A New Perspective on Listeria monocytogenes Evolution

Listeria monocytogenes is a model organism for cellular microbiology and host–pathogen interaction studies and an important food-borne pathogen widespread in the environment, thus representing an attractive model to study the evolution of virulence. The phylogenetic structure of L. monocytogenes was determined by sequencing internal portions of seven housekeeping genes (3,288 nucleotides) in 360 representative isolates. Fifty-eight of the 126 disclosed sequence types were grouped into seven well-demarcated clonal complexes (clones) that comprised almost 75% of clinical isolates. Each clone had a unique or dominant serotype (4b for clones 1, 2 and 4, 1/2b for clones 3 and 5, 1/2a for clone 7, and 1/2c for clone 9), with no association of clones with clinical forms of human listeriosis. Homologous recombination was extremely limited (r/m<1 for nucleotides), implying long-term genetic stability of multilocus genotypes over time. Bayesian analysis based on 438 SNPs recovered the three previously defined lineages, plus one unclassified isolate of mixed ancestry. The phylogenetic distribution of serotypes indicated that serotype 4b evolved once from 1/2b, the likely ancestral serotype of lineage I. Serotype 1/2c derived once from 1/2a, with reference strain EGDe (1/2a) likely representing an intermediate evolutionary state. In contrast to housekeeping genes, the virulence factor internalin (InlA) evolved by localized recombination resulting in a mosaic pattern, with convergent evolution indicative of natural selection towards a truncation of InlA protein. This work provides a reference evolutionary framework for future studies on L. monocytogenes epidemiology, ecology, and virulence

Metagenome-based diversity analyses suggest a significant contribution of non-cyanobacterial lineages to carbonate precipitation in modern microbialites

Frontiers in Microbiology 6 (2015): 797 This Document is Protected by copyright and was first published by Frontiers. All rights reserved. It is reproduced with permissionCyanobacteria are thought to play a key role in carbonate formation due to their metabolic activity, but other organisms carrying out oxygenic photosynthesis (photosynthetic eukaryotes) or other metabolisms (e.g., anoxygenic photosynthesis, sulfate reduction), may also contribute to carbonate formation. To obtain more quantitative information than that provided by more classical PCR-dependent methods, we studied the microbial diversity of microbialites from the Alchichica crater lake (Mexico) by mining for 16S/18S rRNA genes in metagenomes obtained by direct sequencing of environmental DNA. We studied samples collected at the Western (AL-W) and Northern (AL-N) shores of the lake and, at the latter site, along a depth gradient (1, 5, 10, and 15 m depth). The associated microbial communities were mainly composed of bacteria, most of which seemed heterotrophic, whereas archaea were negligible. Eukaryotes composed a relatively minor fraction dominated by photosynthetic lineages, diatoms in AL-W, influenced by Si-rich seepage waters, and green algae in AL-N samples. Members of the Gammaproteobacteria and Alphaproteobacteria classes of Proteobacteria, Cyanobacteria, and Bacteroidetes were the most abundant bacterial taxa, followed by Planctomycetes, Deltaproteobacteria (Proteobacteria), Verrucomicrobia, Actinobacteria, Firmicutes, and Chloroflexi. Community composition varied among sites and with depth. Although cyanobacteria were the most important bacterial group contributing to the carbonate precipitation potential, photosynthetic eukaryotes, anoxygenic photosynthesizers and sulfate reducers were also very abundant. Cyanobacteria affiliated to Pleurocapsales largely increased with depth. Scanning electron microscopy (SEM) observations showed considerable areas of aragonite-encrusted Pleurocapsa-like cyanobacteria at microscale. Multivariate statistical analyses showed a strong positive correlation of Pleurocapsales and Chroococcales with aragonite formation at macroscale, and suggest a potential causal link. Despite the previous identification of intracellularly calcifying cyanobacteria in Alchichica microbialites, most carbonate precipitation seems extracellular in this systemWe are grateful to Eleonor Cortés for help and good company during the field trip and to Eberto Novelo for helpful discussions at the UNAM lab. This research was funded by the European Research Council Grants ProtistWorld (PI PL-G., Grant Agreement no. 322669) and CALCYAN (PI KB, Grant Agreement no. 307110) under the European Union’s Seventh Framework Program and the RTP Génomique environnementale of the CNRS (project MetaStrom, PI DM

