Antibiotic resistance (AR) is a pervasive crisis that is intricately woven into social and environmental systems. Its escalation is fueled by factors such overuse, poverty, climate change, and the heightened interconnectedness characteristic of our era of globalization. In this dissertation, the impact of antibiotic usage is addressed from the perspective of wastewater-based surveillance (WBS) at the wastewater treatment plant (WWTP) and microbial ecology. Antibiotic usage and contamination was found to influence the prevalence of antibiotic resistance genes (ARGs) and resistant bacteria in both lab-scale and full-scale wastewater treatment settings. Through application of novel bioinformatic approaches developed herein, metagenomics revealed associations between sewage-associated microbes and community antibiotic use that were in part mediated by microbial ecological processes and horizontal gene transfer (HGT). In sum, this dissertation increases the arsenal of bioinformatic tools for AR surveillance in wastewater environments and advances knowledge with respect to the contribution of antibiotic use to the spread of antibiotic resistance at the community-scale. Three studies served to evaluate and/or develop bioinformatic resources for molecular characterization of AR in wastewater. Hybrid assembly combining emerging long read DNA sequencing and short read sequencing was evaluated and found to improve accuracy relative to assembly of long or short reads alone. A novel database of mobile genetic element (MGE) marker genes, mobileOG-db, was compiled in order to address short-comings with pre-existing resources. A pipeline for detecting HGT in metagenomes, Kairos, was created in order to facilitate the detection of HGT in metagenome assemblies which greatly amplified coverage of ARGs. In Chapter 5, a lab-scale study of WWTP bioreactors revealed that elevated antibiotic contamination was correlated with increased prevalence of corresponding ARGs. In addition, multiple in situ HGT events of ARGs encoding resistance to the elevated antibiotics were predicted, including one HGT event likely mediated by a novel bacteriophage. In Chapter 6, influent and effluent from a full-scale municipal WWTP were collected twice-weekly for one year and subjected to deep shotgun metagenomic sequencing. In parallel, collaboration with clinicians enabled statistical modeling of antibiotic usage and resistance, revealing associations between antibiotic prescriptions patterns in the region and resistance at the WWTP. Finally, Chapter 7 details bioinformatic recovery of diverse extended spectrum beta-lactamase gene recovery from the influent and effluent metagenomes, shedding light on the dynamics of circulating resistance genes. In sum, this dissertation identifies bioinformatic evidence for the selection of AR in wastewater environments as a result of antibiotic use in the community and advances hypotheses for explaining the mechanisms of the observed phenomena. / Doctor of Philosophy / Antibiotics are key lifesaving drugs that have dramatically improved life expectancy throughout the 20th and 21st centuries. However, there has been an increased incidence of resistance among many important bacterial pathogens in recent decades. The more antibiotics are used, the more chance that resistant bacteria can evolve, survive, and spread. Outpatient care accounts for the vast majority of therapeutic antibiotic use, with more than 200 million prescriptions written for antibiotics in 2021 in the United States. While performing a vital function in combatting disease, oral antibiotics can inadvertently harm the resident microbes of the intestinal tract (i.e., the gut microbiome) by decreasing the diversity of the microbes present and increasing the number of resistant bacteria. At a community level, antibiotic usage also has the potential to induce increased prevalence of antibiotics and antibiotic resistant bacteria in the environment as well, primarily via human excreta (urine and feces). Wastewater represents a key interface between human-derived contaminants and the environment. In regions with centralized wastewater management, antibiotics- and resistant bacteria-containing excreta are typically transported via sewage conveyance systems to a wastewater treatment plant (WWTP). At the WWTP, diverse microbes interact with and degrade various organic contaminants in a series of processes combining physical, chemical, and biological treatments. Due to the intermingling of environmental microbes, antibiotics, and antibiotic resistant bacteria, wastewater is increasingly being recognized as an important venue for antibiotic resistance surveillance and for potential interventions. Awareness of wastewater-based surveillance and epidemiology has surged as a result of the COVID-19 pandemic and such efforts are enshrined in the National COVID-19 Preparedness Plan. However, such a task is fundamentally more challenging for antibiotic resistance than for SARS-CoV-2, as it comprises multiple bacterial strains, antibiotic resistance genes, and resistance mechanisms. In this respect, DNA sequencing of wastewater, i.e., "metagenomics," holds promise as a broad monitoring tool with an unprecedented degree of biological granularity. In this dissertation, we address the impact of antibiotic usage at the WWTP from the perspective of wastewater-based surveillance. We evaluate antibiotic usage at the community-scale as a selective force among bacteria inhabiting WWTPs and identify microbial interactions that influence the escape of resistant bacteria in the effluent. A field-study of wastewater entering the WWTP and cleaned effluent water discharged by the WWTP revealed certain antibiotics and corresponding forms of antibiotic resistance were particularly prone to proliferation in the WWTP. Novel bioinformatic tools were developed and applied to the study of wastewater to reveal these associations. In sum, this dissertation advances knowledge of wastewater as both a mediator of environmental health and as a reflection of community-health in the form of antibiotic resistance.

