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The MucR/Ros family protein is conserved in alpha-proteobacteria and characterized by its zinc-finger motif that has been proposed as the ancestral domain from which the eukaryotic C2H2 zinc-finger structure evolved. In the past decades, accumulated evidences have revealed MucR as a pleiotropic transcriptional regulator that integrating multiple functions such as virulence, symbiosis, cell cycle and various physiological processes. Scattered reports indicate that MucR mainly acts as a repressor, through oligomerization and binding to multiple sites of AT-rich target promoters. The N-terminal region and zinc-finger bearing C-terminal region of MucR mediate oligomerization and DNA-binding, respectively. These features are convergent to those of xenogeneic silencers such as H-NS, MvaT, Lsr2 and Rok, which are mainly found in other lineages. Phylogenetic analysis of MucR homologs suggests an ancestral origin of MucR in alpha- and delta-proteobacteria. Multiple independent duplication and lateral gene transfer events contribute to the diversity and phyletic distribution of MucR. Finally, we posed questions which remain unexplored regarding the putative roles of MucR as a xenogeneic silencer and a general manager in balancing adaptation and regulatory integration in the pangenome context.

Cyclic-di-GMP plays crucial role in the cell cycle regulation of the α-Proteobacterium Caulobacter crescentus. Here we investigated its role in the α-Proteobacterium Brucella abortus, a zoonotic intracellular pathogen. Surprisingly, deletion of all predicted cyclic-di-GMP synthesizing or degrading enzymes did not drastically impair the growth of B. abortus, nor its ability to grow inside cell lines. As other Rhizobiales, B. abortus displays unipolar growth from the new cell pole generated by cell division. We found that the phosphodiesterase PdeA, the ortholog of the essential polar growth factor RgsP of the Rhizobiale Sinorhizobium meliloti, is required for rod shape integrity but is not essential for B. abortus growth. Indeed, the radius of the pole is increased by 31 ± 1.7% in a ΔpdeA mutant, generating a coccoid morphology. A mutation in the cyclic-di-GMP phosphodiesterase catalytic site of PdeA does not generate the coccoid morphology and the ΔpdeA mutant kept the ability to recruit markers of new and old poles. However, the presence of PdeA is required in an intra-nasal mouse model of infection. In conclusion, we propose that PdeA contributes to bacterial morphology and virulence in B. abortus, but it is not crucial for polarity and asymmetric growth.

Intracellular bacteria of the genus Wolbachia (alpha-Proteobacteria) are the most widespread endosymbionts of insects. Host infection is usually associated with alterations in reproduction, such as cytoplasmic incompatibility, the induction of parthenogenesis and offspring sex ratio bias: all phenomena that may influence host speciation. In the present study, by using well-established molecular tools, we investigated the presence of Wolbachia in leaf beetles of the genus Crioceris and their host plants, which are various species of Asparagus. Multilocus sequence typing of bacterial genes showed that despite their occurrence in the same habitat and feeding on the same plant, two species of Crioceris, C. quinquepunctata and C. quatuordecimpunctata, are infected by two different strains of Wolbachia. C. asparagi, C. paracenthesis and C. duodecimpunctata, which are sympatric with the infected species, do not harbour the bacterium. Interestingly, DNA of Wolbachia was detected in host plant tissues that are exploited by the beetles, providing evidence for the horizontal transmission of the bacterium between beetles and their host plants. Moreover, Wolbachia was detected in species of Crioceris that are not closely related.

