A major 6 Mb superlocus is involved in pyrethroid resistance in the common bed bug Cimex lectularius.

In the last few years, the bed bug Cimex lectularius has been an increasing problem worldwide, mainly due to the development of insecticide resistance to pyrethroids. The characterization of resistance alleles is a prerequisite to improve surveillance and resistance management. To identify genomic variants associated with pyrethroid resistance in Cimex lectularius, we compared the genetic composition of two recent and resistant populations with that of two ancient-susceptible strains using a genome-wide pool-seq design. We identified a large 6 Mb "superlocus" showing particularly high genetic differentiation and association with the resistance phenotype. This superlocus contained several clustered resistance genes and was also characterized by a high density of structural variants (inversions, duplications). The possibility that this superlocus constitutes a resistance "supergene" that evolved after the clustering of alleles adapted to insecticide and after reduction in recombination is discussed.

Artifical light at night triggers slight transcriptomic effects on melatonin signaling but not synthesis in tadpoles of two anuran species.

The worldwide expansion of artificial light at night (ALAN) is acknowledged as a threat to biodiversity through alterations of the natural photoperiod triggering the disruption of physiological functions. In vertebrates, melatonin production during the dark phase can be decreased or suppressed by nocturnal light as shown in many taxa. But the effect of ALAN at low intensity mimicking light pollution in peri-urban area has never been investigated in amphibians. We filled this gap by studying the impact of low ALAN levels on the expression of genes related to melatonin synthesis and signaling in two anurans (agile frog, Rana dalmatina, and common toad, Bufo bufo). Circadian expression of genes encoding enzymes catalyzing melatonin synthesis (aralkylamine N-acetyltransferase, AANAT and acetylserotonin O-methyltransferase, ASMT) or melatonin receptors (Mel1a, Mel1b and Mel1c) was investigated using RT-qPCR after 23 days of nocturnal exposure to control (< 0.01 lx) or low ALAN (3 lx). We showed that the relative abundance of most transcripts was low in late afternoon and early evening (06 pm and 08 pm) and increased throughout the night in R. dalmatina. However, a clear and ample nocturnal pattern of target gene expression was not detected in control tadpoles of both species. Surprisingly, a low ALAN level had little influence on the relative expression of most melatonin-related genes. Only Mel1c expression in R. dalmatina and Mel1b expression in B. bufo were affected by ALAN. This target gene approach provides experimental evidence that melatonin signaling pathway was slightly affected by low ALAN level in anuran tadpoles.

Population genomics of Corsican wildcats: Paving the way toward a new subspecies within the Felis silvestris spp. complex?

In the context of the current extinction crisis, identifying new conservation units is pivotal to the development of sound conservation measures, especially in highly threatened taxa such as felids. Corsican wildcats are known by Corsican people since a very long time but have been little studied. Meaningful information about their phylogenetic position is lacking. We used ddRADseq to genotype phenotypically homogenous Corsican wildcats at 3671 genome-wide SNPs and reported for the first time their genetic identity. We compared this genomic information to domestic cats Felis silvestris catus from Corsica and mainland France, European wildcats F. s. silvestris and Sardinian wildcats F. s. lybica. Our premise was that if the Corsican wildcat, as a phenotypic entity, also represents a genetic entity, it deserves conservation measures and to be recognized as a conservation unit. Corsican wildcats appeared highly genetically differentiated from European wildcats and genetically closer to Sardinian wildcats than to domestic cats. Domestic cats from Corsica and mainland France were closer to each other and Sardinian wildcats were intermediate between Corsican wildcats and domestic cats. This suggested that Corsican wildcats do not belong to the F. s. silvestris or catus lineages. The inclusion of more high-quality Sardinian samples and Near-Eastern mainland F. s. lybica would constitute the next step toward assessing the status of Corsican wildcat as a subspecies and/or evolutionarily significant unit and tracing back wildcat introduction history of in Corsica.

Cytotype Affects the Capability of the Whitefly Bemisia tabaci MED Species To Feed and Oviposit on an Unfavorable Host Plant.