The antimicrobial resistance (AMR) crisis, where new antibiotic discovery is not keeping pace with the emergence of resistant pathogens, is driven by mobile genetic elements (MGEs). MGEs can autonomously transfer between bacteria, along with AMR genes. The widespread use of antibiotics in the clinic, in agriculture, and animal husbandry, has accelerated the MGE-mediated transfer of AMR genes in the environment. However, despite playing such an important role in the AMR crisis, the dynamics and mechanisms behind the transmission of genes are poorly understood. Furthermore, which natural and man-made compounds inhibit or promote their movement in these environments is unknown. One method to combat the rise in AMR is to identify small molecules as probes to understand the molecular basis of transmission and apply this information to prevent MGE-mediated resistance dissemination. Since conjugation is the main mechanism for AMR gene transfer, targeting MGEs that use conjugation, such as conjugative plasmids (e.g. Tn1549) and conjugative transposons (e.g. F plasmid), has the potential to prevent the emergence of multi-drug resistant pathogens. In this work, a high throughput assay modeled after Tn1549 excision was screened against a library of known bioactive compounds to find modulators of the integrase and excisionase activity. Several fluoroquinolone antibiotics including ciprofloxacin were identified as dose-dependent inhibitors of excision, which acted by changing supercoiling levels in the cell. Ciprofloxacin enhanced conjugation frequency of Tn1549 at sub-MIC concentrations relative to an untreated control and inhibited conjugation frequency at higher concentrations. A second project was focused on a high throughput conjugation assay based on the separation of the lux operon between a donor and recipient cell, such that only transconjugants produce luminescence to reflect active gene transfer. This work furthers our understanding of the development of assays to target MGEs and screening for inhibitors of their movement. / Thesis / Master of Science (MSc) / Antibiotics are small molecules that cure bacterial infections. However, their efficacy is fading as a result of the ability of mobile genetic elements (MGEs) to spread antimicrobial resistance genes between bacteria. Conjugative plasmids (CPs) and conjugative transposons (CTns) are two of the major types of MGEs that contribute to the dissemination of antimicrobial resistance in pathogens. The goal of this research is to search for inhibitors of CTns and CPs in order to prevent the emergence of multi-drug resistant bacteria. High throughput assays were designed to model both a CTn (Tn1549) and a CP (F plasmid) to find small molecules targeting their movement. A screen of the Tn1549 excision assay identified fluoroquinolone antibiotics that inhibit excision in a dose-dependent manner and indirectly inhibit the integrase used to excise the CTn. Ciprofloxacin, a fluoroquinolone, inhibited the conjugation frequency of Tn1549. Future work will focus on identifying new inhibitors of these MGEs and their characterization.

