Echoes of ancient introgression punctuate stable genomic lineages in the evolution of figs.

Studies investigating the evolution of flowering plants have long focused on isolating mechanisms such as pollinator specificity. Some recent studies have proposed a role for introgressive hybridization between species, recognizing that isolating processes such as pollinator specialization may not be complete barriers to hybridization. Occasional hybridization may therefore lead to distinct yet reproductively connected lineages. We investigate the balance between introgression and reproductive isolation in a diverse clade using a densely sampled phylogenomic study of fig trees (Ficus, Moraceae). Codiversification with specialized pollinating wasps (Agaonidae) is recognized as a major engine of fig diversity, leading to about 850 species. Nevertheless, some studies have focused on the importance of hybridization in Ficus, highlighting the consequences of pollinator sharing. Here, we employ dense taxon sampling (520 species) throughout Moraceae and 1,751 loci to investigate phylogenetic relationships and the prevalence of introgression among species throughout the history of Ficus. We present a well-resolved phylogenomic backbone for Ficus, providing a solid foundation for an updated classification. Our results paint a picture of phylogenetically stable evolution within lineages punctuated by occasional local introgression events likely mediated by local pollinator sharing, illustrated by clear cases of cytoplasmic introgression that have been nearly drowned out of the nuclear genome through subsequent lineage fidelity. The phylogenetic history of figs thus highlights that while hybridization is an important process in plant evolution, the mere ability of species to hybridize locally does not necessarily translate into ongoing introgression between distant lineages, particularly in the presence of obligate plant-pollinator relationships.

The evolution of parasitism from mutualism in wasps pollinating the fig, Ficus microcarpa, in Yunnan Province, China.

Theory identifies factors that can undermine the evolutionary stability of mutualisms. However, theory's relevance to mutualism stability in nature is controversial. Detailed comparative studies of parasitic species that are embedded within otherwise mutualistic taxa (e.g., fig pollinator wasps) can identify factors that potentially promote or undermine mutualism stability. We describe results from behavioral, morphological, phylogenetic, and experimental studies of two functionally distinct, but closely related, Eupristina wasp species associated with the monoecious host fig, Ficus microcarpa, in Yunnan Province, China. One (Eupristina verticillata) is a competent pollinator exhibiting morphologies and behaviors consistent with observed seed production. The other (Eupristina sp.) lacks these traits, and dramatically reduces both female and male reproductive success of its host. Furthermore, observations and experiments indicate that individuals of this parasitic species exhibit greater relative fitness than the pollinators, in both indirect competition (individual wasps in separate fig inflorescences) and direct competition (wasps of both species within the same fig). Moreover, phylogenetic analyses suggest that these two Eupristina species are sister taxa. By the strictest definition, the nonpollinating species represents a "cheater" that has descended from a beneficial pollinating mutualist. In sharp contrast to all 15 existing studies of actively pollinated figs and their wasps, the local F. microcarpa exhibit no evidence for host sanctions that effectively reduce the relative fitness of wasps that do not pollinate. We suggest that the lack of sanctions in the local hosts promotes the loss of specialized morphologies and behaviors crucial for pollination and, thereby, the evolution of cheating.

East‒West genetic differentiation across the Indo-Burma hotspot: evidence from two closely related dioecious figs.

BACKGROUND: Understanding biodiversity patterns and their underlying mechanisms is of interest to ecologists, biogeographers and conservationists and is critically important for conservation efforts. The Indo-Burma hotspot features high species diversity and endemism, yet it also faces significant threats and biodiversity losses; however, few studies have explored the genetic structure and underlying mechanisms of Indo-Burmese species. Here, we conducted a comparative phylogeographic analysis of two closely related dioecious Ficus species, F. hispida and F. heterostyla, based on wide and intensive population sampling across Indo-Burma ranges, using chloroplast (psbA-trnH, trnS-trnG) and nuclear microsatellite (nSSR) markers, as well as ecological niche modeling. RESULTS: The results indicated large numbers of population-specific cpDNA haplotypes and nSSR alleles in the two species. F. hispida showed slightly higher chloroplast diversity but lower nuclear diversity than F. heterostyla. Low-altitude mountainous areas of northern Indo-Burma were revealed to have high genetic diversity and high habitat suitability, suggesting potential climate refugia and conservation priority areas. Strong phylogeographic structure and a marked east‒west differentiation pattern were observed in both species, due to the interactions between biotic and abiotic factors. Interspecific dissimilarities at fine-scale genetic structure and asynchronized historical dynamics of east‒west differentiation between species were also detected, which were attributed to different species-specific traits. CONCLUSIONS: We confirm hypothesized predictions that interactions between biotic and abiotic factors largely determine the patterns of genetic diversity and phylogeographic structure of Indo-Burmese plants. The east‒west genetic differentiation pattern observed in two targeted figs can be generalized to some other Indo-Burmese plants. The results and findings of this work will contribute to the conservation of Indo-Burmese biodiversity and facilitate targeted conservation efforts for different species.