The Yin-Yang of the Green Fluorescent Protein:Impact on Saccharomyces cerevisiae stress resistance

International audienceAlthough fluorescent proteins are widely used as biomarkers (Yin), no study focuses on their influence on the microbial stress response. Here, the Green Fluorescent Protein (GFP) was fused to two proteins of interest in Saccharomyces cerevisiae. Pab1p and Sur7p, respectively involved in stress granules structure and in Can1 membrane domains. These were chosen since questions remain regarding the understanding of the behavior of S. cerevisiae facing different heat kinetics or oxidative stresses. The main results showed that Pab1p-GFP fluorescent mutant displayed a higher resistance than that of the wild type under a heat shock. Moreover, fluorescent mutants exposed to oxidative stresses displayed changes in the cultivability compared to the wild type strain. In silico approaches showed that the presence of the GFP did not influence the structure and so the functionality of the tagged proteins meaning that changes in yeast resistance were certainly related to GFP ROS-scavenging ability (Yang)

Genes but Not Genomes Reveal Bacterial Domestication of Lactococcus Lactis

BACKGROUND: The population structure and diversity of Lactococcus lactis subsp. lactis, a major industrial bacterium involved in milk fermentation, was determined at both gene and genome level. Seventy-six lactococcal isolates of various origins were studied by different genotyping methods and thirty-six strains displaying unique macrorestriction fingerprints were analyzed by a new multilocus sequence typing (MLST) scheme. This gene-based analysis was compared to genomic characteristics determined by pulsed-field gel electrophoresis (PFGE). METHODOLOGY/PRINCIPAL FINDINGS: The MLST analysis revealed that L. lactis subsp. lactis is essentially clonal with infrequent intra- and intergenic recombination; also, despite its taxonomical classification as a subspecies, it displays a genetic diversity as substantial as that within several other bacterial species. Genome-based analysis revealed a genome size variability of 20%, a value typical of bacteria inhabiting different ecological niches, and that suggests a large pan-genome for this subspecies. However, the genomic characteristics (macrorestriction pattern, genome or chromosome size, plasmid content) did not correlate to the MLST-based phylogeny, with strains from the same sequence type (ST) differing by up to 230 kb in genome size. CONCLUSION/SIGNIFICANCE: The gene-based phylogeny was not fully consistent with the traditional classification into dairy and non-dairy strains but supported a new classification based on ecological separation between "environmental" strains, the main contributors to the genetic diversity within the subspecies, and "domesticated" strains, subject to recent genetic bottlenecks. Comparison between gene- and genome-based analyses revealed little relationship between core and dispensable genome phylogenies, indicating that clonal diversification and phenotypic variability of the "domesticated" strains essentially arose through substantial genomic flux within the dispensable genome

Sunlight-Exposed Biofilm Microbial Communities Are Naturally Resistant to Chernobyl Ionizing-Radiation Levels

BACKGROUND: The Chernobyl accident represents a long-term experiment on the effects of exposure to ionizing radiation at the ecosystem level. Though studies of these effects on plants and animals are abundant, the study of how Chernobyl radiation levels affect prokaryotic and eukaryotic microbial communities is practically non-existent, except for a few reports on human pathogens or soil microorganisms. Environments enduring extreme desiccation and UV radiation, such as sunlight exposed biofilms could in principle select for organisms highly resistant to ionizing radiation as well. METHODOLOGY/PRINCIPAL FINDINGS: To test this hypothesis, we explored the diversity of microorganisms belonging to the three domains of life by cultivation-independent approaches in biofilms developing on concrete walls or pillars in the Chernobyl area exposed to different levels of radiation, and we compared them with a similar biofilm from a non-irradiated site in Northern Ireland. Actinobacteria, Alphaproteobacteria, Bacteroidetes, Acidobacteria and Deinococcales were the most consistently detected bacterial groups, whereas green algae (Chlorophyta) and ascomycete fungi (Ascomycota) dominated within the eukaryotes. Close relatives to the most radio-resistant organisms known, including Rubrobacter species, Deinococcales and melanized ascomycete fungi were always detected. The diversity of bacteria and eukaryotes found in the most highly irradiated samples was comparable to that of less irradiated Chernobyl sites and Northern Ireland. However, the study of mutation frequencies in non-coding ITS regions versus SSU rRNA genes in members of a same actinobacterial operational taxonomic unit (OTU) present in Chernobyl samples and Northern Ireland showed a positive correlation between increased radiation and mutation rates. CONCLUSIONS/SIGNIFICANCE: Our results show that biofilm microbial communities in the most irradiated samples are comparable to non-irradiated samples in terms of general diversity patterns, despite increased mutation levels at the single-OTU level. Therefore, biofilm communities growing in sunlight exposed substrates are capable of coping with increased mutation rates and appear pre-adapted to levels of ionizing radiation in Chernobyl due to their natural adaptation to periodical desiccation and ambient UV radiation