The obligate intracellular microbe, Wolbachia pipientis (Rickettsiales; Anaplasmataceae), is a Gram-negative member of the alpha proteobacteria that infects arthropods and filarial worms. Although closely related to the genera Anaplasma and Ehrlichia, which include pathogens of humans, Wolbachia is uniquely associated with invertebrate hosts in the clade Ecdysozoa. Originally described in Culex pipiens mosquitoes, Wolbachia is currently represented by 17 supergroups and is believed to occur in half of all insect species. In mosquitoes, Wolbachia acts as a gene drive agent, with the potential to modify vector populations; in filarial worms, Wolbachia functions as a symbiont, and is a target for drug therapy. A small number of Wolbachia strains from supergroups A, B, and F have been maintained in insect cell lines, which are thought to provide a more permissive environment than the natural host. When transferred back to an insect host, Wolbachia produced in cultured cells are infectious and retain reproductive phenotypes. Here, I review applications of insect cell lines in Wolbachia research and describe conditions that facilitate Wolbachia infection and replication in naive host cells. Progress in manipulation of Wolbachia in vitro will enable genetic and biochemical advances that will facilitate eventual genetic engineering of this important biological control agent.

It has become clear that sRNAs play relevant regulatory functions in bacteria. However, a comprehensive understanding of their biological roles considering evolutionary aspects has not been achieved for most of them. Thus, we have characterized the evolutionary and phylogenetic aspects of the Sinorhizobium meliloti mmgR gene encoding the small RNA MmgR, which has been recently reported to be involved in the regulation of polyhydroxybutyrate accumulation in this bacterium. We constructed a covariance model from a multiple sequence and structure alignment of mmgR close homologs that allowed us to extend the search and to detect further remote homologs of the sRNA gene. From our results, mmgR seemed to evolve from a common ancestor of the α-proteobacteria that diverged from the order of Rickettsiales. We have found mmgR homologs in most current species of α-proteobacteria, with a few exceptions in which genomic reduction events or gene rearrangements seem to explain its absence. Furthermore, a strong microsyntenic relationship was found between a large set of mmgR homologs and homologs of a gene encoding a putative N-formyl glutamate amidohydrolase (NFGAH) that allowed us to trace back the evolutionary path of this group of mmgR orthologs. Among them, structure and sequence traits have been completely conserved throughout evolution, namely a Rho-independent terminator and a 10-mer (5′-UUUCCUCCCU-3′) that is predicted to remain in a single-stranded region of the sRNA. We thus propose the definition of the new family of α-proteobacterial sRNAs αr8, as well as the subfamily αr8s1 which encompass S. meliloti mmgR orthologs physically linked with the downstream open reading frame encoding a putative NFGAH. So far, mmgR is the trans-encoded small RNA with the widest phylogenetic distribution of well recognized orthologs among α-proteobacteria. Expression of the expected MmgR transcript in rhizobiales other than S. meliloti (Sinorhizobium fredii, Rhizobium leguminosarum and Rhizobium etli) was confirmed by Northern blot. These findings will contribute to the understanding of the biological role(s) of mmgR in the α-proteobacteria. Fil: Lagares, Antonio. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Investigación en Interacciones Biológicas; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina Fil: Roux, Indra. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Investigación en Interacciones Biológicas; Argentina Fil: Valverde, Claudio Fabián. Universidad Nacional de Quilmes. Departamento de Ciencia y Tecnología. Laboratorio de Investigación en Interacciones Biológicas; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina

The α-proteobacteria are a fascinating group of free-living, symbiotic and pathogenic organisms, including the Brucella genus, which is responsible for a worldwide zoonosis. One common feature of α-proteobacteria is the presence of a conserved response regulator called CtrA, first described in the model bacterium Caulobacter crescentus, where it controls gene expression at different stages of the cell cycle. Here, we focus on Brucella abortus and other intracellular α-proteobacteria in order to better assess the potential role of CtrA in the infectious context. Comparative genomic analyses of the CtrA control pathway revealed the conservation of specific modules, as well as the acquisition of new factors during evolution. The comparison of CtrA regulons also suggests that specific clades of α-proteobacteria acquired distinct functions under its control, depending on the essentiality of the transcription factor. Other CtrA-controlled functions, for instance motility and DNA repair, are proposed to be more ancestral. Altogether, these analyses provide an interesting example of the plasticity of a regulation network, subject to the constraints of inherent imperatives such as cell division and the adaptations to diversified environmental niches.