Changes in temperature alter competitive interactions and overall structure of fig wasp communities.

Organisms exist within ecological networks, connected through interactions such as parasitism, predation and mutualism which can modify their abundance and distribution within habitat patches. Differential species responses make it hard to predict the influence of climate change at the community scale. Understanding the interplay between climate and biotic interactions can improve our predictions of how ecosystems will respond to current global warming. We aim to understand how climate affects the multitrophic biotic interactions as well as the community structure using the enclosed communities of wasps associated with figs as study system. To examine the presence and strength of multitrophic species interactions, we first characterized the multitrophic community of fig wasps associated with Ficus racemosa and then applied hierarchical joint species distribution models, fitted to community monitoring data. We further evaluated the effect of climate on individual species trends as well as interspecific interactions. We found that the competitive balance shifted to favour non-pollinating galling wasps and disadvantage the dominant pollinator in suboptimal conditions. Furthermore, suboptimal conditions for galling wasps facilitated the occurrence of their specialized parasitoid, as changes cascaded across trophic levels and led to alternative community structures. Our results highlight the role of how species interactions can be modified across multiple trophic levels in a fig wasp community according to climate.

The nature of interspecific interactions and co-diversification patterns, as illustrated by the fig microcosm.

Interactions between mutualists, competitors, and antagonists have contrasting ecological effects that, sustained over generations, can influence micro- and macroevolution. Dissimilar benefits and costs for these interactions should cause contrasting co-diversification patterns between interacting clades, with prevalent co-speciation by mutualists, association loss by competitors, and host switching by antagonists. We assessed these expectations for a local assemblage of 26 fig species (Moraceae: Ficus), 26 species of mutualistic (pollinating), and 33 species of parasitic (galling) wasps (Chalcidoidea). Using newly acquired gene sequences, we inferred the phylogenies for all three clades. We then compared the three possible pairs of phylogenies to assess phylogenetic congruence and the relative frequencies of co-speciation, association duplication, switching, and loss. The paired phylogenies of pollinators with their mutualists and competitors were significantly congruent, unlike that of figs and their parasites. The distributions of macroevolutionary events largely agreed with expectations for mutualists and antagonists. By contrast, that for competitors involved relatively frequent association switching, as expected, but also unexpectedly frequent co-speciation. The latter result likely reflects the heterogeneous nature of competition among fig wasps. These results illustrate the influence of different interspecific interactions on co-diversification, while also revealing its dependence on specific characteristics of those interactions.

Enhanced and asymmetric signatures of hybridization at climatic margins: Evidence from closely related dioecious fig species.

Hybridization plays a significant role in biological evolution. However, it is not clear whether ecological contingency differentially influences likelihood of hybridization, particularly at ecological margins where parental species may exhibit reduced fitnesses. Moreover, it is unknown whether future ecosystem change will increase the prevalence of hybridization. Ficus heterostyla and F. squamosa are closely related species co-distributed from southern Thailand to southwest China where hybridization, yielding viable seeds, has been documented. As a robust test of ecological factors driving hybridization, we investigated spatial hybridization signatures based on nuclear microsatellites from extensive population sampling across a widespread contact range. Both species showed high population differentiation and strong patterns of isolation by distance. Admixture estimates exposed asymmetric interspecific gene flow. Signatures of hybridization increase significantly towards higher latitude zones, peaking at the northern climatic margins. Geographic variation in reproductive phenology combined with ecologically challenging marginal habitats may promote this phenomenon. Our work is a first systematic evaluation of such patterns in a comprehensive, latitudinally-based clinal context, and indicates that tendency to hybridize appears strongly influenced by environmental conditions. Moreover, that future climate change scenarios will likely alter and possibly augment cases of hybridization at ecosystem scales.

