalis clinical isolate V583. These TAs are homologous to the type I txpA-ratA system and the type II mazEF, respectively. We have shown that the putative MazF is toxic for E. coli and triggers RNA degradation, a...

Standard RNA-seq has a well know tendency to generate "ghost" antisense reads due to formation of spurious second strand cDNA in the sequencing process. We recently reported on a novel variant of RNA-seq coined "tagRNA-seq" introduced for the purpose of distinguishing primary from processed transcripts in bacteria. Incidentally, the additional info...

Enterococcus faecalis is the third cause of nosocomial infections. To obtain the first snapshot of transcriptional organizations in this bacterium, we used a modified RNA-seq approach enabling to discriminate primary from processed 5' RNA ends. We also validated our approach by confirming known features in Escherichia coli. We mapped 559 transcript...

Regulatory noncoding RNAs are found in 5′ untranslated regions of messengers RNAs, or as individual molecules transcribed from the complementary DNA strand to annotated genes (antisense RNAs) or from empty chromosomal regions (noncoding RNAs). They can regulate each step of gene expression by directly acting on transcription or translation processe...

Regulatory noncoding RNAs are found in 5′ untranslated regions of messengers RNAs, or as individual molecules transcribed from the complementary DNA strand to annotated genes (antisense RNAs) or from empty chromosomal regions (noncoding RNAs). They can regulate each step of gene expression by directly acting on transcription or translation processe...

pp1 interference with pp4 excision. PCR detection of pp4 circular forms in different isogenic strains (see Figure 2A): WT, pp3−, pp3− pp5−, pp1− pp3− pp5−, pp4+ pp6+ and pp4+ (see Table S1). Circular forms of pp4 are detected only in the absence of pp1. (PDF)

V583 phages infection and immunity. (DOC)

Growth of ciprofloxacin-treated strains. The WT and isogenic strains pp−, pp1+ and pp3+ pp5+ were grown at early exponential growth phase (OD600∼0.2) before treatment with ciprofloxacin at 4 or 6 µg/ml. Relative optical density (OD600) was calculated for each strain as the ratio of OD600 of the ciprofloxacin-induced cultures (Cip 4 and Cip 6) with...

Capsid size distribution of virions produced by strains pp1+ pp7+ and pp1+. Scatter plot of the capsid width (nm) measured particles for strains pp1+ pp7+ (n = 42) and pp1+ (n = 24). Strain pp1+ pp7+ produced two groups of different capsid size, small and large with a mean width of 46.1±3.7 nm and 61.1±1.3 nm, respectively. Strain pp1+ produced hom...

Mice gastro-intestinal tract colonization by strains WT and pp1−. After three days of subcutaneous administration of clindamycin, 1×1010 CFUs of each strain (WT or pp1−) were force-fed in five mice. E. faecalis burden in stools was monitored daily for four days after oral gavage. No significant differences in the efficiency of colonization between...

Bacterial strains and plasmids used in this study. (DOC)

Primers used in this study. (DOC)

Polylysogeny is frequently considered to be the result of an adaptive evolutionary process in which prophages confer fitness and/or virulence factors, thus making them important for evolution of both bacterial populations and infectious diseases. The Enterococcus faecalis V583 isolate belongs to the high-risk clonal complex 2 that is particularly w...

The Enterococcus faecalis leucine-rich protein ElrA promotes virulence by stimulating bacterial persistence in macrophages and production of the interleukin-6 (IL-6) cytokine. The ElrA protein is encoded within an operon that is poorly expressed under laboratory conditions but induced in vivo. In this study, we identify ef2687 (renamed elrR), which...

RNA maturation is a key event regulating genes at post-transcriptional level. In bacteria, it is employed to adjust the amounts of proteins and functional RNAs, often in response to environmental constraints. During the process of RNA maturation, enzymes and factors that would otherwise promote RNA degradation convert a labile RNA into a stable and...

Enterococcus faecalis is a commensal bacterium and a major opportunistic human pathogen. In this study, we combined in silico predictions with a novel 5′RACE-derivative method coined ‘5′tagRACE’, to perform the first search for non-coding RNAs (ncRNAs) encoded on the E. faecalis chromosome. We used the 5′tagRACE to simultaneously probe and characte...