The small RNA AbcR1 regulates the expression of ABC transporters in the plant pathogen Agrobacterium tumefaciens, the plant symbiont Sinorhizobium meliloti, and the human pathogen Brucella abortus. A combination of proteomic and bioinformatic approaches suggested dozens of AbcR1 targets in A. tumefaciens. Several of these newly discovered targets are involved in the uptake of amino acids, their derivatives, and sugars. Among the latter is the periplasmic sugar-binding protein ChvE, a component of the virulence signal transduction system. We examined 16 targets and their interaction with AbcR1 in close detail. In addition to the previously described mRNA interaction site of AbcR1 (M1), the CopraRNA program predicted a second functional module (M2) as target-binding site. Both M1 and M2 contain single-stranded anti-SD motifs. Using mutated AbcR1 variants, we systematically tested by band shift experiments, which sRNA region is responsible for mRNA binding and gene regulation. On the target site, we find that AbcR1 interacts with some mRNAs in the translation initiation region and with others far into their coding sequence. Our data show that AbcR1 is a versatile master regulator of nutrient uptake systems in A. tumefaciens and related bacteria.

Wolbachia (Hertig) endosymbionts are extensively studied in a wide range of organisms and are known to be transmitted through the egg cytoplasm to the offsping. Wolbachia may cause several types of reproductive modifications in arthropods. In Trichogramma species, parthenogenesis-inducing Wolbachia bacteria allow females wasps to produce daughters from unfertilized eggs and these bacteria are present in at least 9% of all Trichogramma species. Phylogenetic studies have led to the subdivision of the Wolbachia clade in five supergroups (A, B, C, D and E) and Wolbachia from Trichogramma belong to supergroup B. Here, using the wsp gene, four groups of Wolbachia that infect Trichogramma species were distinguished and the addition of a new group “Ato” was suggested due to the addition of Wolbachia from Trichogramma atopovirilia (Oatman and Platner). Specific primers were designed and tested for the “Ato” group. Seventy-five percent of all evaluated Wolbachia strains from Trichogramma fell within “Sib” group. Resumo Endosimbiontes do gênero Wolbachia (Hertig) são extensivamente estudados em uma ampla gama de organismos e são conhecidos por serem transmitidos via citoplasma do ovo hospedeiro para seu descendente. Wolbachia pode causar vários tipos de alterações reprodutivas nos artrópodes. Nas espécies de Trichogramma, a reprodução partenogenética induzida por Wolbachia, possibilita as fêmeas dos parasitoides a produção de fêmeas a partir de ovos não fertilizados e estas bactérias estão presentes em pelo menos 9% de todas as espécies de Trichogramma. Estudos filogenéticos têm levado a subdivisão do clado Wolbachia em cinco supergrupos (A, B, C, D and E). Wolbachia em Trichogramma pertence ao supergrupo B. Com o gene wsp foi possível se distinguir quatro grupos de Wolbachia que infectam Trichogramma e adicionar um novo grupo (Ato) devido a inclusão de Wolbachia detectada em Trichogramma atopovirilia (Oatman and Platner, 1983). Primers específicos foram construídos e testados para o grupo “Ato”. Setenta e cinco por cento de todas as linhagens de Wolbachia que infectam Trichogramma se enquadraram dentro do grupo “Sib”.

Bacterial endosymbionts gave rise to mitochondria in a process that depended on the acquisition of protein import pathways. Modification and in some cases major re-tooling of the endosymbiont's cellular machinery produced these pathways, establishing mitochondria as organelles common to all eukaryotic cells. The legacy of this evolutionary tinkering can be seen in the homologies and structural similarities between mitochondrial protein import machinery and modern day bacterial proteins. Comparative analysis of these systems is revealing both possible routes for the evolution of the mitochondrial membrane translocases and a greater understanding of the mechanisms behind mitochondrial protein import. This article is part of a Special Issue entitled Protein translocation across or insertion into membranes.