An extreme case of plant-insect codiversification: figs and fig-pollinating wasps.

It is thought that speciation in phytophagous insects is often due to colonization of novel host plants, because radiations of plant and insect lineages are typically asynchronous. Recent phylogenetic comparisons have supported this model of diversification for both insect herbivores and specialized pollinators. An exceptional case where contemporaneous plant-insect diversification might be expected is the obligate mutualism between fig trees (Ficus species, Moraceae) and their pollinating wasps (Agaonidae, Hymenoptera). The ubiquity and ecological significance of this mutualism in tropical and subtropical ecosystems has long intrigued biologists, but the systematic challenge posed by >750 interacting species pairs has hindered progress toward understanding its evolutionary history. In particular, taxon sampling and analytical tools have been insufficient for large-scale cophylogenetic analyses. Here, we sampled nearly 200 interacting pairs of fig and wasp species from across the globe. Two supermatrices were assembled: on an average, wasps had sequences from 77% of 6 genes (5.6 kb), figs had sequences from 60% of 5 genes (5.5 kb), and overall 850 new DNA sequences were generated for this study. We also developed a new analytical tool, Jane 2, for event-based phylogenetic reconciliation analysis of very large data sets. Separate Bayesian phylogenetic analyses for figs and fig wasps under relaxed molecular clock assumptions indicate Cretaceous diversification of crown groups and contemporaneous divergence for nearly half of all fig and pollinator lineages. Event-based cophylogenetic analyses further support the codiversification hypothesis. Biogeographic analyses indicate that the present-day distribution of fig and pollinator lineages is consistent with a Eurasian origin and subsequent dispersal, rather than with Gondwanan vicariance. Overall, our findings indicate that the fig-pollinator mutualism represents an extreme case among plant-insect interactions of coordinated dispersal and long-term codiversification. [Biogeography; coevolution; cospeciation; host switching; long-branch attraction; phylogeny.].

Description of two new species of Walkerella (Pteromalidae, Otitesellinae) from China with a key to species of the genus.

Walkerella is an Old World genus of non-pollinating fig wasps in the subfamily Otitesellinae (Chalcidoidea, Pteromalidae). It is the most widely distributed genus of the subfamily, though there has been only six known world species. This paper describes two new species associated with the host Ficus subsection Conosycea of section Urostigma, subgenus Urostigma in Xishuangbanna, China. The two new species, Walkerella nigrabdomina Ma & Yang sp. nov. and W. curtipedis Ma & Yang sp. nov., are described from specimens reared from Ficus pisocarpa and Ficus curtipes, respectively. A key to all described species of Walkerella found is provided.

Two new species of Ismarus Haliday (Hymenoptera, Ismaridae) from Yunnan, China.

The genus Ismarus Haliday are rarely collected parasitoids in the small family Ismaridae. In this study, two new species are described from China's Yunnan Province: Ismarusrobustus Chen & Yan, sp. nov. and Ismarusunisulcus Chen & Yan, sp. nov. An updated key to the Chinese species of the genus is provided.

Host insect specificity and interspecific competition drive parasitoid diversification in a plant-insect community.

Ecological interactions among plants, insect herbivores, and parasitoids are pervasive in nature and play important roles in community assembling, but the codiversification of tri-trophic interactions has received less attention. Here we compare pairwise codiversification patterns between a set of 22 fig species, their herbivorous pollinating and galling wasps, and their parasitoids. The parasitoid phylogeny showed significant congruence and more cospeciation events with host insects phylogeny than with host plants. These results suggest that parasitoid phylogeny and speciation is more closely related to their host insects than to their host plants. The pollinating wasps hosted more parasitoid species than gallers and indicated a more intense interspecific competition among parasitoids associated with pollinators. Closer matching and fewer evolutionary host shifts were found between parasitoids and galler hosts than between parasitoids and pollinator hosts. These results suggest that interspecific competition among parasitoids, rather than resource availability of host wasps, is the main driver of the codiversification pattern in this community. Therefore, our study highlights the important role of interspecific competition among high trophic level insects in plant-insect tri-trophic community assembling.