The acquisition of nutritional obligate primary endosymbionts (P-symbionts) allowed phloemo-phageous insects to feed on plant sap and thus colonize novel ecological niches. P-symbionts often coexist with facultative secondary endosymbionts (S-symbionts), which may also influence their hosts' niche utilization ability. The whitefly Bemisia tabaci is a highly diversified species complex harboring, in addition to the P-symbiont "Candidatus Portiera aleyrodidarum," seven S-symbionts whose roles remain poorly understood. Here, we compare the phenotypic and metabolic responses of three B. tabaci lines differing in their S-symbiont community, reared on three different host plants, hibiscus, tobacco, or lantana, and address whether and how S-symbionts influence insect capacity to feed and produce offspring on those plants. We first show that hibiscus, tobacco, and lantana differ in their free amino acid composition. Insects' performance, as well as free amino acid profile and symbiotic load, were shown to be plant dependent, suggesting a critical role for the plant nutritional properties. Insect fecundity was significantly lower on lantana, indicating that it is the least favorable plant. Remarkably, insects reared on this plant show a specific amino acid profile and a higher symbiont density compared to the two other plants. In addition, this plant was the only one for which fecundity differences were observed between lines. Using genetically homogeneous hybrids, we demonstrate that cytotype (mitochondria and symbionts), and not genotype, is a major determinant of females' fecundity and amino acid profile on lantana. As cytotypes differ in their S-symbiont community, we propose that these symbionts may mediate their hosts' suitable plant range. IMPORTANCE Microbial symbionts are universal in eukaryotes, and it is now recognized that symbiotic associations represent major evolutionary driving forces. However, the extent to which symbionts contribute to their hosts' ecological adaptation and subsequent diversification is far from being fully elucidated. The whitefly Bemisia tabaci is a sap feeder associated with multiple coinfecting intracellular facultative symbionts. Here, we show that plant species simultaneously affect whiteflies' performance, amino acid profile, and symbiotic density, which could be partially explained by differences in plant nutritional properties. We also demonstrate that, on lantana, the least favorable plant used in our study, whiteflies' performance is determined by their cytotype. We propose that the host plant utilization in B. tabaci is influenced by its facultative symbiont community composition, possibly through its impact on the host dietary requirements. Altogether, our data provide new insights into the impact of intracellular microorganisms on their animal hosts' ecological niche range and diversification.

How consistent is RAD-seq divergence with DNA-barcode based clustering in insects?

Promoted by the barcoding approach, mitochondrial DNA is more than ever used as a molecular marker to identify species boundaries. Yet, it has been repeatedly argued that it may be poorly suited for this purpose, especially in insects where mitochondria are often associated with invasive intracellular bacteria that may promote their introgression. Here, we inform this debate by assessing how divergent nuclear genomes can be when mitochondrial barcodes indicate very high proximity. To this end, we obtained RAD-seq data from 92 barcode-based species-like units (operational taxonomic units [OTUs]) spanning four insect orders. In 100% of the cases, the observed median nuclear divergence was lower than 2%, a value that was recently estimated as one below which nuclear gene flow is not uncommon. These results suggest that although mitochondria may occasionally leak between species, this process is rare enough in insects to make DNA barcoding a reliable tool for clustering specimens into species-like units.

An efficient RNA-seq-based segregation analysis identifies the sex chromosomes of Cannabis sativa.

Cannabis sativa-derived tetrahydrocannabinol (THC) production is increasing very fast worldwide. C. sativa is a dioecious plant with XY Chromosomes, and only females (XX) are useful for THC production. Identifying the sex chromosome sequence would improve early sexing and better management of this crop; however, the C. sativa genome projects have failed to do so. Moreover, as dioecy in the Cannabaceae family is ancestral, C. sativa sex chromosomes are potentially old and thus very interesting to study, as little is known about old plant sex chromosomes. Here, we RNA-sequenced a C. sativa family (two parents and 10 male and female offspring, 576 million reads) and performed a segregation analysis for all C. sativa genes using the probabilistic method SEX-DETector. We identified >500 sex-linked genes. Mapping of these sex-linked genes to a C. sativa genome assembly identified the largest chromosome pair being the sex chromosomes. We found that the X-specific region (not recombining between X and Y) is large compared to other plant systems. Further analysis of the sex-linked genes revealed that C. sativa has a strongly degenerated Y Chromosome and may represent the oldest plant sex chromosome system documented so far. Our study revealed that old plant sex chromosomes can have large, highly divergent nonrecombining regions, yet still be roughly homomorphic.