International audience; Bacterial accessory genes are genomic symbionts with an evolutionary history and future that is different from that of their hosts. Packages of accessory genes move from strain to strain and confer important adaptations, such as interaction with eukaryotes. The ability to fix nitrogen with legumes is a remarkable example of a complex trait spread by horizontal transfer of a few key symbiotic genes, converting soil bacteria into legume symbionts. Rhizobia belong to hundreds of species restricted to a dozen genera of the Alphaproteobacteria and Betaproteobacteria, suggesting infrequent successful transfer between genera but frequent successful transfer within genera. Here we review the genetic and environmental conditions and selective forces that have shaped evolution of this complex symbiotic trait.

The RNA chaperone Hfq is a global post-transcriptional regulator in bacteria. Here, we used RNAseq to analyze RNA populations from the legume symbiont Sinorhizobium meliloti that were co-immunoprecipitated (CoIP-RNA) with a FLAG-tagged Hfq in five growth/stress conditions. Hfq-bound transcripts (1315) were largely identified in stressed bacteria and derived from small RNAs (sRNAs), both trans-encoded (6.4%) and antisense (asRNAs; 6.3%), and mRNAs (86%). Pull-down with Hfq recovered a small proportion of annotated S. meliloti sRNAs (14% of trans-sRNAs and 2% of asRNAs) suggesting a discrete impact of this protein in sRNA pathways. Nonetheless, Hfq selectively stabilized CoIP-enriched sRNAs, anticipating that these interactions are functionally significant. Transcription of 26 Hfq-bound sRNAs was predicted to occur from promoters recognized by the major stress s factors sigma(E2) or sigma(H1/2). Recovery rates of sRNAs in each of the CoIP-RNA libraries suggest a large impact of Hfq-assisted riboregulation in S. meliloti osmoadaptation. Hfq directly targeted 18% of the predicted S. meliloti mRNAs, which encode functionally diverse proteins involved in transport and metabolism, sigma(E2)-dependent stress responses, quorum sensing, flagella biosynthesis, ribosome, and membrane assembly or symbiotic nitrogen fixation. Canonical targeting of the 5' regions of two of the ABC transporter mRNAs by the homologous Hfq-binding AbcR1 and AbcR2 sRNAs leading to inhibition of protein synthesis was confirmed in vivo. We therefore provide a comprehensive resource for the systems-level deciphering of hitherto unexplored S. meliloti stress and symbiotic post-transcriptional regulons and the identification of Hfq-dependent sRNA-mRNA regulatory pairs.

The symbiotic alpha-rhizobia Sinorhizobium meliloti, Bradyrhizobium japonicum, Rhizobium etli and the related plant pathogen Agrobacterium tumefaciens are important model organisms for studying plant-microbe interactions. These metabolically versatile soil bacteria are characterized by complex lifestyles and large genomes. Here we summarize the recent knowledge on their small non-coding RNAs (sRNAs) including conservation, function, and interaction of the sRNAs with the RNA chaperone Hfq. In each of these organisms, an inventory of hundreds of cis- and trans-encoded sRNAs with regulatory potential was uncovered by high-throughput approaches and used for the construction of 39 sRNA family models. Genome-wide analyses of hfq mutants and co-immunoprecipitation with tagged Hfq revealed a major impact of the RNA chaperone on the physiology of plant-associated alpha-proteobacteria including symbiosis and virulence. Highly conserved members of the SmelC411 family are the AbcR sRNAs, which predominantly regulate ABC transport systems. AbcR1 of A. tumefaciens controls the uptake of the plant-generated signaling molecule GABA and is a central regulator of nutrient uptake systems. It has similar functions in S. meliloti and the human pathogen Brucella abortus. As RNA degradation is an important process in RNA-based gene regulation, a short overview on ribonucleases in plant-associated alpha-proteobacteria concludes this review.

