Multilocus phylogeny and ecological differentiation of the "Eupelmus urozonus species group" (Hymenoptera, Eupelmidae) in the West-Palaearctic.

BACKGROUND: The ecological differentiation of insects with parasitic life-style is a complex process that may involve phylogenetic constraints as well as morphological and/or behavioural adaptations. In most cases, the relative importance of these driving forces remains unexplored. We investigate here this question for the "Eupelmus urozonus species group" which encompasses parasitoid wasps of potential interest in biological control. This was achieved using seven molecular markers, reliable records on 91 host species and a proxy of the ovipositor length. RESULTS: After using an adequate partitioning scheme, Maximum likelihood and Bayesian approaches provide a well-resolved phylogeny supporting the monophyly of this species group and highlighting its subdivision into three sub-groups. Great variations of both the ovipositor length and the host range (specialist versus generalist) were observed at this scale, with these two features being not significantly constrained by the phylogeny. Ovipositor length was not shown as a significant predictor of the parasitoid host range. CONCLUSIONS: This study provides firstly the first evidence for the strong lability of both the ovipositor's length and the realised host range in a set of phylogenetically related and sympatric species. In both cases, strong contrasts were observed between sister species. Moreover, no significant correlation was found between these two features. Alternative drivers of the ecological differentiation such as interspecific interactions are proposed and the consequences on the recruitment of these parasitoids on native and exotic pests are discussed.

Ficus racemosa is pollinated by a single population of a single agaonid wasp species in continental South-East Asia.

High specificity in the Ficus-agaonid wasp mutualism has lead to the assumption of a mostly 'one-to-one' relationship, albeit with some exceptions. This view has been challenged by new molecular data in recent years, but surprisingly little is known about local and spatial genetic structuring of agaonid wasp populations. Using microsatellite markers, we analysed genetic structuring of Ceratosolen fusciceps, the fig wasp pollinating Ficus racemosa, a fig tree species widely distributed from India to Australia. In sampling stretching from the south of China to the south of Thailand we found evidence for only a single pollinating wasp species in continental South-East Asian mainland. We found no evidence for the co-occurrence of cryptic species within our subcontinent sampling zone. We observed no spatial genetic structure within sites and only limited structuring over the whole sampling zone, suggesting that F. racemosa is pollinated by a single population of a single agaonid wasp species all over continental South-East Asia. An additional sample of wasps collected on F. racemosa in Australia showed clear-cut genetic differentiation from the Asian continent, suggesting allopatric divergence into subspecies or species. We propose that the frequent local co-occurrence of sister species found in the literature mainly stems from contact zones between biogeographic regions, and that a single pollinator species over wide areas might be the more common situation everywhere else.

Phylogeny of the genus Aphis Linnaeus, 1758 (Homoptera: Aphididae) inferred from mitochondrial DNA sequences.

Aphis is the largest aphid genus in the world and contains several of the most injurious aphid pests. It is also the most reluctant aphid genus to any comprehensive taxonomic treatment: while most species are easily classified into "species groups" that form well defined entities, numerous species within these groups are difficult to tell apart morphologically and identification keys remain ambiguous and mostly rely on host plant affiliation. In this paper, we used partial sequences of COI/COII and CytB genes to reconstruct the first phylogeny of Aphis and discuss the present systematics. The monophyly of the subgenus Bursaphis and of the tree major species groups, Black aphid, Black backed aphid and frangulae-like species was recovered by all phylogenetic analyses. However our data suggested that the nominal subgenus was not monophyletic. Relationships between major species groups were often ambiguous but "Black" and "Black backed" species groups appeared as sister clades. The most striking result of this study was that our molecular data met the same limits as the morphological characters used in classifications: mitochondrial DNA did not allow the differentiation of species that are difficult to identify. Further, interspecies relationships within groups of species for which taxonomic treatment is difficult stayed unresolved. This suggests that species delineation in the genus Aphis is often ambiguous and that diversification might have been a rapid process.

Patterns of diversification of Afrotropical Otiteselline fig wasps: phylogenetic study reveals a double radiation across host figs and conservatism of host association.