O reconhecimento da resistência antimicrobiana como um fenômeno emergente em saúde pública, tem constituído um problema em nível mundial. O abuso na utilização de antibióticos na medicina humana e veterinária, e na agricultura, tem originado incremento na diversidade de micro-organismos resistentes, refletindo em falha terapêutica. Os mecanismos de resistência a antibióticos em micro-organismos são mediados principalmente por genes adquiridos de DNA exógeno. A dinâmica da transferência horizontal é realizada por meio de elementos genéticos móveis que carregam genes de resistência. A ampla distribuição deste tipo de estruturas, como o elemento SXT, isolado inicialmente em V. cholerae, tem contribuído para a disseminação de complexos específicos clonais em determinadas áreas geográficas. Este estudo pioneiro no Brasil pesquisou a presença de elementos SXT, em espécies bacterianas do grupo das gama proteobactérias em espécies ambientais, determinou suas características estruturais e funcionais, incluindo genes de resistência a antibióticos, bem como a sensibilidade aos antibióticos dentre os isolados bacterianos que os abrigam. O resultado foi a classificação de 43 elementos SXT obtidos no Brasil, através da comparação com aqueles descritos na literatura. Dentre os elementos SXT obtidos, quatro são albergados por Morganella morganii, fato inédito na literatura. O conhecimento da evolução bacteriana constitui importante ferramenta para estabelecer estratégias eficazes de controle e tratamento de infecções, sem aumentar a pressão seletiva sobre os micro-organismos, bem como instrumento preciso e de grande importância para subsidiar estudos epidemiológicos. / Recognition of antimicrobial resistance as an emerging phenomenon in public health has been a problem worldwide. The abuse in the use of antibiotics in human and veterinary medicine, and agriculture, has caused an increase in the diversity of resistant microorganisms, reflecting in treatment failure. The mechanisms of antibiotic resistance in microorganisms are primarily mediated by genes acquired from exogenous DNA. The dynamics of the horizontal transfer is performed by mobile genetic elements which carry resistance genes. The wide distribution of these structures, such as the SXT element originally isolated from V. cholerae, has contributed to the spread of specific clonal complexes in certain geographical areas. This pioneering study in Brazil researched the presence of SXT elements in the group of bacterial species in environmental gamma-proteobacteria species, determined their structural and functional characteristics, including genes for resistance to antibiotics and the antibiotic susceptibility among bacterial isolates that harbor them. The result was the classification of 43 SXT elements found in Brazil, by comparison with those found in the literature. Among the SXT elements found, four are sheltered by Morganella morganii, unprecedented in the literature. Knowledge of bacterial evolution is an important to establish effective strategies to control and treat infections without increasing the selective pressure on microorganisms, as well as a precise instrument and very important tool to support epidemiological studies.

O reconhecimento da resistência antimicrobiana como um fenômeno emergente em saúde pública, tem constituído um problema em nível mundial. O abuso na utilização de antibióticos na medicina humana e veterinária, e na agricultura, tem originado incremento na diversidade de micro-organismos resistentes, refletindo em falha terapêutica. Os mecanismos de resistência a antibióticos em micro-organismos são mediados principalmente por genes adquiridos de DNA exógeno. A dinâmica da transferência horizontal é realizada por meio de elementos genéticos móveis que carregam genes de resistência. A ampla distribuição deste tipo de estruturas, como o elemento SXT, isolado inicialmente em V. cholerae, tem contribuído para a disseminação de complexos específicos clonais em determinadas áreas geográficas. Este estudo pioneiro no Brasil pesquisou a presença de elementos SXT, em espécies bacterianas do grupo das gama proteobactérias em espécies ambientais, determinou suas características estruturais e funcionais, incluindo genes de resistência a antibióticos, bem como a sensibilidade aos antibióticos dentre os isolados bacterianos que os abrigam. O resultado foi a classificação de 43 elementos SXT obtidos no Brasil, através da comparação com aqueles descritos na literatura. Dentre os elementos SXT obtidos, quatro são albergados por Morganella morganii, fato inédito na literatura. O conhecimento da evolução bacteriana constitui importante ferramenta para estabelecer estratégias eficazes de controle e tratamento de infecções, sem aumentar a pressão seletiva sobre os micro-organismos, bem como instrumento preciso e de grande importância para subsidiar estudos epidemiológicos. / Recognition of antimicrobial resistance as an emerging phenomenon in public health has been a problem worldwide. The abuse in the use of antibiotics in human and veterinary medicine, and agriculture, has caused an increase in the diversity of resistant microorganisms, reflecting in treatment failure. The mechanisms of antibiotic resistance in microorganisms are primarily mediated by genes acquired from exogenous DNA. The dynamics of the horizontal transfer is performed by mobile genetic elements which carry resistance genes. The wide distribution of these structures, such as the SXT element originally isolated from V. cholerae, has contributed to the spread of specific clonal complexes in certain geographical areas. This pioneering study in Brazil researched the presence of SXT elements in the group of bacterial species in environmental gamma-proteobacteria species, determined their structural and functional characteristics, including genes for resistance to antibiotics and the antibiotic susceptibility among bacterial isolates that harbor them. The result was the classification of 43 SXT elements found in Brazil, by comparison with those found in the literature. Among the SXT elements found, four are sheltered by Morganella morganii, unprecedented in the literature. Knowledge of bacterial evolution is an important to establish effective strategies to control and treat infections without increasing the selective pressure on microorganisms, as well as a precise instrument and very important tool to support epidemiological studies.