Diversity and microbial colonization process in biofilms on mineral substractes

Mes travaux de recherche ont eu pour but d’analyser la diversité des microorganismes des trois domaines du vivant présents dans des biofilms phototrophes exposés à l’air, se développant sur des substrats minéraux divers, afin d’essayer, d’une part, de répondre à des questions de diversité et de biogéographie et, d’autre part, d’étudier le processus de colonisation par le biais d’expériences d’exposition contrôlées.J’ai ainsi caractérisé, essentiellement par des approches moléculaires basées sur l'analyse des banques des gènes d'ARNr de la petite sous-unité (SSU rDNAs) et sur des analyses d'empreintes communautaires, la diversité microbienne (procaryote et eucaryote) formant des biofilms matures (exposés depuis plusieurs années) dans plusieurs sites géographiques en Irlande du Nord, en France et en Ukraine, dans la région de Chernobyl. Dans ces biofilms soumis à forte pression sélective, nous avons mis en évidence beaucoup de microorganismes hétérotrophes et phototrophes, mais avec une diversité relativement restreinte en comparaison à d’autres milieux comme les sols ou les systèmes aquatiques. Les archées étaient absentes. Les conditions environnementales auxquelles ce type de biofilm est constamment exposé comme l’irradiation, la dessiccation et la limitation des nutriments sélectionnent des microorganismes qui développent des stratégies pour s’adapter comme, entre autres, la production de pigments. Ce sont des microorganismes fréquemment retrouvés dans des milieux désertiques extrêmes et résistants aussi aux radiations ionisantes qui ont ainsi été identifiés, notamment des Deinococcales et des Actinobacteria, ou encore des champignons ascomycètes (Ascomycota). Parmi les organismes phototrophes, nous avons dénombré des Cyanobacteria, des algues vertes (Chlorophyta) et des Streptophyta. Nous avons mis en évidence que les facteurs environnementaux influencent la composition des biofilms. Toutefois, tandis que la composition de la communauté bactérienne est fortement dépendante de la nature du substrat ou elle se développe, la composition des communautés microbiennes eucaryotes dépend de la distance géographique. Nous avons également mené des expériences de colonisation en exposant un même substrat minéral dans trois sites géographiques en Irlande du Nord et en France. L'analyse de la diversité microbienne lors du processus de colonisation a révélé des changements importants dans la composition des communautés, que ce soit pour les procaryotes ou pour les eucaryotes avec, cependant, des comportements différents de ces deux groupes de microorganismes. Dans le cas des bactéries, on observe une transition des Gammaproteobacteria, qui dominent les temps 0-6 mois et qui correspondent vraisemblablement aux cellules inactives en dispersion, vers des Betaproteobacteria, Bacteroidetes, Alphaproteobacteria et Actinobacteria dans des phases successives de formation du biofilm. Par contre, dès leur détection sur le substrat minéral, les eucaryotes sont massivement dominés par des champignons ascomycètes et basidiomycètes, des algues vertes ainsi que d'autres composantes minoritaires comme des ciliés, étant détectées dans des stades plus tardifs. Nos résultats montrent que les organismes hétérotrophes sont pionniers dans la formation de ces biofilms, ce qui permet d'émettre l'hypothèse qu'ils facilitent l'installation des cyanobactéries et surtout des algues vertes. Ils montrent aussi que le processus d'assemblage des communautés bactériennes dépend du temps de colonisation, alors que le site géographique détermine celui des microorganismes eucaryotes. Ces différences majeures de comportement pourraient être expliquées par des modes de vie différents entre les organismes de ces deux grands groupes.The major objective of my PhD work was the analysis of the diversity of microorganisms from the three domains of life associated with phototrophic biofilms developing on different mineral substrates exposed outdoors. These studies aimed at answering questions about microbial diversity and biogeography and also at studying the colonization process through controlled exposure experiments. I have thus characterized, essentially by molecular methods based on small subunit (SSU) rRNA gene libraries and fingerprinting analyses the diversity of prokaryote and eukaryote microorganisms forming mature biofilms (exposed for several years) in various geographic sites in Northern Ireland, France and Ukraine, in the Chernobyl area. In these biofilms, subjected to strong selective pressure, we found many heterotrophic and phototrophic microorganisms, but their diversity was limited when compared to that of other environments such as soils or aquatic systems. Archaea were absent from all biofilms. The environmental conditions to which these biofilms are constantly exposed, such as irradiation, desiccation and nutrient limitation select for organisms that develop particular adaptive strategies including, among others, pigment production. The microorganisms identified in these biofilms are also frequently found in extreme, desert environments and are known for their resistance also to ionizing radiation, such as Deinococcales and Actinobacteria or ascomycete fungi (Ascomycota). Among phototrophic lineages, we identified Cyanobacteria, Chlorophyta (green algae) and sometimes Streptophyta. We showed that environmental parameters influenced biofilm microbial communities. However, whereas the bacterial community composition depends on the nature of the substrate, the microbial eukaryotic community composition depends on the geographic distance. We also carried out colonization experiences exposing outdoors the same mineral substrate in three different sites in Northern Ireland and France. The analysis of microbial diversity along the colonization process revealed important changes in community composition both for prokaryotes and eukaryotes, although the behavior of the two groups was different. In the case of bacteria, we observed a transition from Gammaproteobacteria, which dominated the initial 0-6 months and which likely corresponded to inactive dispersive cells, towards Betaproteobacteria, Bacteroidetes, Alphaproteobacteria and Actinobacteria in successive steps of biofilm formation. By contrast, since their detection on mineral substrates, eukaryotes were massively dominated by ascomycete and basidiomycete fungi, green algae and other minor components such as ciliates were detected in later stages of biofilm formation. Our results show that heterotrophic organisms are pioneers in the formation of these biofilms, leading to the hypothesis that they facilitate the settlement of Cyanobacteria and, especially, of green algae. They also show that the process of bacteria community assembly depends on colonization time whereas the geographic site determines that of eukaryotic microorganisms. These major differences might be explained by different lifestyles between organisms of the two group