We studied the phylogenetic relationships of Otiteselline fig waSPS associated with Ficus in the Afrotropical region using rDNA sequences. African fig species usually host two species of Otiteselline fig waSPS. Phylogenetic analyses reveal that this pattern of association results from the radiation of two clades of waSPS superimposed on the fig system. Within each clade, wasp species generally cluster according to their host classification. The phylogenies of the two clades are also generally more congruent than expected by chance. Together these results suggest that Otiteselline wasp speciation is largely constrained by the diversification of their hosts. Finally, we show a difference in ovipositor length between the two Otiteselline species coexisting in the same Ficus species, which probably corresponds to ecological differences. The diversification of ecological niches within the fig is probably, with cospeciation, one of the key factors explaining the diversification and maintenance of species of parasites of the fig/pollinator system.

When morphometry meets genetics: inferring the phylogeography of Carabus solieri using Fourier analyses of pronotum and male genitalia.

Population differentiation is a crucial step in the speciation process and is therefore a central subject in studies of microevolution. Assessing divergence and inferring its dynamics in space and time generally require a wide array of markers. Until now however, most studies of population structure are based on molecular markers and those concerning morphological traits are more scarce. In the present work, we studied morphological differentiation among populations of the ground beetle Carabus solieri, and tested its congruence with genetic population structure. The shape of pronotum and aedeagus was assessed using Dual Axis Fourier Shape Analysis. manova on Fourier coefficients revealed highly significant morphological variation between populations and a similar geographical pattern of differentiation for both structures. On the whole, morphological and genetic patterns were also found to be congruent. Our analysis confirms the phylogeographical scenario proposing that two entities of C. solieri differentiated during the last glaciation events before recolonizing the actual range of the species. It also indicates a large introgression between the two differentiated entities in the centre of the range.

Nest architecture and genetic differentiation in a species complex of Australian stingless bees.

We investigated the taxonomic significance of nest shape and its putative role in speciation in Trigona (Heterotrigona) carbonaria and T. (H.) hockingsi, two sibling species of stingless bee species from eastern Australia. These species are primarily distinguished by their nest architecture, as in all other respects they are nearly identical. We genotyped 130 colonies from six locations in Queensland at 13 microsatellite loci together with 106 additional colonies from six other Indo-Pacific Trigona species. Whether they were present in allopatry or in sympatry, colonies that displayed the T. carbonaria or the T. hockingsi nest architecture could be unambiguously differentiated at the genetic level. However, T. hockingsi colonies were classifiable into two highly differentiated paraphyletic and geographically separate populations, one in northern and one in southern Queensland. These two populations probably belong to two distinct species, T. hockingsi and T. davenporti nov. sp. Our results suggest that nest architecture characters are relevant but not sufficient criteria to identify species in this group. Consequently, modifications of nest architecture are probably not of prime importance in the speciation process of Australian stingless bees, although nest architecture differences probably result from relatively simple mechanisms. The rare interspecific hybrid colonies detected did not display a nest with an intermediate form between T. hockingsi and T. carbonaria.

Specialization and habitat: spatial and environmental effects on abundance and genetic diversity of forest generalist and specialist Carabus species.

Habitat specialist species are supposed to be more susceptible to variations in local environmental characteristics than generalists. To test this hypothesis, we conducted a comparative analysis on abundance and genetic diversity of forest carabids differing in their habitat requirements. Four species were sampled in forests characterized by abiotic, landscape and biotic environmental variables. A statistical framework based on canonical correspondence analysis was used for one habitat generalist and one habitat specialist species to determine the relative contribution of environmental variables in structuring inter- and intrapopulational genetic diversity depicted by microsatellites. Our results showed that sympatric species differed in their sensitivity to environmental variables. The same variables were found to be important in analyses of abundance and genetic data. However, specialization was not related to a greater sensitivity to local environmental characteristics. The strong impact of spatial variables on genetic data suggested that genetic variation among populations would largely reflect the response of individual species to dispersal opportunities more than the effect of habitat quality.

Isolation by distance and sharp discontinuities in gene frequencies: implications for the phylogeography of an alpine insect species, Carabus solieri.