Bacteria are social organisms which participate in multiple cooperative and group behaviours. They moreover have peculiar genetic systems, as they often bear mobile genetic elements like plasmids, molecular symbionts that are the cause of widespread horizontal gene transfer and play a large role in bacterial evolution. Both cooperation and horizontal transfer have consequences for human health: cooperative behaviours are very often involved in the virulence of pathogens, and horizontal gene transfer leads to the spread of antibiotic resistance. The evolution of plasmid transfer has mainly been analyzed in terms of infectious benefits for selfish mobile elements. However, chromosomal genes can also modulate horizontal transfer. A huge diversity in transfer rates is observed among bacterial isolates, suggesting a complex co-evolution between plasmids and hosts. Moreover, plasmids are enriched in genes involved in social behaviours, and so could play a key role in bacterial cooperative behaviours. We study here the coevolution of gene mobility and sociality in bacteria. To investigate the selective pressures acting on plasmid transfer and public good production, we use both mathematical modelling and a synthetic system that we constructed where we can independently control public good cooperation and plasmid conjugation in Escherichia coli. We first show experimentally that horizontal transfer allows the specific maintenance of public good alleles in a structured population by increasing relatedness at the gene-level. We further demonstrate experimentally and theoretically that this in turn allows for second-order selection of transfer ability: when cooperation is needed, alleles promoting donor and recipient abilities for public good traits can be selected both on the plasmid and on the chromosome in structured populations. Moreover, donor ability for private good traits can also be selected on the chromosome, provided that transfer happens towards kin. The interactions between transfer and cooperation can finally lead to an association between transfer and public good production alleles, explaining the high frequency of genes related to cooperation that are located on plasmids. Globally, these results provide insight into the mechanisms maintaining cooperation in bacteria, and may suggest ways to target cooperative virulence.

Nos connaissances sur la diversité virale associée aux micro-organismes présents dans les sources hydrothermales océaniques profondes restent encore limitées. Seules quelques études concernant l’abondance virale et l’impact de ceux-ci sur la mortalité microbienne dans ces écosystèmes sont disponibles. En effet, seuls 6 bactériovirus et 2 archéovirus provenant de ces écosystèmes ont été caractérisés à l’heure actuelle. Les deux archéovirus infectent des archées anaérobies hyperthermophiles appartenant à l’ordre des Thermococcales et ont été décrits au laboratoire.Afin d’étendre nos connaissances sur la diversité virale associée aux micro-organismes colonisant ces environnements, il a été décidé d’élargir les recherches à l’ordre bactérien des Thermotogales. Cet ordre bactérien est composé de bactéries chimio-organotrophes anaérobies en majorité thermophiles ou hyperthermophiles. De nombreux transferts latéraux de gènes ont contribué à l’histoire évolutive des Thermotogales supposant une forte implication des virus dans celle-ci. Ces travaux de thèse ont permis la caractérisation fonctionnelle et génomique de deux nouveaux siphovirus, MCV1 et MCV2, infectant deux souches de Marinitoga camini. Ces souches ont été isolées de deux sites hydrothermaux profonds (Menez Gwen et Lucky Strike) au niveau de la dorsale médio-atlantique. Ces virus mettent en oeuvre un cycle lysogénique avec une production basale sans induction relativement haute (>107 virions/ml). Une comparaison de ces deux génomes viraux à celui de MPV1, virus précédemment isolé de Marinitoga piezophila, a été réalisée, révélant la présence de nombreuses similarités. Un core genome de 35 ORFs partagé par ces trois génomes a été identifié, incluant des protéines impliquées dans le métabolisme de l’ADN, l’assemblage des virions et le cycle lysogénique. Des protéines hypothétiques ont aussi été identifiées parmi ces gènes communs, elles portent donc probablement des fonctions importantes pour ces bactériovirus. Par ailleurs, 60% des gènes de ces virus ayant une correspondance dans les bases de données, après exclusion des Thermotogales, partagent des similarités avec lesFirmicutes et les bactériovirus qui leurs sont associés. Le génome d’une autre Thermotogales, Thermosipho sp. AT1244-VC14 a été étudié ainsi que son système CRISPR-cas. Ces résultats indiquent que cette souche, qui porte un système CRISPR-cas qui semble complet et fonctionnel, a probablement déjà été infectée par MCV1, MCV2 ou un virus similaire. Ces travaux permettent d’étendre nos connaissances sur les virus portés par les bactéries du phylum Thermotogae, encore peu décrits à ce jour. Les