Does host socio-spatial behavior lead to a fine-scale spatial genetic structure in its associated parasites?

Gastro-intestinal nematodes, especially Haemonchus contortus, are widespread pathogenic parasites of small ruminants. Studying their spatial genetic structure is as important as studying host genetic structure to fully understand host-parasite interactions and transmission patterns. For parasites having a simple life cycle (e.g., monoxenous parasites), gene flow and spatial genetic structure are expected to strongly rely on the socio-spatial behavior of their hosts. Based on five microsatellite loci, we tested this hypothesis for H. contortus sampled in a wild Mediterranean mouflon population (Ovis gmelini musimon × Ovis sp.) in which species- and environment-related characteristics have been found to generate socio-spatial units. We nevertheless found that their parasites had no spatial genetic structure, suggesting that mouflon behavior was not enough to limit parasite dispersal in this study area and/or that other ecological and biological factors were involved in this process, for example other hosts, the parasite life cycle, or the study area history.

TITLE: Le comportement socio-spatial de l'hôte conduit-il à une structure génétique à fine échelle de ses parasites ? ABSTRACT: Les nématodes gastro-intestinaux, et plus particulièrement Haemonchus contortus, sont cosmopolites et pathogènes chez les petits ruminants. Étudier leur structure génétique spatiale est aussi important que d'étudier celle des hôtes pour pleinement comprendre les interactions hôtes-parasites et les processus de transmission. Pour les parasites ayant des cycles de vie simples (par exemple, les parasites monoxènes), on s'attend à ce que les flux de gènes et la structure génétique spatiale dépendent fortement du comportement socio-spatial de leurs hôtes. En utilisant cinq loci microsatellites, nous avons testé cette hypothèse pour des H. contortus échantillonnés dans une population sauvage de mouflons méditerranéens (Ovis gmelini musimon × Ovis sp.) dans laquelle les caractéristiques de l'espèce et de l'environnement génèrent des unités socio-spatiales. Nous avons néanmoins mis en évidence que leurs parasites ne présentent pas de structure génétique spatiale, ce qui suggère que le comportement des mouflons ne restreint pas la dispersion des parasites dans cette aire d'étude et/ou que d'autres facteurs biologiques et écologiques tels que d'autres hôtes, le cycle de vie du parasite, ou l'histoire de l'aire d'étude jouent un rôle dans ce processus.

Phenotypic plasticity in the invasive pest Drosophila suzukii: activity rhythms and gene expression in response to temperature.

Phenotypic plasticity may contribute to the invasive success of an alien species in a new environment. A highly plastic species may survive and reproduce in more diverse environments, thereby supporting establishment and colonization. We focused on plasticity in the circadian rhythm of activity, which can favour species coexistence in invasion, for the invasive species Drosophila suzukii, which is expected to be a weaker direct competitor than other Drosophila species of the resident community. We compared the circadian rhythms of the locomotor activity in adults and the expression of clock genes in response to temperature in the invasive D. suzukii and the resident Drosophila melanogaster. We showed that D. suzukii is active in a narrower range of temperatures than D. melanogaster and that the activities of the two species overlap during the day, regardless of the temperature. Both species are diurnal and exhibit rhythmic activity at dawn and dusk, with a much lower activity at dawn for D. suzukii females. Our results show that the timeless and clock genes are good candidates to explain the plastic response that is observed in relation to temperature. Overall, our results suggest that thermal phenotypic plasticity in D. suzukii activity is not sufficient to explain the invasive success of D. suzukii and call for testing other hypotheses, such as the release of competitors and/or predators.

Development of a PCR-RFLP assay to identify Drosophila melanogaster among field-collected larvae.

