48 research outputs found
A New Integrative and Mobilizable Element Is a Major Contributor to Tetracycline Resistance in Streptococcus dysgalactiae subsp. equisimilis
Tetracycline resistance in streptococci is mainly due to ribosomal protection mediated by the tet(M) gene that is usually located in the integrative and conjugative elements (ICEs) of the Tn916-family. In this study, we analyzed the genes involved in tetracycline resistance and the associated mobile genetic elements (MGEs) in Streptococcus dysgalactiae subsp. equisimilis (SDSE) causing invasive disease. SDSE resistant to tetracycline collected from 2012 to 2019 in a single hospital and from 2018 in three other hospitals were analyzed by whole genome sequencing. Out of a total of 84 SDSE isolates, 24 (28.5%) were resistant to tetracycline due to the presence of tet(M) (n = 22), tet(W) (n = 1), or tet(L) plus tet(W) (n = 1). The tet(M) genes were found in the ICEs of the Tn916-family (n = 10) and in a new integrative and mobilizable element (IME; n = 12). Phylogenetic analysis showed a higher genetic diversity among the strains carrying Tn916 than those having the new IME, which were closely related, and all belonged to CC15. In conclusion, tetracycline resistance in SDSE is mostly due to the tet(M) gene associated with ICEs belonging to the Tn916-family and a new IME. This new IME is a major cause of tetracycline resistance in invasive Streptococcus dysgalactiae subsp. equisimilis in our settings
Les régulateurs transcriptionnels Rgg de Streptococcus thermophilus LMG18311 (étude du rÎle de la protéine Rgg0182)
Cette thÚse a pour objectif la caractérisation des gÚnes rgg de S. thermophilus LMG18311 codant des régulateurs transcriptionnels et l'étude de leur implication dans l'adaptation à l'environnement. Ce travail montre que le gÚne rgg0182 code un régulateur transcriptionnel activant la transcription de ses gÚnes adjacents. Par ailleurs, le couple Rgg0182/Shp0182 participerait à un mécanisme de quorum sensing. De plus, la protéine Rgg0182 participe à la tolérance au stress chaud. En outre, les cellules du mutant [delta]rgg0182 présentent un phénotype d'adhésion thermo-induite via des interactions de types hydrophobes. L'analyse par microscopie à force atomique des cellules de la souche LMG18311 et du mutant [delta]rgg0182 révÚle la présence de polymÚres de surface uniquement chez la souche sauvage, suggérant que la protéine Rgg0182 régulerait l'expression de protéines de surface et de la division cellulaire. Une étude protéomique couplée à une analyse transcriptomique ont permis d'identifier plusieurs cibles de Rgg0182 qui participeraient à diverses fonctions biologiques. L'ensemble des données obtenues démontre que la protéine Rgg0182 de S. thermophilus LMG18311 est un régulateur global de l'expression génique. Par ailleurs, la transcription des 7 gÚnes rgg présents au sein du génome de S. thermophilus LMG18311 est modulée par les conditions environnementales. Les profils de transcription des 7 gÚnes rgg diffÚrent les uns par rapport aux autres, suggérant que chacun d'eux seraient requis dans des conditions de croissances différentes. Ces données posent l'hypothÚse que les protéines Rgg participeraient à la régulation fine et complexe de l'expression génique de S. thermophilusThis thesis aims to characterize the rgg genes of S. thermophilus LMG18311 coding transcriptional regulator and their involvement in environmental adaptation. This work shows that rgg0182 gene encodes a transcriptional regulator controlling the transcription of its flanking genes. The Rgg0182/Shp0182 pair could be involved in a quorum sensing mechanism. This work also demonstrates that the Rgg0182 protein is involved in S. thermophilus tolerance to heat stresses. In addition, the mutant delta rgg0182 cells exhibit a thermo-induced adhesion phenotype via hydrophobic interactions. Analyses by atomic force microscopy of LMG18311 cells of the wildtype and its derivative rgg0182 mutant reveal the presence of polymers only on the surface of the wild-type strain, suggesting that the protein Rgg0182 would regulate the expression of surface proteins and proteins of cell division. A proteomic study coupled with transcriptomic analysis led to the identification of several targets of Rgg0182 involving in various biological functions. The data obtained in this work have shown that the S. thermophilus LMG18311 rgg0182 genes encodes a global regulator of gene expression. Furthermore, transcriptional analyses, in different growth conditions, of the 7 rgg genes present in the genome of S. thermophilus LMG18311 showed that they display different expression profiles that are modulated by environmental conditions. This suggests that these genes would be required in distinct growth conditions. These data raise the hypothesis that Rgg proteins participate in the fine and complex regulation of S. thermophilus gene expressionMETZ-SCD (574632105) / SudocNANCY1-Bib. numérique (543959902) / SudocNANCY2-Bibliotheque electronique (543959901) / SudocNANCY-INPL-Bib. électronique (545479901) / SudocSudocFranceF
