Personne : Tremblay, Denise
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Groupe de recherche en écologie buccale, Faculté de médecine dentaire, Université Laval
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- PublicationAccès libreProphages of the genus Bifidobacterium as modulating agents of the infant gut microbiota(2016-07-08) Lugli, Gabriele Andrea; Tremblay, Denise; Milani, Christian; Moineau, Sylvain; Turroni, Francesca; Priego, Sabrina.; Mancabelli, Leonardo; Ward, Doyle V.; Ossiprandi, Maria Cristina; Sinderen, Douwe van; Ventura, MarcoPhage predation is one of the key forces that shape genetic diversity in bacterial genomes. Phages are also believed to act as modulators of the microbiota composition and, consequently, as agents that drive bacterial speciation in complex bacterial communities. Very little is known about the occurrence and genetic variability of (pro)phages within the genus, a dominant bacterial group of the human infant microbiota. Here, we performed cataloguing of the predicted prophage sequences from the genomes of all currently recognized bifidobacterial type strains. We analysed their genetic diversity and deduced their evolutionary development, thereby highlighting an intriguing origin. Furthermore, we assessed infant gut microbiomes for the presence of (pro)phage sequences and found compelling evidence that these viral elements influence the composition of bifidobacterial communities in the infant gut microbiota
- PublicationAccès libreCharacterization and diversity of phages infecting Aeromonas salmonicida subsp. salmonicida(Nature Publishing Group, 2017-08-01) Tremblay, Denise; Paquet, Valérie; Bernatchez, Alex; Moineau, Sylvain; Vincent, Antony; Charette, StevePhages infecting Aeromonas salmonicida subsp. salmonicida, the causative agent of the fsh disease furunculosis, have been isolated for decades but very few of them have been characterized. Here, the host range of 12 virulent phages, including three isolated in the present study, was evaluated against a panel of 65A. salmonicida isolates, including representatives of the psychrophilic subspecies salmonicida, smithia, masoucida, and the mesophilic subspecies pectinolytica. This bacterial set also included three isolates from India suspected of being members of a new subspecies. Our results allowed to elucidate a lytic dichotomy based on the lifestyle of A. salmonicida (mesophilic or psychrophilic) and more generally, on phage types (lysotypes) for the subspecies salmonicida. The genomic analyses of the 12 phages from this study with those available in GenBank led us to propose an A. salmonicida phage pan-virome. Our comparative genomic analyses also suggest that some phage genes were under positive selection and A. salmonicida phage genomes having a discrepancy in GC% compared to the host genome encode tRNA genes to likely overpass the bias in codon usage. Finally, we propose a new classifcation scheme for A. salmonicida phages.
- PublicationRestreintGenomic diversity of phages infecting probiotic strains of Lactobacillus paracasei(American Society for Microbiology, 2015-12-22) Mercanti, Diego J.; Tremblay, Denise; Moineau, Sylvain; Capra, María Luján; Labrie, Simon; Luján Quiberoni, Andrea del; Rousseau, Geneviève M.Strains of the Lactobacillus casei group have been extensively studied because some are used as probiotics in foods. Conversely, their phages have received much less attention. We analyzed the complete genome sequences of five L. paracasei temperate phages: CL1, CL2, iLp84, iLp1308, and iA2. Only phage iA2 could not replicate in an indicator strain. The genome lengths ranged from 34,155 bp (iA2) to 39,474 bp (CL1). Phages iA2 and iLp1308 (34,176 bp) possess the smallest genomes reported, thus far, for phages of the L. casei group. The GC contents of the five phage genomes ranged from 44.8 to 45.6%. As observed with many other phages, their genomes were organized as follows: genes coding for DNA packaging, morphogenesis, lysis, lysogeny, and replication. Phages CL1, CL2, and iLp1308 are highly related to each other. Phage iLp84 was also related to these three phages, but the similarities were limited to gene products involved in DNA packaging and structural proteins. Genomic fragments of phages CL1, CL2, iLp1308, and iLp84 were found in several genomes of L. casei strains. Prophage iA2 is unrelated to these four phages, but almost all of its genome was found in at least four L. casei strains. Overall, these phages are distinct from previously characterized Lactobacillus phages. Our results highlight the diversity of L. casei phages and indicate frequent DNA exchanges between phages and their hosts.