Riboswitches are RNA elements acting in cis, controlling expression of their downstream genes through a metabolite-induced alteration of their secondary structure. Here, we demonstrate that two S-adenosylmethionine (SAM) riboswitches, SreA and SreB, can also function in trans and act as noncoding RNAs in Listeria monocytogenes. SreA and SreB contro...

Regulatory ncRNAs (non-coding RNAs) adjust bacterial physiology in response to environmental cues. ncRNAs can base-pair to mRNAs and change their translation efficiency and/or their stability, or they can bind to proteins and modulate their activity. ncRNAs have been discovered in several species throughout the bacterial kingdom. This review illust...

Infectious diseases are a leading cause of mortality worldwide. A major challenge in achieving their eradication is a better understanding of bacterial pathogenesis processes. The recent discovery of small noncoding RNAs (sRNAs) as modulators of gene expression in response to environmental cues has brought a new insight into bacterial regulation. s...

To identify noncoding RNAs (ncRNAs) in the pathogenic bacterium Listeria monocytogenes, we analyzed the intergenic regions (IGRs) of strain EGD-e by in silico-based approaches. Among the twelve ncRNAs found, nine are novel and specific to the Listeria genus, and two of these ncRNAs are expressed in a growth-dependent manner. Three of the ncRNAs are...

Enterohemorrhagic (EHEC) and enteropathogenic (EPEC) Escherichia coli strains carry a pathogenicity island termed locus of enterocyte effacement (LEE) responsible for attaching and effacing lesions on epithelial cells. The expression of LEE varies among isolates and is dependent on environmental cues. In the EHEC O157:H7 Sakaï isolate (RIMD-0509952...

Synthesis of the small regulatory RNA DsrA is under temperature control. The minimal dsrA promoter of 36 bp contains sufficient information to ensure such regulation. In vivo, we have analyzed the critical elements responsible for the temperature control of dsrA by using a collection of chimeric promoters combining various elements of the dsrA prom...

Adaptation to the changing environment requires both the integration of external signals and the co-ordination of internal responses. Around 50 non-coding small RNAs (sRNAs) have been described in Escherichia coli; the levels of many of these vary with changing environmental conditions. This suggests that they play a role in cell adaptation. In thi...

A burgeoning list of small RNAs with a variety of regulatory functions has been identified in both prokaryotic and eukaryotic cells. However, it remains difficult to identify small RNAs by sequence inspection. We used the high conservation of small RNAs among closely related bacterial species, as well as analysis of transcripts detected by high-den...

Many environmental parameters modulate the amount of the RpoS sigma factor in Escherichia coli. Temperature control of RpoS depends on the untranslated RNA DsrA. DsrA activates RpoS translation by pairing with the leader of the mRNA. We find that temperature affects both the rate of transcription initiation of thedsrA gene and the stability of DsrA...

Excerpt In the last few years, it has become increasingly clearthat small RNAs (sRNAs) can act as important regulators, transmitting environmental signals to up- or downregulate the synthesis and activity of proteins. However,sRNA genes are not immediately obvious from genomicsequences. Here we review what has been learned aboutthe activities of th...

Transcription of the gene osmE of Escherichia coli is inducible by elevated osmotic pressure and during the decelerating phase of growth. osmE expression is directed by a single promoter, osmEp. Decelerating phase induction of osmEp is dependent on the sigmas (RpoS) factor, whereas its osmotic induction is independent of sigmas. Purified Esigmas an...

Polymerase chain reaction analysis of a large collection of bacteriophages with T-even morphology revealed four phages that are distantly related to all the others. The genomes of these pseudo T-even phages hybridized under stringent conditions to only a limited portion of the T4 genome that encodes virus head, head-to-tail joining and contractile...

The adsorption specificity of T4 is determined by the tip of the gene 37 tail fibers which bind to receptors on the bacterial surface. T4 infects only Escherichia coli and closely related Shigella species, but rare host range mutants can be isolated that infect Yersinia pseudotuberculosis I, an evolutionally distant bacterium. Some of these mutatio...