Pollinator sharing and hybridization in a pair of dioecious figs sheds light on the pathways to speciation.

The dynamics and processes underlying the codiversification of plant-pollinator interactions are of great interest to researchers of biodiversity and evolution. Cospeciation is generally considered a key process driving the diversity of figs and their pollinating wasps. Groups of closely related figs pollinated by separate wasps occur frequently and represent excellent opportunities to study ongoing diversification in this textbook mutualism. We study two closely related sympatric dioecious figs (Ficus heterostyla and Ficus squamosa) in Xishuangbanna, southwest China, and aim to document what is likely to be the final stages of speciation between these species using a combination of trait data and experimental manipulation. Volatile profiles at the receptive phase, crucial for attracting pollinators, were analyzed. In total, 37 and 29 volatile compounds were identified from receptive F. heterostyla and F. squamosa figs, respectively. Despite significant interspecific dissimilarity, 25 compounds were shared. Ovipositor lengths lie well within range required for access to heterospecific ovules, facilitating hybridization. Cross introduction of wasps into figs was conducted and hybrid seeds were generated for all donor/recipient combinations. F. heterostyla wasps produce adult offspring in F. squamosa figs. While F. squamosa wasps induce gall development in F. heterostyla figs and their offspring fail to mature in synchrony with their novel host. We record limited geographic barriers, minimal volatile dissimilarity, compatible morphology, complementary reproductive phenologies, and the production of hybrid seeds and wasp offspring. These findings suggest ongoing wasp specialization and reproductive isolation, potentially applicable to other related fig species.

The genus Eurymeros Bhat (Hymenoptera, Braconidae, Alysiinae) newly recorded from China.

Background: Alysiinae Leach is a species-rich subfamily in Braconidae, of which several species play an important role in biological control. The monotypic genus Eurymerostumespiraculum Bhat, 1980 was discovered in Tibet and Yunnan provinces for the first time, representing the first record of the genus Eurymeros Bhat, 1980 (Braconidae, Alysiinae) in China. New information: The rare genus Eurymeros Bhat, 1980 (Braconidae, Alysiinae) and its only known species, E.tumespiraculum Bhat, 1980, are newly recorded from China. The morphological variation of the Chinese specimens is described and illustrated.

Phylogenetic analyses suggest a hybrid origin of the figs (Moraceae: Ficus) that are endemic to the Ogasawara (Bonin) Islands, Japan.

The Ogasawara Islands are oceanic islands and harbor a unique endemic flora. There are three fig species (Ficus boninsimae, F. nishimurae and F. iidaiana) endemic to the Ogasawara Islands, and these species have been considered to be closely related to Ficus erecta, and to have diverged within the islands. However, this hypothesis remains uncertain. To investigate this issue, we assessed the phylogenetic relationships of the Ogasawara figs and their close relatives occurring in Japan, Taiwan and South China based on six plastid genome regions, nuclear ITS region and two nuclear genes. The plastid genome-based tree indicated a close relationship between the Ogasawara figs and F. erecta, whereas some of the nuclear gene-based trees suggested this relationship was not so close. In addition, the phylogenetic analyses of the pollinating wasps associated with these fig species based on the nuclear 28S rRNA and mitochondrial cytB genes suggested that the fig-pollinating wasps of F. erecta are not sister to those of the Ogasawara figs These results suggest the occurrence of an early hybridization event(s) in the lineage leading to the Ogasawara figs.

Portable microsatellite primers for Ficus (Moraceae).

PREMISE OF THE STUDY: Highly portable microsatellite primers were developed for Ficus to facilitate investigation of genetic structure of complete regional floras using a single set of markers. METHODS AND RESULTS: Pyrosequencing of five species of Ficus produced a library of 5723 potential primers. Potential primers found in at least two species and presenting identical annealing temperatures were tested on a set of five additional Ficus species. A set of 20 primer pairs producing well-defined and easily readable peaks was retained and tests showed their potential utility for analyzing population genetic structure of 24 Ficus species from Taiwan. Numbers of alleles per locus ranged from one to six in the least variable species and from one to 17 in the most variable species. CONCLUSIONS: The results indicate that our set of primers can be used to analyze polymorphism and compare levels of polymorphism among Ficus species.