- PublicationRestreintStructure, adsorption to host, and infection mechanism of virulent lactococcal phage p2(American Society for Microbiology, 2013-09-11) Bebeacua, Cecilia; Tremblay, Denise; Moineau, Sylvain; Farenc, Carine; Chapot-Chartier, Marie-Pierre; Sadovskaya, Irina; Heel, Marin van; Veesler, David; Cambillau, ChristianLactococcal siphophages from the 936 and P335 groups infect the Gram-positive bacterium Lactococcus lactis using receptor binding proteins (RBPs) attached to their baseplate, a large multiprotein complex at the distal part of the tail. We have previously reported the crystal and electron microscopy (EM) structures of the baseplates of phages p2 (936 group) and TP901-1 (P335 group) as well as the full EM structure of the TP901-1 virion. Here, we report the complete EM structure of siphophage p2, including its capsid, connector complex, tail, and baseplate. Furthermore, we show that the p2 tail is characterized by the presence of protruding decorations, which are related to adhesins and are likely contributed by the major tail protein C-terminal domains. This feature is reminiscent of the tail of Escherichia coli phage and Bacillus subtilis phage SPP1 and might point to a common mechanism for establishing initial interactions with their bacterial hosts. Comparative analyses showed that the architecture of the phage p2 baseplate differs largely from that of lactococcal phage TP901-1. We quantified the interaction of its RBP with the saccharidic receptor and determined that specificity is due to lower koff values of the RBP/saccharidic dissociation. Taken together, these results suggest that the infection of L. lactis strains by phage p2 is a multistep process that involves reversible attachment, followed by baseplate activation, specific attachment of the RBPs to the saccharidic receptor, and DNA ejection.
- PublicationRestreintLactococcus lactis type III-A CRISPR-Cas system cleaves bacteriophage RNA(Tandfonline, 2018-10-02) Millen, Anne M.; Tremblay, Denise; Moineau, Sylvain; Samson, Julie; Magadán, Alfonso H.; Rousseau, Geneviève M.; Romero, Dennis A.CRISPR-Cas defends microbial cells against invading nucleic acids including viral genomes. Recent studies have shown that type III-A CRISPR-Cas systems target both RNA and DNA in a transcriptiondependent manner. We previously found a type III-A system on a conjugative plasmid in Lactococcus lactis which provided resistance against virulent phages of the Siphoviridae family. Its naturally occurring spacers are oriented to generate crRNAs complementary to target phage mRNA, suggesting transcription-dependent targeting. Here, we show that only constructs whose spacers produce crRNAs complementary to the phage mRNA confer phage resistance in L. lactis. In vivo nucleic acid cleavage assays showed that cleavage of phage dsDNA genome was not detected within phage-infected L. lactis cells. On the other hand, Northern blots indicated that the lactococcal CRISPR-Cas cleaves phage mRNA in vivo. These results cannot exclude that single-stranded phage DNA is not being targeted, but phage DNA replication has been shown to be impaired.
- PublicationAccès libreDiversity and host specificity revealed by biological characterization and whole genome sequencing of bacteriophages infecting salmonella enterica(MDPI, 2019-09-14) Fong, Karen; Tremblay, Denise; Moineau, Sylvain; Delaquis, Pascal; Levesque, Roger; Goodridge, Lawrence; Suttle, Curtis Arnold; Wang, SiyunPhages infecting members of the opportunistic human pathogen, Salmonella enterica, are widespread in natural environments and offer a potential source of agents that could be used for controlling populations of this bacterium; yet, relatively little is known about these phages. Here we describe the isolation and characterization of 45 phages of Salmonella enterica from disparate geographic locations within British Columbia, Canada. Host-range profiling revealed host-specific patterns of susceptibility and resistance, with several phages identified that have a broad-host range (i.e., able to lyse >40% of bacterial hosts tested). One phage in particular, SE13, is able to lyse 51 out of the 61 Salmonella strains tested. Comparative genomic analyses also revealed an abundance of sequence diversity in the sequenced phages. Alignment of the genomes grouped the phages into 12 clusters with three singletons. Phages within certain clusters exhibited extraordinarily high genome homology (>98% nucleotide identity), yet between clusters, genomes exhibited a span of diversity (<50% nucleotide identity). Alignment of the major capsid protein also supported the clustering pattern observed with alignment of the whole genomes. We further observed associations between genomic relatedness and the site of isolation, as well as genetic elements related to DNA metabolism and host virulence. Our data support the knowledge framework for phage diversity and phage–host interactions that are required for developing phage-based applications for various sectors, including biocontrol, detection and typing.
- PublicationAccès libreComplete genome sequence of Streptococcus thermophilus SMQ-301, a model strain for phage-host interactions(American Society for Microbiology, 2015-05-21) Tremblay, Denise; Moineau, Sylvain; Labrie, Simon; Wasserscheid, Jessica; Dewar, Ken; Plante, Pier-Luc; Corbeil, JacquesStreptococcus thermophilus is used by the dairy industry to manufacture yogurt and several cheeses. Using PacBio and Illumina platforms, we sequenced the genome of S. thermophilus SMQ-301, the host of several virulent phages. The genome is composed of 1,861,792 bp and contains 2,037 genes, 67 tRNAs, and 18 rRNAs.