éléments génétiques mobiles associés à ce phylum sont particulièrement intéressants car ils ont probablement eu un impact important dans l’évolution de ces communautés microbiennes ainsi que dans leur adaptation aux conditions physico-chimiques extrêmes et fluctuantes présentes dans les écosystèmes qu’elles colonisent. / Our knowledge of the viral diversity associated to microorganisms inhabiting the deep-sea hydrothermal vents is still limited. Only a few studies have focused on viral abundance and impact on microbial mortality within these ecosystems. A limited number of viruses (6 bacterioviruses and 2 archaeoviruses) were isolated from these environments and characterized. Two viruses associated to hyperthermophilic anaerobic Archaea, from the Thermococcales order, have been described in our laboratory. In order to deepen our knowledge on the viral diversity of these extreme environments, we have extended our investigation to the bacterial order of Thermotogales. This order is composed of anaerobic chemoorganotrophic bacteria that are, for the most part, hyper/thermophilic. Numerous lateral gene transfers have contributed to the evolutionary history of the Thermotogales, implying the potential involvement of viruses. Here, we will report the characterization of two new siphoviruses MCV1 and MCV2 that infect two strains of Marinitoga camini. Those bacterial strains were isolated from two deep-sea hydrothermal vents sites (Menez Gwen and Lucky strike) in the Mid Atlantic Ridge. These viruses are temperate with a high basal production of virions (>107 virions/mL). Comparative genomics with MPV1, a virus isolated from M. piezophila, was performed and show that those bacterioviruses share numerous similarities. A set of “core genes” shared by all these three viruses was identified and includes proteins involved in DNA metabolism, head and tail assembly and lysogenic cycle. Shared hypothetical proteins were also identified, suggesting that these unknown proteins probably provide important functions for these viruses. Interestingly, for genes with blastp matches in Genbank, over 60% have their top matches, outside the Thermotoga, to genes from Firmicutes and bacterioviruses associated to Firmicutes. We also analyzed the genome of Thermosipho sp. 1244 and studied his CRISPR-cas system. Our results indicated that thisThermosipho strain, with a complete and functional CRISPR-cas system, had already been infected by MCV1, MCV2 or a similar virus. The analyses presented here extend our knowledges about these newly discovered viruses in the deeply branching bacterial phylum Thermotogae. This bacterial order and associated mobile genetic elements are significant for addressing long-term evolutionary adaptation to fluctuant and extreme physicochemical conditions.

Les bactéries sont des organismes extrêmement sociaux, qui présentent de multiples comportements de coopération. De plus, les génomes bactériens sont caractérisés par la présence de nombreux éléments génétiques mobiles, tels que les plasmides. Ces éléments mobiles sont la cause de transferts génétiques horizontaux, et jouent un rôle important dans l'évolution bactérienne. La coopération et le transfert horizontal ont tous deux des conséquences importantes sur la santé humaine: des comportements coopératifs sont souvent à l'origine de propriétés de virulence chez les bactéries pathogènes, et le transfert horizontal entraîne la dissémination de gènes de résistance aux antibiotiques. L'évolution du transfert horizontal a jusqu'ici été analysée essentiellement en termes de bénéfices infectieux apportés à des éléments génétiques égoïstes. Cependant, le taux de transfert des plasmides est extrêmement variable et partiellement contrôlé par les gènes des bactéries hôtes, suggérant une co-évolution complexe entre hôtes et plasmides. De plus, les plasmides sont particulièrement riches en gènes liés à des comportements coopératifs, et semblent donc jouer un rôle-clé dans les phénomènes de socialité bactérienne. Ce travail porte sur la coévolution entre mobilité génétique et socialité chez les bactéries. Nous analysons ici les pressions de sélection agissant sur le transfert de plasmides et la production de biens publics, à l'aide de modèles mathématiques et d'un système synthétique que nous avons construit chez Escherichia coli, dans lequel nous pouvons contrôler indépendamment la coopération et la conjugaison. Dans un premier temps, nous montrons expérimentalement que le transfert horizontal favorise le maintien de la coopération dans une population structurée, en augmentant la sélection de parentèle agissant au niveau des gènes transférés. Dans un second temps, nous montrons expérimentalement et théoriquement que l'échange génétique lui-même peut être sélectionné: les bactéries transférant des plasmides codant pour des biens publics sont favorisées dans une population structurée. Le transfert de gènes codant pour des biens privés peut également être sélectionné, à condition que ce transfert