The Obscure World of Integrative and Mobilizable Elements, Highly Widespread Elements that Pirate Bacterial Conjugative Systems
Conjugation is a key mechanism of bacterial evolution that involves mobile genetic elements. Recent findings indicated that the main actors of conjugative transfer are not the well-known conjugative or mobilizable plasmids but are the integrated elements. This paper reviews current knowledge on âintegrative and mobilizable elementsâ (IMEs) that have recently been shown to be highly diverse and highly widespread but are still rarely described. IMEs encode their own excision and integration and use the conjugation machinery of unrelated co-resident conjugative element for their own transfer. Recent studies revealed a much more complex and much more diverse lifecycle than initially thought. Besides their main transmission as integrated elements, IMEs probably use plasmid-like strategies to ensure their maintenance after excision. Their interaction with conjugative elements reveals not only harmless hitchhikers but also hunters that use conjugative elements as target for their integration or harmful parasites that subvert the conjugative apparatus of incoming elements to invade cells that harbor them. IMEs carry genes conferring various functions, such as resistance to antibiotics, that can enhance the fitness of their hosts and that contribute to their maintenance in bacterial populations. Taken as a whole, IMEs are probably major contributors to bacterial evolution
Vers une approche intégrée d'annotation des ICE et IME dans les génomes bactériens
International audienc
Effects of rodA and pbp2b Disruption on Cell Morphology and Oxidative Stress Response of Streptococcus thermophilus CNRZ368
Insertional mutagenesis was used to isolate clones from Streptococcus thermophilus CNRZ368 that were modified in their abilities to tolerate oxidative stress. During this process, two menadione-sensitive clones (6G4 and 18C3) were found to display abnormal cell morphologies and distorted chain topologies and were further studied. Molecular characterization of both 6G4 and 18C3 mutants indicated that they were disrupted in open reading frames homologous to rodA and pbp2b, respectively. Both genes encoded proteins in Escherichia coli that were described as being implicated in peptidoglycan synthesis during the process of cell elongation and to function in determining the rod shape of the cell. This work reports a possible connection between peptidoglycan biosynthesis and oxidative stress defense in S. thermophilus CNRZ368
Characterisation of Streptococcus thermophilus CNRZ368 oxidative stress-resistant mutants: involvement of a potential Rgg-like transcriptional regulator
International audienceEight mutants of Streptococcus thermophilus CNRZ368 presenting a menadione-resistant phenotype were selected and the locus mutated in each mutant was identified. Among these clones, 5 were disrupted in a gene of unknown function, 2 were impaired in genes involved in cellular metabolism and the last one (the mutant 15H7) was disrupted in rgg encoding a putative transcriptional regulator. To determine if rgg could be a key regulator of the superoxide defence of S. thermophilus, the 15H7 mutant was further characterised. The results from our work indicate that the rgg gene, mutated in the 15H7 genome, is a pseudogene composed of 2 ORFs (rggA and rggB) that are potentially translated in two different frames. Moreover, transcription analysis demonstrated the existence of a transcript containing rggA, rggB and orf2. Construction of rggA, rggB and orf2 mutants and their phenotypic analysis confirmed the involvement of rggA and rggB in the oxidative stress response. The question of the role of the pseudogene rgg is still open.Caractérisation de mutants résistants au stress oxydant chez Streptococcus thermophilus CNZR368 : implication d'un régulateur transcriptionnel putatif de type Rgg. Huit mutants de Streptococcus thermophilus CNRZ368, présentant un phénotype de résistance à la ménadione ont été sélectionnés et le locus muté a été identifié pour chacun d'eux. Parmi ces clones, 5 sont interrompus dans un gÚne de fonction