The fruit fly Drosophila melanogaster is a model organism to study several aspects of metazoan biology. Most of the work has been conducted in adult fruit flies, including laboratory and field-derived specimens, but Drosophila melanogaster larvae recently became a valuable model to better understand animal physiology, development, or host-microbe interactions. While adult flies can be easily assigned to a given Drosophila species based on morphological characteristics, such visual identification is more intricate at the larval stage. This could explain the limited number of studies focusing on larvae, especially field-derived samples. Here, we developed a polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) assay that discriminates D. melanogaster from other ecologically relevant Drosophila species at the larval stage. The method, which targets the cytochrome oxidase I (COI) gene, was validated using laboratory-derived larvae from seven D. melanogaster populations originating from different geographic areas as well as six Drosophila species. We further validated this PCR-RFLP assay in a natural context, by identifying wild larvae collected in two locations in France. Notably, among all PCR-RFLP profiles that matched the D. melanogaster species, 100% were correctly identified, as confirmed by COI sequencing. In summary, our work provides a rapid, simple, and accurate molecular tool to identify D. melanogaster from field-collected larvae.

Abundance of Bemisia tabaci Gennadius (Hemiptera: Aleyrodidae) and its parasitoids on vegetables and cassava plants in Burkina Faso (West Africa).

The whitefly Bemisia tabaci is a pest of many agricultural and ornamental crops worldwide and particularly in Africa. It is a complex of cryptic species, which is extremely polyphagous with hundreds of host plants identified around the world. Previous surveys in western Africa indicated the presence of two biotypes of the invasive MED species (MED-Q1 and MED-Q3) living in sympatry with the African species SSA and ASL. This situation constitutes one of the rare cases of local coexistence of various genetic entities within the B. tabaci complex. In order to study the dynamics of the distribution and abundance of genetic entities within this community and to identify potential factors that could contribute to coexistence, we sampled B. tabaci populations in Burkina Faso in 2015 and 2016 on various plants, and also their parasitoids. All four genetic entities were still recorded, indicating no exclusion of local species by the MED species. While B. tabaci individuals were found on 55 plant species belonging to eighteen (18) families showing the high polyphagy of this pest, some species/biotypes exhibited higher specificity. Two parasitoid species (Eretmocerus mundus and Encarsia vandrieschei) were also recorded with E. mundus being predominant in most localities and on most plants. Our data indicated that whitefly abundance, diversity, and rate of parasitism varied according to areas, plants, and years, but that parasitism rate was globally highly correlated with whitefly abundance suggesting density dependence. Our results also suggest dynamic variation in the local diversity of B. tabaci species/biotypes from 1 year to the other, specifically with MED-Q1 and ASL species. This work provides relevant information on the nature of plant-B. tabaci-parasitoid interactions in West Africa and identifies that coexistence might be stabilized by niche differentiation for some genetic entities. However, MED-Q1 and ASL show extensive niche overlap, which could ultimately lead to competitive exclusion.

High-throughput sequencing of transposable element insertions suggests adaptive evolution of the invasive Asian tiger mosquito towards temperate environments.

Invasive species represent unique opportunities to evaluate the role of local adaptation during colonization of new environments. Among these species, the Asian tiger mosquito, Aedes albopictus, is a threatening vector of several human viral diseases, including dengue and chikungunya, and raises concerns about the Zika fever. Its broad presence in both temperate and tropical environments has been considered the reflection of great "ecological plasticity." However, no study has been conducted to assess the role of adaptive evolution in the ecological success of Ae. albopictus at the molecular level. In the present study, we performed a genomic scan to search for potential signatures of selection leading to local adaptation in one-hundred-forty field-collected mosquitoes from native populations of Vietnam and temperate invasive populations of Europe. High-throughput genotyping of transposable element insertions led to the discovery of more than 120,000 polymorphic loci, which, in their great majority, revealed a virtual absence of structure between the biogeographic areas. Nevertheless, 92 outlier loci showed a high level of differentiation between temperate and tropical populations. The majority of these loci segregate at high insertion frequencies among European populations, indicating that this pattern could have been caused by recent adaptive evolution events in temperate areas. An analysis of the overlapping and neighbouring genes highlighted several candidates, including diapause, lipid and juvenile hormone pathways.

Two host clades, two bacterial arsenals: evolution through gene losses in facultative endosymbionts.