s'effectue entre bactéries apparentées. Finalement, ces interactions entre transfert horizontal et coopération peuvent mener à une association entre allèles de coopération et de transfert, expliquant la fréquence élevée de gènes sociaux situés sur des plasmides.Ces résultats permettent de mieux comprendre le maintien de comportements coopératifs chez les bactéries, et suggèrent des moyens de cibler certains cas de virulence bactérienne. / Bacteria are social organisms which participate in multiple cooperative and group behaviours. They moreover have peculiar genetic systems, as they often bear mobile genetic elements like plasmids, molecular symbionts that are the cause of widespread horizontal gene transfer and play a large role in bacterial evolution. Both cooperation and horizontal transfer have consequences for human health: cooperative behaviours are very often involved in the virulence of pathogens, and horizontal gene transfer leads to the spread of antibiotic resistance. The evolution of plasmid transfer has mainly been analyzed in terms of infectious benefits for selfish mobile elements. However, chromosomal genes can also modulate horizontal transfer. A huge diversity in transfer rates is observed among bacterial isolates, suggesting a complex co-evolution between plasmids and hosts. Moreover, plasmids are enriched in genes involved in social behaviours, and so could play a key role in bacterial cooperative behaviours. We study here the coevolution of gene mobility and sociality in bacteria. To investigate the selective pressures acting on plasmid transfer and public good production, we use both mathematical modelling and a synthetic system that we constructed where we can independently control public good cooperation and plasmid conjugation in Escherichia coli. We first show experimentally that horizontal transfer allows the specific maintenance of public good alleles in a structured population by increasing relatedness at the gene-level. We further demonstrate experimentally and theoretically that this in turn allows for second-order selection of transfer ability: when cooperation is needed, alleles promoting donor and recipient abilities for public good traits can be selected both on the plasmid and on the chromosome in structured populations. Moreover, donor ability for private good traits can also be selected on the chromosome, provided that transfer happens towards kin. The interactions between transfer and cooperation can finally lead to an association between transfer and public good production alleles, explaining the high frequency of genes related to cooperation that are located on plasmids. Globally, these results provide insight into the mechanisms maintaining cooperation in bacteria, and may suggest ways to target cooperative virulence.

Mobile genetic elements are occupied by this project. It is aimed at their features, which can be used for their detection. It also deals with issue of conversion of symbolic sequence into numerical form. Classifications of mobile genetic elements are explained, basic types of mobile genetic sequences are described, and principles of numerical maps and detection in symbolic represetation are also clarified. Conversion of symbolic genetical sequences by chosen numerical map and calculation of normalized correlation values for set of mobile genetic elements are compiled. Analysis of the mobile genetic elements properties is performed for design of detector. The library of themes is created at the end for usage by designed detector.

Le transfert horizontal participe à l'évolution rapide des génomes bactériens. Les éléments intégratifs et conjugatifs (ICE) sont des îlots génomiques capables de se transférer par conjugaison vers une bactérie receveuse. Streptococcus agalactiae est une bactérie pathogène opportuniste qui est à l'origine de problèmes sanitaires et économiques majeurs. Des études ont révélé la présence de nombreux ICE chez cette espèce, notamment à l'extrémité 3' d?un gène codant un ARNtLys. La fonctionnalité de l'ICE intégré à ce locus chez la souche 515 de S. agalactiae a été démontrée. Les fonctions véhiculées par ICE_515_tRNALys et pouvant conférer un avantage adaptatif ont été caractérisées et leur transfert vers d'autres espèces a été évalué. Les résultats ont montré que l'ICE confère à S. agalactiae des propriétés d'adhésion à l'hôte et de formation de biofilm et pourrait être impliqué dans l'agrégation cellulaire. Un antigène I/II codé par l'ICE est impliqué dans des phénotypes d'adhésion. De plus, un nouveau facteur co-hémolytique de type CAMP, codé par l'ICE et qui pourrait être impliqué dans la virulence et la survie des souches, a été caractérisé. La fonctionnalité de ces facteurs de virulence chez des espèces bactériennes pathogènes et non pathogènes a été établie. Les travaux ont également révélé la prévalence et la dynamique évolutive des ICE appartenant à la famille d'ICE_515_tRNALys et des fonctions adaptatives codées par ces éléments chez