Diversité et processus de colonisation microbienne sur des substrats minéraux

The major objective of my PhD work was the analysis of the diversity of microorganisms from the three domains of life associated with phototrophic biofilms developing on different mineral substrates exposed outdoors. These studies aimed at answering questions about microbial diversity and biogeography and also at studying the colonization process through controlled exposure experiments. I have thus characterized, essentially by molecular methods based on small subunit (SSU) rRNA gene libraries and fingerprinting analyses the diversity of prokaryote and eukaryote microorganisms forming mature biofilms (exposed for several years) in various geographic sites in Northern Ireland, France and Ukraine, in the Chernobyl area. In these biofilms, subjected to strong selective pressure, we found many heterotrophic and phototrophic microorganisms, but their diversity was limited when compared to that of other environments such as soils or aquatic systems. Archaea were absent from all biofilms. The environmental conditions to which these biofilms are constantly exposed, such as irradiation, desiccation and nutrient limitation select for organisms that develop particular adaptive strategies including, among others, pigment production. The microorganisms identified in these biofilms are also frequently found in extreme, desert environments and are known for their resistance also to ionizing radiation, such as Deinococcales and Actinobacteria or ascomycete fungi (Ascomycota). Among phototrophic lineages, we identified Cyanobacteria, Chlorophyta (green algae) and sometimes Streptophyta. We showed that environmental parameters influenced biofilm microbial communities. However, whereas the bacterial community composition depends on the nature of the substrate, the microbial eukaryotic community composition depends on the geographic distance. We also carried out colonization experiences exposing outdoors the same mineral substrate in three different sites in Northern Ireland and France. The analysis of microbial diversity along the colonization process revealed important changes in community composition both for prokaryotes and eukaryotes, although the behavior of the two groups was different. In the case of bacteria, we observed a transition from Gammaproteobacteria, which dominated the initial 0-6 months and which likely corresponded to inactive dispersive cells, towards Betaproteobacteria, Bacteroidetes, Alphaproteobacteria and Actinobacteria in successive steps of biofilm formation. By contrast, since their detection on mineral substrates, eukaryotes were massively dominated by ascomycete and basidiomycete fungi, green algae and other minor components such as ciliates were detected in later stages of biofilm formation. Our results show that heterotrophic organisms are pioneers in the formation of these biofilms, leading to the hypothesis that they facilitate the settlement of Cyanobacteria and, especially, of green algae. They also show that the process of bacteria community assembly depends on colonization time whereas the geographic site determines that of eukaryotic microorganisms. These major differences might be explained by different lifestyles between organisms of the two groupsMes travaux de recherche ont eu pour but d’analyser la diversité des microorganismes des trois domaines du vivant présents dans des biofilms phototrophes exposés à l’air, se développant sur des substrats minéraux divers, afin