Bacterial endosymbiosis is an important evolutionary process in insects, which can harbor both obligate and facultative symbionts. The evolution of these symbionts is driven by evolutionary convergence, and they exhibit among the tiniest genomes in prokaryotes. The large host spectrum of facultative symbionts and the high diversity of strategies they use to infect new hosts probably impact the evolution of their genome and explain why they undergo less severe genomic erosion than obligate symbionts. Candidatus Hamiltonella defensa is suitable for the investigation of the genomic evolution of facultative symbionts because the bacteria are engaged in specific relationships in two clades of insects. In aphids, H. defensa is found in several species with an intermediate prevalence and confers protection against parasitoids. In whiteflies, H. defensa is almost fixed in some species of Bemisia tabaci, which suggests an important role of and a transition toward obligate symbiosis. In this study, comparisons of the genome of H. defensa present in two B. tabaci species (Middle East Asia Minor 1 and Mediterranean) and in the aphid Acyrthosiphon pisum revealed that they belong to two distinct clades and underwent specific gene losses. In aphids, it contains highly virulent factors that could allow protection and horizontal transfers. In whiteflies, the genome lost these factors and seems to have a limited ability to acquire genes. However it contains genes that could be involved in the production of essential nutrients, which is consistent with a primordial role for this symbiont. In conclusion, although both lineages of H. defensa have mutualistic interactions with their hosts, their genomes follow distinct evolutionary trajectories that reflect their phenotype and could have important consequences on their evolvability.

Spodoptera frugiperda (Lepidoptera: Noctuidae) host-plant variants: two host strains or two distinct species?

The moth Spodoptera frugiperda is a well-known pest of crops throughout the Americas, which consists of two strains adapted to different host-plants: the first feeds preferentially on corn, cotton and sorghum whereas the second is more associated with rice and several pasture grasses. Though morphologically indistinguishable, they exhibit differences in their mating behavior, pheromone compositions, and show development variability according to the host-plant. Though the latter suggest that both strains are different species, this issue is still highly controversial because hybrids naturally occur in the wild, not to mention the discrepancies among published results concerning mating success between the two strains. In order to clarify the status of the two host-plant strains of S. frugiperda, we analyze features that possibly reflect the level of post-zygotic isolation: (1) first generation (F1) hybrid lethality and sterility; (2) patterns of meiotic segregation of hybrids in reciprocal second generation (F2), as compared to the meiosis of the two parental strains. We found a significant reduction of mating success in F1 in one direction of the cross and a high level of microsatellite markers showing transmission ratio distortion in the F2 progeny. Our results support the existence of post-zygotic reproductive isolation between the two laboratory strains and are in accordance with the marked level of genetic differentiation that was recovered between individuals of the two strains collected from the field. Altogether these results provide additional evidence in favor of a sibling species status for the two strains.

Optimization of multiplexed RADseq libraries using low-cost adaptors.

Reduced representation genomics approaches, of which RADseq is currently the most popular form, offer the possibility to produce genome wide data from potentially any species, without previous genomic information. The application of RADseq to highly multiplexed libraries (including numerous specimens, and potentially numerous different species) is however limited by technical constraints. First, the cost of synthesis of Illumina adaptors including molecular identifiers (MIDs) becomes excessive when numerous specimens are to be multiplexed. Second, the necessity to empirically adjust the ratio of adaptors to genomic DNA concentration impedes the high throughput application of RADseq to heterogeneous samples, of variable DNA concentration and quality. In an attempt to solve these problems, we propose here some adjustments regarding the adaptor synthesis. First, we show that the common and unique (MID) parts of adaptors can be synthesized separately and subsequently ligated, which drastically reduces the synthesis cost, and thus allows multiplexing hundreds of specimens. Second, we show that self-ligation of adaptors, which makes the adaptor concentration so critical, can be simply prevented by using unphosphorylated adaptors, which significantly improves the ligation and sequencing yield.

Detection of genetically isolated entities within the Mediterranean species of Bemisia tabaci: new insights into the systematics of this worldwide pest.