The complete chloroplast genome of two related fig species Ficus squamosa and Ficus heterostyla.

The Ficus squamosa and Ficus heterostyla share an undescribed pollinating fig wasp Ceratosolen sp. in Xishuangbanna region, which constitutes the most excellent model to study the role of convergent evolution and hybridization in the species-specific fig-wasp mutualism. The plastomes were 160,350 bp for Ficus squamosa and 160,300 bp for F. heterostyla, both in length with the typical quadripartite structure. In the two genomes, the LSC region was 88,615 bp (F. squamosa) and 88,535 bp (F. heterostyla), the SSC region was 20,071 bp (F. squamosa) and 20,101 bp (F. heterostyla), and the IR regions of both figs were 25,832 bp. They contained 113 unique genes, including a set of 78 protein-coding genes, 30 transfer RNA (tRNA) genes, four ribosomal RNA (rRNA) genes, and one pseudogene (infA). Phylogenetic analysis based on the complete chloroplast (cp) genomes within the Ficus genus suggests that they are closely related sister species.

Diversity begets diversity: Low resource heterogeneity reduces the diversity of nut-nesting ants in rubber plantations.

One of the most general patterns in ecology is the positive relationship between environmental heterogeneity and local diversity. On the one hand, increased resource heterogeneity provides more resources for diverse consumers in the community. On the other hand, increased structural heterogeneity creates variation in the environment's physical structure, thus allowing the coexistence of diverse species with different environmental requirements. Here, we examined the relative importance of resource and structural heterogeneity in determining the taxonomic, functional, and phylogenetic diversity of nut-nesting ants in natural rainforest and rubber plantation. The species richness of nut-nesting ants was 70% higher in rainforest than in rubber plantation. The clustered functional and phylogenetic structure in rubber plantation suggested a strong effect of environmental filtering in shaping ant functional and phylogenetic structure. Nesting heterogeneity (nut diversity) was the major factor explaining variation in taxonomic, functional, and phylogenetic diversity, suggesting that resource heterogeneity plays a major role in shaping the biodiversity patterns of nut-nesting ants. Overall, these results indicate that decreased resource diversity following the conversion of rainforest to rubber plantation can drive biodiversity loss in nut-nesting ants, through its effect on reducing both ant species, functional, and phylogenetic diversity. The decline in species richness and functional and phylogenetic diversity in the local ant community might have major effects on ecosystem functioning.

Updating the taxonomy of the ant genus Myrmecina (Hymenoptera, Formicidae) in China with descriptions of three new species.

In this study, we present a taxonomic update for the ant genus Myrmecina in China that includes 14 species in total. A recent survey of the leaf litter ant fauna in Chinas Hengduan Mountains collected three unknown Myrmecina species, which we describe here as Myrmecina eowilsoni sp. nov., M. gaoligongensis sp. nov., and Myrmecina pierceae sp. nov. These new species are clearly distinguishable from all the other species in the genus. We provide an update to the identification key of Chinese Myrmecina, as well as a diagnostic discussion and high-quality specimen images. In addition, Myrmecina raviwonghei Jaitrong, Samung, Waengsothorn Okido 2019 is reported for the first time from China.

Overlaps in olfactive signalling coupled with geographic variation may result in localised pollinator sharing between closely related Ficus species.