d’essayer, d’une part, de répondre à des questions de diversité et de biogéographie et, d’autre part, d’étudier le processus de colonisation par le biais d’expériences d’exposition contrôlées.J’ai ainsi caractérisé, essentiellement par des approches moléculaires basées sur l'analyse des banques des gènes d'ARNr de la petite sous-unité (SSU rDNAs) et sur des analyses d'empreintes communautaires, la diversité microbienne (procaryote et eucaryote) formant des biofilms matures (exposés depuis plusieurs années) dans plusieurs sites géographiques en Irlande du Nord, en France et en Ukraine, dans la région de Chernobyl. Dans ces biofilms soumis à forte pression sélective, nous avons mis en évidence beaucoup de microorganismes hétérotrophes et phototrophes, mais avec une diversité relativement restreinte en comparaison à d’autres milieux comme les sols ou les systèmes aquatiques. Les archées étaient absentes. Les conditions environnementales auxquelles ce type de biofilm est constamment exposé comme l’irradiation, la dessiccation et la limitation des nutriments sélectionnent des microorganismes qui développent des stratégies pour s’adapter comme, entre autres, la production de pigments. Ce sont des microorganismes fréquemment retrouvés dans des milieux désertiques extrêmes et résistants aussi aux radiations ionisantes qui ont ainsi été identifiés, notamment des Deinococcales et des Actinobacteria, ou encore des champignons ascomycètes (Ascomycota). Parmi les organismes phototrophes, nous avons dénombré des Cyanobacteria, des algues vertes (Chlorophyta) et des Streptophyta. Nous avons mis en évidence que les facteurs environnementaux influencent la composition des biofilms. Toutefois, tandis que la composition de la communauté bactérienne est fortement dépendante de la nature du substrat ou elle se développe, la composition des communautés microbiennes eucaryotes dépend de la distance géographique. Nous avons également mené des expériences de colonisation en exposant un même substrat minéral dans trois sites géographiques en Irlande du Nord et en France. L'analyse de la diversité microbienne lors du processus de colonisation a révélé des changements importants dans la composition des communautés, que ce soit pour les procaryotes ou pour les eucaryotes avec, cependant, des comportements différents de ces deux groupes de microorganismes. Dans le cas des bactéries, on observe une transition des Gammaproteobacteria, qui dominent les temps 0-6 mois et qui correspondent vraisemblablement aux cellules inactives en dispersion, vers des Betaproteobacteria, Bacteroidetes, Alphaproteobacteria et Actinobacteria dans des phases successives de formation du biofilm. Par contre, dès leur détection sur le substrat minéral, les eucaryotes sont massivement dominés par des champignons ascomycètes et basidiomycètes, des algues vertes ainsi que d'autres composantes minoritaires comme des ciliés, étant détectées dans des stades plus tardifs. Nos résultats montrent que les organismes hétérotrophes sont pionniers dans la formation de ces biofilms, ce qui permet d'émettre l'hypothèse qu'ils facilitent l'installation des cyanobactéries et surtout des algues vertes. Ils montrent aussi que le processus d'assemblage des communautés bactériennes dépend du temps de colonisation, alors que le site géographique détermine celui des microorganismes eucaryotes. Ces différences majeures de comportement pourraient être expliquées par des modes de vie différents entre les organismes de ces deux grands groupes