BACKGROUND: The taxonomy of the species complex Bemisia tabaci, a serious agricultural pest worldwide, is not well resolved yet, even though species delimitation is critical for designing effective control strategies. Based on a threshold of 3.5% mitochondrial (mtCOI) sequence divergence, recent studies have identified 28 putative species. Among them, mitochondrial variability associated with particular symbiotic compositions (=cytotypes) can be observed, as in MED, which raises the question of whether it is a single or a complex of biological species. RESULTS: Using microsatellites, an investigation was made of the genetic relatedness of Q1 and ASL cytotypes that belong to MED. Samples of the two cytotypes were collected in West Africa where they live in sympatry on the same hosts. Genotyping revealed a high level of differentiation, without evidence of gene flow. Moreover, they differed highly in frequencies of resistance alleles to insecticides, which were much higher in Q1 than in ASL. CONCLUSION: Q1 and ASL are sufficiently reproductively isolated for the introgression of neutral alleles to be prevented, suggesting that they are actually different species. This indicates that nuclear genetic differentiation must be investigated within groups with less than 3.5% mtCOI divergence in order to elucidate the taxonomy of B. tabaci at a finer level. Overall, these data provide important information for pest management.

Specific activation of an I-like element in Drosophila interspecific hybrids.

The non-long terminal repeat (LTR) retrotransposon I, which belongs to the I superfamily of non-LTR retrotransposons, is well known in Drosophila because it transposes at a high frequency in the female germline cells in I-R hybrid dysgenic crosses of Drosophila melanogaster. Here, we report the occurrence and the upregulation of an I-like element in the hybrids of two sister species belonging to the repleta group of the genus Drosophila, D. mojavensis, and D. arizonae. These two species display variable degrees of pre- and postzygotic isolation, depending on the geographic origin of the strains. We took advantage of these features to explore the transposable element (TE) dynamics in interspecific crosses. We fully characterized the copies of this TE family in the D. mojavensis genome and identified at least one complete copy. We showed that this element is transcriptionally active in the ovaries and testes of both species and in their hybrids. Moreover, we showed that this element is upregulated in hybrid males, which could be associated with the male-sterile phenotype.

Biomphalysin, a new ß pore-forming toxin involved in Biomphalaria glabrata immune defense against Schistosoma mansoni.

Aerolysins are virulence factors belonging to the ß pore-forming toxin (ß-PFT) superfamily that are abundantly distributed in bacteria. More rarely, ß-PFTs have been described in eukaryotic organisms. Recently, we identified a putative cytolytic protein in the snail, Biomphalaria glabrata, whose primary structural features suggest that it could belong to this ß-PFT superfamily. In the present paper, we report the molecular cloning and functional characterization of this protein, which we call Biomphalysin, and demonstrate that it is indeed a new eukaryotic ß-PFT. We show that, despite weak sequence similarities with aerolysins, Biomphalysin shares a common architecture with proteins belonging to this superfamily. A phylogenetic approach revealed that the gene encoding Biomphalysin could have resulted from horizontal transfer. Its expression is restricted to immune-competent cells and is not induced by parasite challenge. Recombinant Biomphalysin showed hemolytic activity that was greatly enhanced by the plasma compartment of B. glabrata. We further demonstrated that Biomphalysin with plasma is highly toxic toward Schistosoma mansoni sporocysts. Using in vitro binding assays in conjunction with Western blot and immunocytochemistry analyses, we also showed that Biomphalysin binds to parasite membranes. Finally, we showed that, in contrast to what has been reported for most other members of the family, lytic activity of Biomphalysin is not dependent on proteolytic processing. These results provide the first functional description of a mollusk immune effector protein involved in killing S. mansoni.

Endosymbiont diversity among sibling weevil species competing for the same resource.

BACKGROUND: Whereas the impact of endosymbionts on the ecology of their hosts is well known in some insect species, the question of whether host communities are influenced by endosymbionts remains largely unanswered. Notably, the coexistence of host species competing with each other, which is expected to be stabilized by their ecological differences, could be facilitated by differences in their endosymbionts. Yet, the composition of endosymbiotic communities housed by natural communities of competing host species is still almost unknown. In this study, we started filling this gap by describing and comparing the bacterial endosymbiotic communities of four sibling weevil species (Curculio spp.) that compete with each other to lay eggs into oak acorns (Quercus spp.) and exhibit marked ecological differences. RESULTS: All four species housed the primary endosymbiont Candidatus Curculioniphilus buchneri, yet each of these had a clearly distinct community of secondary endosymbionts, including Rickettsia, Spiroplasma, and two Wolbachia strains. Notably, three weevil species harbored their own predominant facultative endosymbiont and possessed the remaining symbionts at a residual infection level. CONCLUSIONS: The four competing species clearly harbor distinct endosymbiotic communities. We discuss how such endosymbiotic communities could spread and keep distinct in the four insect species, and how these symbionts might affect the organization and species richness of host communities.