inconnue, 2 sont mutés dans des gÚnes impliqués dans le métabolisme cellulaire et un autre (15H7) est interrompu dans le gÚne rgg codant potentiellement un régulateur transcriptionnel. Afin de déterminer si rgg est un régulateur majeur de la défense contre le stress superoxyde le mutant 15H7 a été plus amplement étudié. Les résultats de notre travail indiquent que le gÚne rgg, muté dans le génome de 15H7, est un pseudogÚne composé de deux ORF (rggA and rggB) potentiellement traduites dans deux phases de lecture différentes. De plus, l'analyse transcriptionelle démontre l'existence d'un transcrit englobant rggA, rggB et orf2. La construction des mutants rggA, rggB et orf2 ainsi que l'analyse de leur phénotype confirme l'implication de rggA et rggB dans la réponse contre le stress oxydant. La question concernant le rÎle du pseudogÚne rgg est toujours ouverte
Identification of Streptococcus thermophilus CNRZ368 Genes Involved in Defense against Superoxide Stress
To better understand the defense mechanism of Streptococcus thermophilus against superoxide stress, molecular analysis of 10 menadione-sensitive mutants, obtained by insertional mutagenesis, was undertaken. This analysis allowed the identification of 10 genes that, with respect to their putative functions, were classified into five categories: (i) those involved in cell wall metabolism, (ii) those involved in exopolysaccharide translocation, (iii) those involved in RNA modification, (iv) those involved in iron homeostasis, and (v) those whose functions are still unknown. The behavior of the 10 menadione-sensitive mutants exposed to heat shock was investigated. Data from these experiments allowed us to distinguish genes whose action might be specific to oxidative stress defense (tgt, ossF, and ossG) from those whose action may be generalized to other stressful conditions (mreD, rodA, pbp2b, cpsX, and iscU). Among the mutants, two harbored an independently inserted copy of pGh9:ISS1 in two loci close to each other. More precisely, these two loci are homologous to the sufD and iscU genes, which are involved in the biosynthesis of iron-sulfur clusters. This region, called the suf region, was further characterized in S. thermophilus CNRZ368 by sequencing and by construction of ÎsufD and iscU(97) nonpolar mutants. The streptonigrin sensitivity levels of both mutants suggest that these two genes are involved in iron metabolism
A glimpse into an unexplored world: very high diversity of integrative and mobilizable elements in Streptococci
International audienc
Prevalence and mobility of integrative and conjugative elements within a Streptomyces natural population
International audienceHorizontal Gene Transfer (HGT) is a powerful force generating genomic diversity in bacterial populations. HGT in Streptomyces is in large part driven by conjugation thanks to plasmids, Integrative and Conjugative elements (ICEs) and Actinomycete ICEs (AICEs). To investigate the impact of ICE and AICE conjugation on Streptomyces genome evolution, we used in silico and experimental approaches on a set of 11 very closely related strains isolated from a millimeter scale rhizosphere population. Through bioinformatic searches of canonical conjugation proteins, we showed that AICEs are the most frequent integrative conjugative elements, with the central chromosome region being a hotspot for integrative element insertion. Strains exhibited great variation in AICE composition consistent with frequent HGT and/or gene loss. We found that single insertion sites can be home to different elements in different strains (accretion) and conversely, elements belonging to the same family can be found at different insertion sites. A wide variety of cargo genes was present in the AICEs with the potential to mediate strain-specific adaptation (e.g., DNA metabolism and resistance genes to antibiotic and phages). However, a large proportion of AICE cargo genes showed hallmarks of pseudogenization, consistent with deleterious effects of cargo genes on fitness. Pock assays enabled the direct visualization of conjugal AICE transfer and demonstrated the transfer of AICEs between some, but not all, of the isolates. Multiple AICEs were shown to be able to transfer during a single mating event. Although we did not obtain experimental evidence for transfer of the sole chromosomal ICE in this population, genotoxic stress mediated its excision from the chromosome, suggesting its functionality. Our results indicate that AICE-mediated HGT in Streptomyces populations is highly dynamic, with likely impact on strain fitness and the ability to adapt to environmental change