Diversity and microbial colonization process in biofilms on mineral substractes

Mes travaux de recherche ont eu pour but d’analyser la diversité des microorganismes des trois domaines du vivant présents dans des biofilms phototrophes exposés à l’air, se développant sur des substrats minéraux divers, afin d’essayer, d’une part, de répondre à des questions de diversité et de biogéographie et, d’autre part, d’étudier le processus de colonisation par le biais d’expériences d’exposition contrôlées.J’ai ainsi caractérisé, essentiellement par des approches moléculaires basées sur l'analyse des banques des gènes d'ARNr de la petite sous-unité (SSU rDNAs) et sur des analyses d'empreintes communautaires, la diversité microbienne (procaryote et eucaryote) formant des biofilms matures (exposés depuis plusieurs années) dans plusieurs sites géographiques en Irlande du Nord, en France et en Ukraine, dans la région de Chernobyl. Dans ces biofilms soumis à forte pression sélective, nous avons mis en évidence beaucoup de microorganismes hétérotrophes et phototrophes, mais avec une diversité relativement restreinte en comparaison à d’autres milieux comme les sols ou les systèmes aquatiques. Les archées étaient absentes. Les conditions environnementales auxquelles ce type de biofilm est constamment exposé comme l’irradiation, la dessiccation et la limitation des nutriments sélectionnent des microorganismes qui développent des stratégies pour s’adapter comme, entre autres, la production de pigments. Ce sont des microorganismes fréquemment retrouvés dans des milieux désertiques extrêmes et résistants aussi aux radiations ionisantes qui ont ainsi été identifiés, notamment des Deinococcales et des Actinobacteria, ou encore des champignons ascomycètes (Ascomycota). Parmi les organismes phototrophes, nous avons dénombré des Cyanobacteria, des algues vertes (Chlorophyta) et des Streptophyta. Nous avons mis en évidence que les facteurs environnementaux influencent la composition des biofilms. Toutefois, tandis que la composition de la communauté bactérienne est fortement dépendante de la nature du substrat ou elle se développe, la composition des communautés microbiennes eucaryotes dépend de la distance géographique. Nous avons également mené des expériences de colonisation en exposant un même substrat minéral dans trois sites géographiques en Irlande du Nord et en France. L'analyse de la diversité microbienne lors du processus de colonisation a révélé des changements importants dans la composition des communautés, que ce soit pour les procaryotes ou pour les eucaryotes avec, cependant, des comportements différents de ces deux groupes de microorganismes. Dans le cas des bactéries, on observe une transition des Gammaproteobacteria, qui dominent les temps 0-6 mois et qui correspondent vraisemblablement aux cellules inactives en dispersion, vers des Betaproteobacteria, Bacteroidetes, Alphaproteobacteria et Actinobacteria dans des phases successives de formation du biofilm. Par contre, dès leur détection sur le substrat minéral, les eucaryotes sont massivement dominés par des champignons ascomycètes et basidiomycètes, des algues vertes ainsi que d'autres composantes minoritaires comme des ciliés, étant détectées dans des stades plus tardifs. Nos résultats montrent que les organismes hétérotrophes sont pionniers dans la formation de ces biofilms, ce qui permet d'émettre l'hypothèse qu'ils facilitent l'installation des cyanobactéries et surtout des algues vertes. Ils montrent aussi que le processus d'assemblage des communautés bactériennes dépend du temps de colonisation, alors que le site géographique détermine celui des microorganismes eucaryotes. Ces différences majeures de comportement pourraient être expliquées par des modes de vie différents entre les organismes de ces deux grands groupes.The major objective of my PhD work was the analysis of the diversity of microorganisms from the three domains of life associated with phototrophic biofilms developing on different mineral substrates exposed outdoors. These studies aimed at answering questions about microbial diversity and biogeography and also at studying the colonization process through controlled exposure experiments. I have thus characterized, essentially by molecular methods based on small subunit (SSU) rRNA gene libraries and fingerprinting analyses the diversity of prokaryote and eukaryote microorganisms forming mature biofilms (exposed for several years) in various geographic sites in Northern Ireland, France and Ukraine, in the Chernobyl area. In these biofilms, subjected to strong selective pressure, we found many heterotrophic and phototrophic microorganisms, but their diversity was limited when compared to that of other environments such as soils or aquatic systems. Archaea were absent from all biofilms. The environmental conditions to which these biofilms are constantly exposed, such as irradiation, desiccation and nutrient limitation select for organisms that develop particular adaptive strategies including, among others, pigment production. The microorganisms identified in these biofilms are also frequently found in extreme, desert environments and are known for their resistance also to ionizing radiation, such as Deinococcales and Actinobacteria or ascomycete fungi (Ascomycota). Among phototrophic lineages, we identified Cyanobacteria, Chlorophyta (green algae) and sometimes Streptophyta. We showed that environmental parameters influenced biofilm microbial communities. However, whereas the bacterial community composition depends on the nature of the substrate, the microbial eukaryotic community composition depends on the geographic distance. We also carried out colonization experiences exposing outdoors the same mineral substrate in three different sites in Northern Ireland and France. The analysis of microbial diversity along the colonization process revealed important changes in community composition both for prokaryotes and eukaryotes, although the behavior of the two groups was different. In the case of bacteria, we observed a transition from Gammaproteobacteria, which dominated the initial 0-6 months and which likely corresponded to inactive dispersive cells, towards Betaproteobacteria, Bacteroidetes, Alphaproteobacteria and Actinobacteria in successive steps of biofilm formation. By contrast, since their detection on mineral substrates, eukaryotes were massively dominated by ascomycete and basidiomycete fungi, green algae and other minor components such as ciliates were detected in later stages of biofilm formation. Our results show that heterotrophic organisms are pioneers in the formation of these biofilms, leading to the hypothesis that they facilitate the settlement of Cyanobacteria and, especially, of green algae. They also show that the process of bacteria community assembly depends on colonization time whereas the geographic site determines that of eukaryotic microorganisms. These major differences might be explained by different lifestyles between organisms of the two group