Background: Specialized interactions help structure communities, but persistence of specialized organisms is puzzling because a generalist can occupy more environments and partake in more beneficial interactions. The " Jack-of-all-trades is a master of none" hypothesis asserts that specialists persist because the fitness of a generalist utilizing a particular habitat is lower than that of a specialist adapted to that habitat. Yet, there are many reasons to expect that mutualists will generalize on partners.Plant-soil feedbacks help to structure plant and microbial communities, but how frequently are soil-based symbiotic mutualistic interactions sufficiently specialized to influence species distributions and community composition? To address this question, we quantified realized partner richness and phylogenetic breadth of four wild-grown native legumes (Lupinus bicolor, L. arboreus, Acmispon strigosus and A. heermannii) and performed inoculation trials to test the ability of two hosts (L. bicolor and A. strigosus) to nodulate (fundamental partner richness), benefit from (response specificity), and provide benefit to (effect specificity) 31 Bradyrhizobium genotypes.Results: In the wild, each Lupinus species hosted a broader genetic range of Bradyrhizobium than did either Acmispon species, suggesting that Acmispon species are more specialized. In the greenhouse, however, L. bicolor and A. strigosus did not differ in fundamental association specificity: all inoculated genotypes nodulated both hosts. Nevertheless, A. strigosus exhibited more specificity, i.e., greater variation in its response to, and effect on, Bradyrhizobium genotypes. Lupinus bicolor benefited from a broader range of genotypes but averaged less benefit from each. Both hosts obtained more fitness benefit from symbionts isolated from conspecific hosts; those symbionts in turn gained greater fitness benefit from hosts of the same species from which they were isolated.Conclusions: This study affirmed two important tenets of evolutionary theory. First, as predicted by the Jack-of-all-trades is a master of none hypothesis, specialist A. strigosus obtained greater benefit from its beneficial symbionts than did generalist L. bicolor. Second, as predicted by coevolutionary theory, each test species performed better with partner genotypes isolated from conspecifics. Finally, positive fitness feedback between the tested hosts and symbionts suggests that positive plant-soil feedback could contribute to their patchy distributions in this system. © 2014 Ehinger et al.; licensee BioMed Central Ltd.

A core component of the α-proteobacterial general stress response (GSR) is the extracytoplasmic function (ECF) sigma factor EcfG, exclusively present in this taxonomic class. Half of the completed α-proteobacterial genome sequences contain two or more copies of genes encoding σEcfG-like sigma factors, with the primary copy typically located adjacent to genes coding for a cognate anti-sigma factor (NepR) and two-component response regulator (PhyR). So far, the widespread occurrence of additional, non-canonical σEcfG copies has not satisfactorily been explained. This study explores the hierarchical relation between Rhizobium etli σEcfG1 and σEcfG2, canonical and non-canonical σEcfG proteins, respectively. Contrary to reports in other species, we find that σEcfG1 and σEcfG2 act in parallel, as nodes of a complex regulatory network, rather than in series, as elements of a linear regulatory cascade. We demonstrate that both sigma factors control unique yet also shared target genes, corroborating phenotypic evidence. σEcfG1 drives expression of rpoH2, explaining the increased heat sensitivity of an ecfG1 mutant, while katG is under control of σEcfG2, accounting for reduced oxidative stress resistance of an ecfG2 mutant. We also identify non-coding RNA genes as novel σEcfG targets. We propose a modified model for GSR regulation in R. etli, in which σEcfG1 and σEcfG2 function largely independently. Based on a phylogenetic analysis and considering the prevalence of α-proteobacterial genomes with multiple σEcfG copies, this model may also be applicable to numerous other species. This research has been funded by the Interuniversity Attraction Poles Programme initiated by the Belgian Science Policy Office and was supported by grants from the Research Council of the KU Leuven (GOA/011/2008) and from the Fund for Scientific Research-Flanders (G.0412.10).