plusieurs espèces de streptocoques. En conclusion, les ICE de la famille d'ICE_515_tRNALys représentent des vecteurs de traits phénotypiques importants pour la virulence et la survie chez les streptocoques / Horizontal gene transfer is a rapid mechanism of evolution. Integrative and conjugative elements (ICEs) are genomic islands which can transfer by conjugation to recipient bacteria. Streptococcus agalactiae is a human and animal opportunistic pathogen that is responsible for major health and economic problems. Studies revealed the presence of numerous ICEs in S. agalactiae, in particular at the 3' end of a tRNALys encoding gene. The functionality of the element present in strain S. agalactiae 515 was demonstrated and was thus chosen as a model for this study. This work focused on the characterization of adaptive and virulence functions encoded by ICE_515_tRNALys and their transfer to other species. Results indicated that this ICE confers adhesion properties to host, increases biofilm formation and may be involved in cell aggregation. A new protein belonging to the antigens I/II family is involved in fibronectin binding and contributes to the biofilm phenotype. In addition, a new co-hemolytic CAMP factor encoded by ICE_515_tRNALys, which could be involved in virulence and bacterial survival, was identified and characterized. These virulence factors are functional in other bacterial species. This work also revealed the prevalence and evolutionary dynamics of ICE belonging to the family of ICE_515_tRNALys and adaptive functions encoded by these elements in several species of streptococci. In conclusion, ICEs of the ICE_515_tRNALys family represent vectors of phenotypic features important for virulence and survival in streptococci

Les éléments intégratifs conjugatifs (ICE) sont des éléments génétiques mobiles se transférant horizontalement d’une bactérie à une autre. Les ICE sont porteurs de gènes adaptatifs pouvant significativement améliorer le fitness de la bactérie hôte et permettre son adaptation à de nouvelles niches écologiques. Lors de ces travaux, des ICE apparentés à ICESt3 ont été retrouvés chez la bactérie commensale et pathogène opportuniste Streptococcus salivarius. Les analyses in silico réalisées ont démontré la diversité des ICE de cette famille, notamment au niveau de leurs modules de recombinaison, de régulation mais aussi de leurs gènes adaptatifs potentiellement mis à disposition de la communauté microbienne orale et digestive de l’Homme. La fonctionnalité de deux de ces ICE a été mise en évidence expérimentalement à travers l’évaluation de la capacité de ces éléments à se transférer intra- et inter-spécifiquement. Ces travaux ont également permis l’identification de facteurs d’hôte influençant la mobilité d’ICESt3, révélant ainsi l’importance, pour le transfert et l’acquisition de cet ICE, des molécules de surface telles que les lipoprotéines, les acides téichoïques et les exopolysaccharides. En conclusion, il a été démontré que les éléments de la famille ICESt3 participent à l’évolution du génome chez différentes espèces de streptocoques et aux échanges génétiques entre bactéries issues de l’alimentation et bactéries de la flore digestive humaine. Enfin, ces travaux ont contribué à une meilleure compréhension des mécanismes et des facteurs d’hôte influençant la mobilité de ces éléments génétiques mobiles / Integrative Conjugative Elements (ICEs) are mobile genetic elements that can be horizontally transferred from a bacterium to another, eventually regardless of the species or any other classification, allowing them to benefit from a broad host spectrum. ICEs can carry adaptive genes that can significantly improve the bacterial fitness and allow its adaptation to new ecological niches. In this work, ICEs related to ICESt3 were found in the commensal and opportunistic pathogen Streptococcus salivarius. In silico analysis highlighted the diversity of the ICESt3 family within this species, especially concerning their recombination and regulation modules, but also their adaptive genes likely available for the oral and digestive microbial community of the human host. The functionality of two ICEs found in S. salivarius was experimentally confirmed through their ability to transfer in intra- and interspecific manners. This work also allowed the identification of host factors affecting ICESt3 mobility, and revealed the importance, for the transfer and the acquisition of this ICE, of cell surface molecules such as lipoproteins, teichoic acids and exopolysaccharides. In conclusion, this thesis allowed expanding the knowledge regarding the mobile genetic elements of the ICESt3 family. This work demonstrated that these elements contribute to genome evolution of different streptococci species and gene exchanges between bacteria originated from food and the human gut flora. Finally, this study contributes to a better comprehension of the mechanisms and host factors influencing the mobility of these mobile genetic elements