Analysis of genetic isolation by distance (IBD) is of prime importance for the study of processes responsible for spatial population genetic structure and is thus frequently used in case studies. However, the identification of a significant IBD pattern does not necessarily imply the absence of sharp discontinuities in gene frequencies. Therefore, identifying barriers to gene flow and/or secondary contact between differentiated entities remains a major challenge in population biology. Geographical genetic structure of 41 populations (1080 individuals) of an alpine insect species, Carabus solieri, was studied using 10 microsatellite loci. All populations were significantly differentiated and spatially structured according to IBD over the entire range. However, clustering analyses clearly identified three main clusters of populations, which correspond to geographical entities. Whereas IBD also occurs within each cluster, population structure was different according to which group of populations was considered. The southernmost cluster corresponds to the most fragmented part of the range. Consistently, it was characterized by relatively high levels of differentiation associated with low genetic diversity, and the slope of the regression of genetic differentiation against geographical distances was threefold those of the two other clusters. Comparisons of within-cluster and between-cluster IBD patterns revealed barriers to gene flow. A comparison of the two approaches, IBD and clustering analyses, provided us with valuable information with which to infer the phylogeography of the species, and in particular to propose postglacial colonization routes from two potential refugia located in Italy and in southeastern France. Our study highlights strongly the possible confounding contribution of barriers to gene flow to IBD pattern and emphasizes the utility of the model-based clustering analysis to identify such barriers.

Population genetic structure of rock ptarmigan Lagopus mutus in Northern and Western Europe.

Alpine species may be losing habitat because of global warming. Setting management priorities for such species is thus urgent and cannot be achieved without data on population structure. We studied the structure of rock ptarmigan (Lagopus mutus) populations in the Pyrenees, Alps and Norway, using six microsatellites. We found that rock ptarmigan in the Pyrenees were genetically impoverished compared with those in the Alps and Norway, and displayed a greater divergence (Pyrenees vs. Alps or Norway: theta(ST) = 0.16, Alps vs. Norway, theta(ST) = 0.04). In the Alps, despite a weak genetic differentiation between localities up to 200 km apart (theta(ST) = 0.011), a significant isolation-by-distance (IBD) effect was detected. When computed for each sex separately this IBD effect was significant for males but not for females, suggesting that males are highly philopatric.

Fine-scale genetic structure of two carabid species with contrasted levels of habitat specialization.

Using microsatellite markers, we compared the genetic structure of populations of two carabid species, one described as a generalist (commonly found in forest and in open habitats) and the other known as a forest specialist. Both species were sampled in the same forest plots, which were separated from each other by either open or forested areas. At the local scale considered (13.6 km separating the most distant plots), genetic differentiation was substantial for both species studied, but populations of the forest specialist Carabus punctatoauratus appeared to be more spatially structured than those of C. nemoralis. Isolation by distance analyses showed that nonforested areas are partial barriers to gene flow for both species studied, although more clearly for the forest specialist. Between and within forests, dispersal capacity of the generalist C. nemoralis was shown to be higher than that of the specialist C. punctatoauratus.

Molecular phylogenies of fig wasps: partial cocladogenesis of pollinators and parasites.

Figs (Ficus spp., Moraceae) and their pollinating wasps form an obligate mutualism, which has long been considered a classic case of coevolution and cospeciation. Figs are also exploited by several clades of nonpollinating wasps, which are parasites of the mutualism and whose patterns of speciation have received little attention. We used data from nuclear and mitochondrial DNA regions to estimate the phylogenies of 20 species of Pleistodontes pollinating wasps and 16 species of Sycoscapter nonpollinating wasps associated with Ficus species in the section Malvanthera. We compare the phylogenies of 15 matched Pleistodontes/Sycoscapter species pairs and show that the level of cospeciation is significantly greater than that expected by chance. Our estimates of the maximum level of cospeciation (50 to 64% of nodes) are very similar to those obtained in other recent studies of coevolved parasitic and mutualistic associations. However, we also show that there is not perfect congruence of pollinator and parasite phylogenies (for any substantial clade) and argue that host plant switching is likely to be less constrained for Sycoscapter parasites than for Pleistodontes pollinators. There is perfect correspondence between two terminal clades of two sister species in the respective phylogenies, and rates of molecular evolution in these pairs are similar.

Pollination mode in fig wasps: the predictive power of correlated traits.