Unisexual Trichogramma forms have attracted much attention due to their potential advantages as biocontrol agents. Fitness studies have been performed and understanding the cost that Wolbachia may inflict on their hosts will help in deciding if Wolbachia infected (unisexual) forms are indeed better than sexual forms when used in biological control programmes. The influence of Wolbachia on the foraging behaviour (including walking activity and speed) of T. atopovirilia is reported in this paper. Temperature strongly affected T. atopovirilia female walking activity, but Wolbachia infected and uninfected females differed in none of the behavioural components that were measured such as walking activity and walking speed. Walking activity was highest at 25 ºC and differed significantly from that at 20 and 15 ºC. Trichogramma wasps were highly affected at 15 ºC. Behaviour analysis with females showed that female wasps spend most of the time on drilling + ovipositing on host eggs followed by host drumming and walking while drumming. The parasitism rate and number of offspring did not differ significantly between infected and cured Trichogramma females. Biological control implications of these findings are discussed. Formas unissexuais de Trichogramma têm despertado a atenção de pesquisadores devido as potenciais vantagens deste parasitóide como agente de controle biológico. O estudo do "Fitness" tem sido avaliado e entender o custo de ser infectado por Wolbachia ajudará em determinar se formas infectadas por Wolbachia (unisexuais) são realmente melhores que as sexuadas quando utilizadas em programas de controle biológico. A influência de Wolbachia no comportamento de procura (incluindo a atividade e a velocidade de caminhamento) de T. atopovirilia é relatada neste artigo. A temperatura afetou grandemente a atividade de caminhamento de fêmeas de T. atopovirilia, entretanto os componentes de comportamento tais como a atividade e a velocidade de caminhamento não diferiram em relação às fêmeas infectadas e não infectadas. A atividade de caminhamento foi maior a 25 ºC e diferiu significativamente de 20 e 15 ºC. Fêmeas de Trichogramma foram altamente afetados a 15 ºC. Em função da análise do comportamento com fêmeas, pode-se verificar que houve maior tempo gasto no processo de oviposição, seguido pelo toque com as antenas sobre os ovos dos hospedeiros e do caminhamento ao mesmo tempo que toca os ovos com as antenas. A taxa de parasitismo e o número de descendentes não diferiram significativamente entre fêmeas de Trichogramma infectadas e curadas da infecção por Wolbachia. As implicações dos resultados obtidos sobre o controle biológico são discutidas.

Sinorhizobium meliloti is a symbiotic soil bacterium of the alphaproteobacterial subdivision. Like other rhizobia, S. meliloti induces nitrogen-fixing root nodules on leguminous plants. This is an ecologically and economically important interaction, because plants engaged in symbiosis with rhizobia can grow without exogenous nitrogen fertilizers. The S. meliloti–Medicago truncatula (barrel medic) association is an important symbiosis model. The S. meliloti genome was published in 2001, and the M. truncatula genome currently is being sequenced. Many new resources and data have been made available since the original S. meliloti genome annotation and an update was needed. In June 2008, we submitted our annotation update to the EMBL and NCBI databases. Here we describe this new annotation and a new webbased portal RhizoGATE. About 1000 annotation updates were made; these included assigning functions to 313 putative proteins, assigning EC numbers to 431 proteins, and identifying 86 new putative genes. RhizoGATE incorporates the new annotion with the S. meliloti GenDB project, a platform that allows annotation updates in real time. Locations of transposon insertions, plasmid integrations, and array probe sequences are available in the GenDB project. RhizoGATE employs the EMMA platform for management and analysis of transcriptome data and the IGetDB data warehouse to integrate a variety of heterogeneous external data sources. © 2008 Elsevier B.V. All rights reserved.