Influence of Wolbachia on host gene expression in an obligatory symbiosis.

BACKGROUND: Wolbachia are intracellular bacteria known to be facultative reproductive parasites of numerous arthropod hosts. Apart from these reproductive manipulations, recent findings indicate that Wolbachia may also modify the host's physiology, notably its immune function. In the parasitoid wasp, Asobara tabida, Wolbachia is necessary for oogenesis completion, and aposymbiotic females are unable to produce viable offspring. The absence of egg production is also associated with an increase in programmed cell death in the ovaries of aposymbiotic females, suggesting that a mechanism that ensures the maintenance of Wolbachia in the wasp could also be responsible for this dependence. In order to decipher the general mechanisms underlying host-Wolbachia interactions and the origin of the dependence, we developed transcriptomic approaches to compare gene expression in symbiotic and aposymbiotic individuals. RESULTS: As no genetic data were available on A. tabida, we constructed several Expressed Sequence Tags (EST) libraries, and obtained 12,551 unigenes from this species. Gene expression was compared between symbiotic and aposymbiotic ovaries through in silico analysis and in vitro subtraction (SSH). As pleiotropic functions involved in immunity and development could play a major role in the establishment of dependence, the expression of genes involved in oogenesis, programmed cell death (PCD) and immunity (broad sense) was analyzed by quantitative RT-PCR. We showed that Wolbachia might interfere with these numerous biological processes, in particular some related to oxidative stress regulation. We also showed that Wolbachia may interact with immune gene expression to ensure its persistence within the host. CONCLUSIONS: This study allowed us to constitute the first major dataset of the transcriptome of A. tabida, a species that is a model system for both host/Wolbachia and host/parasitoid interactions. More specifically, our results highlighted that symbiont infection may interfere with numerous pivotal processes at the individual level, suggesting that the impact of Wolbachia should also be investigated beyond reproductive manipulations.

Evidence of diversity and recombination in Arsenophonus symbionts of the Bemisia tabaci species complex.

BACKGROUND: Maternally inherited bacterial symbionts infecting arthropods have major implications on host ecology and evolution. Among them, the genus Arsenophonus is particularly characterized by a large host spectrum and a wide range of symbiotic relationships (from mutualism to parasitism), making it a good model to study the evolution of host-symbiont associations. However, few data are available on the diversity and distribution of Arsenophonus within host lineages. Here, we propose a survey on Arsenophonus diversity in whitefly species (Hemiptera), in particular the Bemisia tabaci species complex. This polyphagous insect pest is composed of genetic groups that differ in many ecological aspects. They harbor specific bacterial communities, among them several lineages of Arsenophonus, enabling a study of the evolutionary history of these bacteria at a fine host taxonomic level, in association to host geographical range and ecology. RESULTS: Among 152 individuals, our analysis identified 19 allelic profiles and 6 phylogenetic groups, demonstrating this bacterium's high diversity. These groups, based on Arsenophonus phylogeny, correlated with B. tabaci genetic groups with two exceptions reflecting horizontal transfers. None of three genes analyzed provided evidence of intragenic recombination, but intergenic recombination events were detected. A mutation inducing a STOP codon on one gene in a strain infecting one B. tabaci genetic group was also found. Phylogenetic analyses of the three concatenated loci revealed the existence of two clades of Arsenophonus. One, composed of strains found in other Hemiptera, could be the ancestral clade in whiteflies. The other, which regroups strains found in Hymenoptera and Diptera, may have been acquired more recently by whiteflies through lateral transfers. CONCLUSIONS: This analysis of the genus Arsenophonus revealed a diversity within the B. tabaci species complex which resembles that reported on the larger scale of insect taxonomy. We also provide evidence for recombination events within the Arsenophonus genome and horizontal transmission of strains among insect taxa. This work provides further insight into the evolution of the Arsenophonus genome, the infection dynamics of this bacterium and its influence on its insect host's ecology.