BACKGROUND: In brood site pollination mutualisms, pollinators are attracted by odours emitted at anthesis. In Ficus, odours of receptive figs differ among species and the specific pollinators generally only enter figs of their host species ensuring a pre-zygotic barrier to plant interspecific hybridisation. However, field observations recorded that, in Guangdong province in China, Valisia javana hilli, the local pollinator of F. hirta, entered and reproduced successfully in the figs of the closely related F. triloba on a regular basis. We propose that closely related Ficus species produce similar receptive fig odours. Under particular contexts of odours locally present, the receptive fig odours of non-host figs of a Ficus species may become attractive to pollinators of closely related Ficus species. We used the headspace technique to collect in situ receptive fig odours of F. triloba in a series of locations in China. Under controlled conditions, we tested the attraction of fig pollinating wasps from F. hirta and F. triloba to host figs and non-host figs in Y tube experiments. RESULTS: Receptive fig odours of F. triloba though different from those of F. hirta, were mainly composed of a same set of volatile organic compounds. When given the choice between receptive fig odours and air, the pollinating wasps were only attracted by their host's odours. However, when given a choice between host and non-host figs the pollinators of F. hirta were equally attracted by the two odours while the pollinators of F. triloba tended to be more attracted by their host's fig odours. CONCLUSIONS: Receptive fig odours vary geographically within species and the differentiation of receptive fig odours between closely related Ficus species is often incomplete. This allows localised or occasional pollinator sharing following different modalities. Cross stimulation when wasps are exposed simultaneously to odours of host and non-host species may be important. While occasional pollinator sharing may play a marginal role when wasp populations are robust, it may ensure the provisioning of new pollinators from the closest relative of a Ficus species if its pollinators go extinct.

Comparative chloroplast genome analysis of Ficus (Moraceae): Insight into adaptive evolution and mutational hotspot regions.

As the largest genus in Moraceae, Ficus is widely distributed across tropical and subtropical regions and exhibits a high degree of adaptability to different environments. At present, however, the phylogenetic relationships of this genus are not well resolved, and chloroplast evolution in Ficus remains poorly understood. Here, we sequenced, assembled, and annotated the chloroplast genomes of 10 species of Ficus, downloaded and assembled 13 additional species based on next-generation sequencing data, and compared them to 46 previously published chloroplast genomes. We found a highly conserved genomic structure across the genus, with plastid genome sizes ranging from 159,929 bp (Ficus langkokensis) to 160,657 bp (Ficus religiosa). Most chloroplasts encoded 113 unique genes, including a set of 78 protein-coding genes, 30 transfer RNA (tRNA) genes, four ribosomal RNA (rRNA) genes, and one pseudogene (infA). The number of simple sequence repeats (SSRs) ranged from 67 (Ficus sagittata) to 89 (Ficus microdictya) and generally increased linearly with plastid size. Among the plastomes, comparative analysis revealed eight intergenic spacers that were hotspot regions for divergence. Additionally, the clpP, rbcL, and ccsA genes showed evidence of positive selection. Phylogenetic analysis indicated that none of the six traditionally recognized subgenera of Ficus were monophyletic. Divergence time analysis based on the complete chloroplast genome sequences showed that Ficus species diverged rapidly during the early to middle Miocene. This research provides basic resources for further evolutionary studies of Ficus.

Diversity and metabolic potentials of microbial communities associated with pollinator and cheater fig wasps in fig-fig wasp mutualism system.

Microbial symbionts can influence a myriad of insect behavioral and physiological traits. However, how microbial communities may shape or be shaped by insect interactions with plants and neighboring species remains underexplored. The fig-fig wasp mutualism system offers a unique model to study the roles of microbiome in the interactions between the plants and co-habiting insects because a confined fig environment is shared by two fig wasp species, the pollinator wasp (Eupristina altissima and Eupristina verticillata) and the cheater wasp (Eupristina sp1 and Eupristina sp2). Here, we performed whole genome resequencing (WGS) on 48 individual fig wasps (Eupristina spp.) from Yunnan, China, to reveal the phylogenetic relationship and genetic divergence between pollinator and congeneric cheater wasps associated with the Ficus trees. We then extracted metagenomic sequences to explore the compositions, network structures, and functional capabilities of microbial communities associated with these wasps. We found that the cheaters and pollinators from the same fig species are sister species, which are highly genetically divergent. Fig wasps harbor diverse but stable microbial communities. Fig species dominate over the fig wasp genotype in shaping the bacterial and fungal communities. Variation in microbial communities may be partially explained by the filtering effect from fig and phylogeny of fig wasps. It is worth noting that cheaters have similar microbial communities to their sister pollinators, which may allow cheaters to coexist and gain resources from the same fig species. In terms of metabolic capabilities, some bacteria such as Desulfovibrio and Lachnospiraceae are candidates involved in the nutritional uptake of fig wasps. Our results provide novel insights into how microbiome community and metabolic functions may couple with the fig-wasp mutualistic systems.