We used comparative genomics to investigate the distribution of conserved DNA-binding motifs in the regulatory regions of genes involved in iron and manganese homeostasis in alpha-proteobacteria. Combined with other computational approaches, this allowed us to reconstruct the metal regulatory network in more than three dozen species with available genome sequences. We identified several classes of cis-acting regulatory DNA motifs (Irr-boxes or ICEs, RirA-boxes, Iron-Rhodo-boxes, Fur-alpha-boxes, Mur-box or MRS, MntR-box, and IscR-boxes) in regulatory regions of various genes involved in iron and manganese uptake, Fe-S and heme biosynthesis, iron storage, and usage. Despite the different nature of the iron regulons in selected lineages of alpha-proteobacteria, the overall regulatory network is consistent with, and confirmed by, many experimental observations. This study expands the range of genes involved in iron homeostasis and demonstrates considerable interconnection between iron-responsive regulatory systems. The detailed comparative and phylogenetic analyses of the regulatory systems allowed us to propose a theory about the possible evolution of Fe and Mn regulons in alpha-proteobacteria. The main evolutionary event likely occurred in the common ancestor of the Rhizobiales and Rhodobacterales, where the Fur protein switched to regulating manganese transporters (and hence Fur had become Mur). In these lineages, the role of global iron homeostasis was taken by RirA and Irr, two transcriptional regulators that act by sensing the physiological consequence of the metal availability rather than its concentration per se, and thus provide for more flexible regulation., Synopsis The availability of hundreds of complete genomes allows one to use comparative genomics to describe key metabolic processes and regulatory gene networks. Genome context analyses and comparisons of transcription factor binding sites between genomes offer a powerful approach for functional gene annotation. Reconstruction of transcriptional regulatory networks allows for better understanding of cellular processes, which can be substantiated by direct experimentation. Iron homeostasis in bacteria is conferred by the regulation of various iron uptake transporters, iron storage ferritins, and iron-containing enzymes. In high concentrations, iron is poisonous for the cell, so strict control of iron homeostasis is maintained, mostly at the level of transcription by iron-responsive regulators. Despite their general importance, iron regulatory networks in most bacterial species are not well-understood. In this study, Rodionov and colleagues applied comparative genomic approaches to describe the regulatory network formed by genes involved in iron homeostasis in the alpha subclass of proteobacteria, which have extremely versatile lifestyles. These networks are mediated by a set of various DNA motifs (or regulatory signals) that occur in 5′ gene regions and involve at least six different metal-responsive regulators. This study once again shows the power of comparative genomics in the analysis of complex regulatory networks and their evolution.

Bacterial genomes provide records of the molecular processes associated with emergence and evolution of different bacterial lifestyles. This thesis is based on whole-genome comparisons within the genus Bartonella, an excellent model system for studies of host- and vector-specificity and infection outcome in animal-associated bacteria. The louse-borne human specialist and trench fever agent Bartonella quintana was contrasted to the flea-borne generalist relatives Bartonella henselae and Bartonella grahamii, which cause asymptomatic infection in cat and mouse respectively. While B. henselae is commonly isolated from humans, and causes cat scratch disease, there is only one reported case of B. grahamii human infection. The gene complements of the three species are nested like Russian dolls with the smaller genome (B. quintana) being entirely contained in the medium sized (B. henselae), which in turned is contained in the largest (B. grahamii). Size differences reflect differences in the horizontally and vertically acquired gene content, and in the number of genus- and species- specific genes, owing to differential impact of bacteriophages and plasmids, and to different degrees of genome decay. These processes can be attributed to the three distinct lifestyles. Comparisons with other alpha-proteobacteria suggest that the Bartonella genus as a whole evolved from plant-associated species, and that horizontal transfer, in particular of genes involved in interaction with the host, played a key role in the transition to animal intracellular lifestyle. The long-term genome decay associated with this lifestyle is most advanced in the host-restricted B. quintana. The broad host-range species B. grahamii has the largest genome and the largest proportion of auxiliary DNA of the three, probably because it has access to a larger gene pool. In encodes all the known pathogenicity determinants found in the genomes of B. henselae and B. quintana, suggesting that these genes primarily evolved to facilitate colonization in the reservoir host.