The over 700 species of Ficus are thought to have co-speciated with their obligate pollinators (family Agaonidae). Some of these wasp species pollinate figs actively, while others are passive pollinators. Based on direct observations of mode of pollination in 88 species, we show that mode of pollination can confidently be predicted from fig traits only (anther-to-ovule ratio) or from wasp traits only (presence of coxal combs). The presence of pollen pockets is not a predictor of mode of pollination. Data, direct and indirect, on 142 species, demonstrate numerous cases of the loss of active pollination and suggest one or few origins of active pollination. Hence, active pollination, an impressive example of the sophisticated traits that may result from mutualistic coevolution, depends on selective forces that can be overcome in some species, allowing reversions. Despite frequent loss, active pollination remains the predominant mode of pollination in Ficus.

Distribution and phylogeny of Wolbachia inducing thelytoky in Rhoditini and 'Aylacini' (Hymenoptera: Cynipidae).

Wolbachia are endosymbiotic bacteria responsible for thelytoky in several parasitoid hymenopteran genera. After finding these micro-organisms in some populations of Diplolepis spinosissimae (Hymenoptera: Cynipidae) where they are responsible for thelytoky through gamete duplication, we searched for Wolbachia spp. using specific PCR primers in nineteen other species of the Rhoditini tribe (rose gallwasps) and eight species of the 'Aylacini' tribe (gallwasps associated with herbaceous plants). Wolbachia were found in twelve Rhoditini species and four 'Aylacini' species. The most infected species have very few males (spanandry) and the thelytoky of infected species/arrhenotoky of uninfected species is confirmed by previous research based on the sex of the offspring of virgin females. Phylogenetic analyses based on the partial Wolbachia ftsZ gene sequences indicate that some strains associated with closely related gallwasps are phylogenetically distant, suggesting that cynipids have been affected by several infection events. In contrast, the five infected European species of Diplolepis harbour the same strain of Wolbachia.

Molecular phylogeny of the Ceratosolen species pollinating Ficus of the subgenus Sycomorus sensu stricto: biogeographical history and origins of the species-specificity breakdown cases.

The 14 species of Ficus of the subgenus Sycomorus (Moraceae) are invariably pollinated by Ceratosolen species (Hym. Chalcidoidea), which in turn reproduce in the fig florets. They are distributed mostly in continental Africa, Madagascar, and the Mascarene and Comoro Islands, but 1 species extends its geographical range all over the Oriental region. Fig-pollinator relationships are usually strictly species specific, but exceptions to the 'one-to-one' rule occur within the group we studied. In order to understand both the biogeographical history of the Ceratosolen species associated with Ficus of the subgenus Sycomorus and the origins of the specificity breakdown cases, we have used cytochrome b sequences to reconstruct a phylogeny of the fig wasps. The results show that the pollinators from the Malagasy region and those from continental Africa form two distinct clades, which probably diverged after the crossing of the Mozambique Channel by an ancestral population. The Oriental wasp species show strong affinities with the African species. The two species-specificity exceptions are due to different evolutionary events. The occurrence of the two West African pollinators associated with F. sur can be explained by successive speciation events of the mutualistic partner without plant radiation. In contrast, we hypothesize that C. galili shifted by horizontal transfer from an unknown, presumably extinct, Ficus species to F. sycomorus after this native Malagasy fig species colonized Africa.

Molecular phylogeny of fig wasps Agaonidae are not monophyletic.

According to the present classification, the family Agaonidae contains all fig pollinators as well as five subfamilies of non-pollinating fig wasps. The molecular phylogeny of the family was reconstructed using partial sequences of the 28S rRNA (D1 and D2 domains). Our results show that the family Agaonidae is not monophyletic. As a consequence, we restrict the famiy to the pollinator clade, and assign the non-pollinating subfamilies to various chalcid families. Sycoecinae, Otitesellinae and Sycoryctinae are included in Pteromalidae, whereas Sycophaginae and Epichrysomallinae are left unclassified and will require more in-depth morphological studies. Moreover, we proved that the fig pollination syndrome evolved only once, early in group history. The resource due to the fig-pollinator mutualism has secondarily been colonized independently by different Chalcid lineages.