- PublicationRestreintA new Microviridae phage isolated from a failed biotechnological process driven by Escherichia coli(American Society for Microbiology, 2014-10-17) Tremblay, Denise; Moineau, Sylvain; Dupuis, Marie-Ève; Labrie, Simon; Plante, Pier-Luc; Corbeil, JacquesBacteriophages are present in every environment that supports bacterial growth, including manmade ecological niches. Virulent phages may even slow or, in more severe cases, interrupt bioprocesses driven by bacteria. Escherichia coli is one of the most widely used bacteria for large-scale bioprocesses; however, literature describing phage-host interactions in this industrial context is sparse. Here, we describe phage MED1 isolated from a failed industrial process. Phage MED1 (Microviridae family, with a singlestranded DNA [ssDNA] genome) is highly similar to the archetypal phage phiX174, sharing >95% identity between their genomic sequences. Whole-genome phylogenetic analysis of 52 microvirus genomes from public databases revealed three genotypes (alpha3, G4, and phiX174). Phage MED1 belongs to the phiX174 group. We analyzed the distribution of single nucleotide variants in MED1 and 18 other phiX174-like genomes and found that there are more missense mutations in genes G, B, and E than in the other genes of these genomes. Gene G encodes the spike protein, involved in host attachment. The evolution of this protein likely results from the selective pressure on phages to rapidly adapt to the molecular diversity found at the surface of their hosts.
- PublicationAccès libreStructure of lactococcal phage p2 baseplate and its mechanism of activation(National Academy of Sciences, 2010-04-13) Sciara, Giuliano; Tremblay, Denise; Bebeacua, Cecilia; Moineau, Sylvain; Bron, Patrick; Ortiz-Lombardia, Miguel; Lichière, Julie; Heel, Marin van; Campanaccia, Valérie; Cambillau, ChristianSiphoviridae is the most abundant viral family on earth which infects bacteria as well as archaea. All known siphophages infecting gram+ Lactococcus lactis possess a baseplate at the tip of their tail involved in host recognition and attachment. Here, we report analysis of the p2 phage baseplate structure by X-ray crystallography and electron microscopy and propose a mechanism for the baseplate activation during attachment to the host cell. This ∼1 MDa, Escherichia coli-expressed baseplate is composed of three protein species, including six trimers of the receptor-binding protein (RBP). RBPs host-recognition domains point upwards, towards the capsid, in agreement with the electron-microscopy map of the free virion. In the presence of Ca2þ, a cation mandatory for infection, the RBPs rotated 200° downwards, presenting their binding sites to the host, and a channel opens at the bottom of the baseplate for DNA passage. These conformational changes reveal a novel siphophage activation and host-recognition mechanism leading ultimately to DNA ejection.
- PublicationRestreintReceptor-binding protein of lactococcus lactis phages : identification and characterization of the saccharide receptor-binding site(American Society for Microbiology, 2006-03-17) Tremblay, Denise; Tegoni, Mariella; Labrie, Steve; Spinelli, Silvia; Moineau, Sylvain; Campanacci, Valérie; Blangy, Stéphanie; Huyghe, Céline; Desmyter, Aline; Cambillau, ChristianPhage p2, a member of the lactococcal 936 phage species, infects Lactococcus lactis strains by binding initially to specific carbohydrate receptors using its receptor-binding protein (RBP). The structures of p2 RBP, a homotrimeric protein composed of three domains, and of its complex with a neutralizing llama VH domain (VHH5) have been determined (S. Spinelli, A. Desmyter, C. T. Verrips, H. J. de Haard, S. Moineau, and C. Cambillau, Nat. Struct. Mol. Biol. 13:85–89, 2006). Here, we show that VHH5 was able to neutralize 12 of 50 lactococcal phages belonging to the 936 species. Moreover, escape phage mutants no longer neutralized by VHH5 were isolated from 11 of these phages. All of the mutations (but one) cluster in the RBP/VHH5 interaction surface that delineates the receptor-binding area. A glycerol molecule, observed in the 1.7-Å resolution structure of RBP, was found to bind tightly (Kd = 0.26 M) in a crevice located in this area. Other saccharides bind RBP with comparable high affinity. These data prove the saccharidic nature of the bacterial receptor recognized by phage p2 and identify the position of its binding site in the RBP head domain.