Bacterial genomes provide records of the molecular processes associated with emergence and evolution of different bacterial lifestyles. This thesis is based on whole-genome comparisons within the genus Bartonella, an excellent model system for studies of host- and vector-specificity and infection outcome in animal-associated bacteria. The louse-borne human specialist and trench fever agent Bartonella quintana was contrasted to the flea-borne generalist relatives Bartonella henselae and Bartonella grahamii, which cause asymptomatic infection in cat and mouse respectively. While B. henselae is commonly isolated from humans, and causes cat scratch disease, there is only one reported case of B. grahamii human infection. The gene complements of the three species are nested like Russian dolls with the smaller genome (B. quintana) being entirely contained in the medium sized (B. henselae), which in turned is contained in the largest (B. grahamii). Size differences reflect differences in the horizontally and vertically acquired gene content, and in the number of genus- and species- specific genes, owing to differential impact of bacteriophages and plasmids, and to different degrees of genome decay. These processes can be attributed to the three distinct lifestyles. Comparisons with other alpha-proteobacteria suggest that the Bartonella genus as a whole evolved from plant-associated species, and that horizontal transfer, in particular of genes involved in interaction with the host, played a key role in the transition to animal intracellular lifestyle. The long-term genome decay associated with this lifestyle is most advanced in the host-restricted B. quintana. The broad host-range species B. grahamii has the largest genome and the largest proportion of auxiliary DNA of the three, probably because it has access to a larger gene pool. In encodes all the known pathogenicity determinants found in the genomes of B. henselae and B. quintana, suggesting that these genes primarily evolved to facilitate colonization in the reservoir host.

A symbiotic relationship between the marine ciliate Euplotes magnicirratus and an alpha-proteobacterium has recently been described. The bacterial symbiont, identified by molecular analysis as a new species of the genus Devosia, is referred to with the provisional name Candidatus Devosia euplotis'. This association is constant (it was found in different ciliate strains sampled in different geographical regions), and durable (it persisted in laboratory conditions for years). The physiological significance of this association was investigated through antibiotically produced aposymbiotic E. magnicirratus and attempts to grow the endosymbionts outside their host. Aposymbiotic protozoa completely lost the ability to digest the green alga Dunaliella tertiolecta, their usual food in laboratory conditions, and died. An investigation into the different steps of the digestive process showed that acidification of the digestive vacuoles occurred normally in both symbiotic and aposymbiotic ciliates, but that subsequent activation of acid phosphatase did not occur in aposymbiotic cells. Therefore, the ciliate depends on the symbiont for digestion of its green algal food and, as a consequence, for its survival. Attempts to grow bacterial endosymbionts on diluted culture media and on ciliate homogenate were unsuccessful. The results indicate that this represents a new type of symbiosis between prokaryotes and ciliates, which is mutualistic, and most likely obligatory for both partners.

Item does not contain fulltext Mitochondria are eukaryotic organelles that originated from a single bacterial endosymbiosis some 2 billion years ago. The transition from the ancestral endosymbiont to the modern mitochondrion has been accompanied by major changes in its protein content, the so-called proteome. These changes included complete loss of some bacterial pathways, amelioration of others and gain of completely new complexes of eukaryotic origin such as the ATP/ADP translocase and most of the mitochondrial protein import machinery. This renewal of proteins has been so extensive that only 14-16% of modern mitochondrial proteome has an origin that can be traced back to the bacterial endosymbiont. The rest consists of proteins of diverse origin that were eventually recruited to function in the organelle. This shaping of the proteome content reflects the transformation of mitochondria into a highly specialized organelle that, besides ATP production, comprises a variety of functions within the eukaryotic metabolism. Here we review recent advances in the fields of comparative genomics and proteomics that are throwing light on the origin and evolution of the mitochondrial proteome.