Diversité et processus de colonisation microbienne sur des substrats minéraux

Mes travaux de recherche ont eu pour but d analyser la diversité des microorganismes des trois domaines du vivant présents dans des biofilms phototrophes exposés à l air, se développant sur des substrats minéraux divers, afin d essayer, d une part, de répondre à des questions de diversité et de biogéographie et, d autre part, d étudier le processus de colonisation par le biais d expériences d exposition contrôlées.J ai ainsi caractérisé, essentiellement par des approches moléculaires basées sur l'analyse des banques des gènes d'ARNr de la petite sous-unité (SSU rDNAs) et sur des analyses d'empreintes communautaires, la diversité microbienne (procaryote et eucaryote) formant des biofilms matures (exposés depuis plusieurs années) dans plusieurs sites géographiques en Irlande du Nord, en France et en Ukraine, dans la région de Chernobyl. Dans ces biofilms soumis à forte pression sélective, nous avons mis en évidence beaucoup de microorganismes hétérotrophes et phototrophes, mais avec une diversité relativement restreinte en comparaison à d autres milieux comme les sols ou les systèmes aquatiques. Les archées étaient absentes. Les conditions environnementales auxquelles ce type de biofilm est constamment exposé comme l irradiation, la dessiccation et la limitation des nutriments sélectionnent des microorganismes qui développent des stratégies pour s adapter comme, entre autres, la production de pigments. Ce sont des microorganismes fréquemment retrouvés dans des milieux désertiques extrêmes et résistants aussi aux radiations ionisantes qui ont ainsi été identifiés, notamment des Deinococcales et des Actinobacteria, ou encore des champignons ascomycètes (Ascomycota). Parmi les organismes phototrophes, nous avons dénombré des Cyanobacteria, des algues vertes (Chlorophyta) et des Streptophyta. Nous avons mis en évidence que les facteurs environnementaux influencent la composition des biofilms. Toutefois, tandis que la composition de la communauté bactérienne est fortement dépendante de la nature du substrat ou elle se développe, la composition des communautés microbiennes eucaryotes dépend de la distance géographique. Nous avons également mené des expériences de colonisation en exposant un même substrat minéral dans trois sites géographiques en Irlande du Nord et en France. L'analyse de la diversité microbienne lors du processus de colonisation a révélé des changements importants dans la composition des communautés, que ce soit pour les procaryotes ou pour les eucaryotes avec, cependant, des comportements différents de ces deux groupes de microorganismes. Dans le cas des bactéries, on observe une transition des Gammaproteobacteria, qui dominent les temps 0-6 mois et qui correspondent vraisemblablement aux cellules inactives en dispersion, vers des Betaproteobacteria, Bacteroidetes, Alphaproteobacteria et Actinobacteria dans des phases successives de formation du biofilm. Par contre, dès leur détection sur le substrat minéral, les eucaryotes sont massivement dominés par des champignons ascomycètes et basidiomycètes, des algues vertes ainsi que d'autres composantes minoritaires comme des ciliés, étant détectées dans des stades plus tardifs. Nos résultats montrent que les organismes hétérotrophes sont pionniers dans la formation de ces biofilms, ce qui permet d'émettre l'hypothèse qu'ils facilitent l'installation des cyanobactéries et surtout des algues vertes. Ils montrent aussi que le processus d'assemblage des communautés bactériennes dépend du temps de colonisation, alors que le site géographique détermine celui des microorganismes eucaryotes. Ces différences majeures de comportement pourraient être expliquées par des modes de vie différents entre les organismes de ces deux grands groupes.The major objective of my PhD work was the analysis of the diversity of microorganisms from the three domains of life associated with phototrophic biofilms developing on different mineral substrates exposed outdoors. These studies aimed at answering questions about microbial diversity and biogeography and also at studying the colonization process through controlled exposure experiments. I have thus characterized, essentially by molecular methods based on small subunit (SSU) rRNA gene libraries and fingerprinting analyses the diversity of prokaryote and eukaryote microorganisms forming mature biofilms (exposed for several years) in various geographic sites in Northern Ireland, France and Ukraine, in the Chernobyl area. In these biofilms, subjected to strong selective pressure, we found many heterotrophic and phototrophic microorganisms, but their diversity was limited when compared to that of other environments such as soils or aquatic systems. Archaea were absent from all biofilms. The environmental conditions to which these biofilms are constantly exposed, such as irradiation, desiccation and nutrient limitation select for organisms that develop particular adaptive strategies including, among others, pigment production. The microorganisms identified in these biofilms are also frequently found in extreme, desert environments and are known for their resistance also to ionizing radiation, such as Deinococcales and Actinobacteria or ascomycete fungi (Ascomycota). Among phototrophic lineages, we identified Cyanobacteria, Chlorophyta (green algae) and sometimes Streptophyta. We showed that environmental parameters influenced biofilm microbial communities. However, whereas the bacterial community composition depends on the nature of the substrate, the microbial eukaryotic community composition depends on the geographic distance. We also carried out colonization experiences exposing outdoors the same mineral substrate in three different sites in Northern Ireland and France. The analysis of microbial diversity along the colonization process revealed important changes in community composition both for prokaryotes and eukaryotes, although the behavior of the two groups was different. In the case of bacteria, we observed a transition from Gammaproteobacteria, which dominated the initial 0-6 months and which likely corresponded to inactive dispersive cells, towards Betaproteobacteria, Bacteroidetes, Alphaproteobacteria and Actinobacteria in successive steps of biofilm formation. By contrast, since their detection on mineral substrates, eukaryotes were massively dominated by ascomycete and basidiomycete fungi, green algae and other minor components such as ciliates were detected in later stages of biofilm formation. Our results show that heterotrophic organisms are pioneers in the formation of these biofilms, leading to the hypothesis that they facilitate the settlement of Cyanobacteria and, especially, of green algae. They also show that the process of bacteria community assembly depends on colonization time whereas the geographic site determines that of eukaryotic microorganisms. These major differences might be explained by different lifestyles between organisms of the two groupsPARIS11-SCD-Bib. électronique (914719901) / SudocSudocFranceF

Mise en place du bilan sanitaire volontaire en élevage bovin (l'exemple de la Loire (42))

LYON1-BU Santé (693882101) / SudocTOULOUSE-EN Vétérinaire (315552301) / SudocSudocFranceF