Variation in inflorescence size in a dioecious fig tree and its consequences for the plant and its pollinator fig wasp

The host-specific relationship between fig trees (Ficus) and their pollinator wasps (Agaonidae) is a classic case of obligate mutualism. Pollinators reproduce within highly specialised inflorescences (figs) of fig trees that depend on the pollinator offspring for the dispersal of their pollen. About half of all fig trees are functionally dioecious, with separate male and female plants responsible for separate sexual functions. Pollen and the fig wasps that disperse it are produced within male figs, whereas female figs produce only seeds. Figs vary greatly in size between different species, with female flower numbers varying from tens to many thousands. Within species, the number of female flowers present in each fig is potentially a major determinant of the numbers of pollinator offspring and seeds produced. We recorded variation in female flower numbers within male and female figs of the dioecious Ficus montana growing under controlled conditions, and assessed the sources and consequences of inflorescence size variation for the reproductive success of the plants and their pollinator (Kradibia tentacularis). Female flower numbers varied greatly within and between plants, as did the reproductive success of the plants, and their pollinators. The numbers of pollinator offspring in male figs and seeds in female figs were positively correlated with female flower numbers, but the numbers of male flowers and a parasitoid of the pollinator were not. The significant variation in flower number among figs produced by different individuals growing under uniform conditions indicates that there is a genetic influence on inflorescence size and that this character may be subject to selection.     

Only pollinator fig wasps have males that collaborate to release their females from figs of an Asian fig tree

Male insects rarely collaborate with each other, but pollinator fig wasps (Hymenoptera: Agaonidae) are said to be an exception. Immature fig wasps feed on galled ovules located inside figs, the inflorescences of Ficus species (Moraceae). After mating, adult pollinator males chew communal exit-holes that allow mated females (which are often also their siblings) to escape. Figs also support non-pollinating fig wasps (NPFWs), some of which produce exit-holes independently. We determined whether collaboration between pollinator males (Kradibia tentacularis from Ficus montana) was necessary for the release of their females, and used the relationship between male numbers and likelihood of success to measure the extent of cooperation during exit-hole production. These attributes were then compared with those of an NPFW (Sycoscapter sp.) from the same host plant. Pollinators were more abundant than NPFW, but their more female-biased sex ratio meant male pollinator densities were only slightly higher. Individual males of both species could produce an exit-hole. Single males of the NPFW were just as successful as single male pollinators, but only male pollinators cooperated effectively, becoming more successful as their numbers increased. The lack of cooperation among NPFW may be linked to their earlier period of intense inter-male aggression.     

苹果榕传粉小蜂雌性触角感器及其生态学意义

利用扫描电子显微镜对苹果榕传粉小蜂Ceratosolen sp.雌性触角感器进行观察。结果显示:苹果榕传粉小蜂雌性触角呈膝形,分布着7类13种类型的感器,包括毛型感器、多孔的板型感器、锥型感器(2种类型)、栓锥型乳突状感器、刺型感器(4种类型)、腔锥型感器(3种类型)和一种专一性的角锥型感器。对各感器的形态、数量、分布进行描述,并结合感器的选择性染色(Ag+染色)、传粉小蜂行为及其榕/蜂互惠共生系统,对不同类型感器的功能和生态学意义进行探讨。     

Laticifer distribution in fig inflorescence and its potential role in the fig-fig wasp mutualism

Although in Moraceae the presence of laticifers is considered to be a synapomorphy, little is known about the distribution and morphology of this type of secretory structure in the reproductive organs of its species. Ficus, the largest genus of Moraceae, is characterized by an inflorescence known as syconium and by an obligate mutualistic interaction with pollinating wasps. The objectives of the present study were to evaluate the distribution and morphology of laticifers in syconia of 36 species belonging to different Ficus sections and to survey traits of taxonomic and adaptive value for the group. Syconia containing flowers in a receptive state were collected, fixed and processed for anatomical analysis. All species studied have branched laticifers distributed in the syconium receptacle, in the ostiolar bracts and in the pedicel of staminate flowers. Almost all species show laticifers in the pedicel of shorter-styled flowers. Laticifers also occur in the pedicel of longer-styled flowers in most Ficus sections, except F. curtipes (Conosycea section) and more than 75% of the studied species of the Americanae section. Laticifers are observed in the sepals of 25 of the 36 species studied and occasionally in the pistil. The presence of laticifers in the pedicel of shorter-style flowers and its absence in the pistil suggest that the distribution of this secretory structure in the fig flower was selected by pressures imposed by the fig-fig wasp mutualism. The laticifers in the pedicel of shorter-styled flowers may confer protection to the developing wasp larvae against natural enemies. However, the absence of laticifers in the pistil of most Ficus species studied was probably selected by the mutualistic relationship with the agaonid pollinating wasps since the latex could interfere with oviposition through the style, with the larval development of the pollinating fig wasps, and the emergence of pollinator offspring from their galls.     

Complementary fruiting phenologies facilitate sharing of one pollinator fig wasp by two fig trees

Aims Most pollinator fig wasps are host plant specific, with each species only breeding in the figs of one fig tree species, but increasing numbers of species are known to be pollinated by more than one fig wasp, and in rare instances host switching can result in Ficus species sharing pollinators. In this study, we examined factors facilitating observed host switching at Xishuangbanna in Southwestern (SW) China, where Ficus squamosa is at the northern edge of its range and lacks the fig wasps that pollinate it elsewhere, and its figs are colonized by a Ceratosolen pollinator that routinely breeds in figs of F. heterostyla .Methods We recorded the habitat preferences of F. squamosa and F. heterostyla at Xishuangbanna, and compared characteristics such as fig size, location and colour at receptive phase. Furthermore, the vegetative and reproductive phenologies in the populations of F. squamosa and F. heterostyla were recorded weekly at Xishuangbanna Tropical Botanical Garden for 1 year.Important findings Ficus squamosa is a shrub found near fast-flowing rivers, F. heterostyla is a small tree of disturbed forest edges. Although preferring different habitats, they can be found growing close together. Both species have figs located at or near ground level, but they differ in size when pollinated. Fig production in F. squamosa was concentrated in the colder months. F. heterostyla produced more figs in summer but had some throughout the year. The absence of its normal pollinators, in combination with similarly located figs and partially complementary fruiting patterns appear to have facilitated colonization of F. squamosa by the routine pollinator of F. heterostyla. The figs probably also share similar attractant volatiles. This host switching suggests one mechanism whereby fig trees can acquire new pollinators and emphasizes the likely significance of edges of ranges in the genesis of novel fig tree–fig wasp relationships.     

Direct evidence for the cycling of fig wasps within one male fig tree

Fig wasps can only survive when flowering fig trees are present all the year around. Ficus trees can only reproduce if they are pollinated by highly specific wasps. In highly seasonal habitats, when only few trees occur at a specific site, gaps in fruiting may lead to the extinction of the local pollinator population. This paper demonstrates that in a dioecious fig tree, Ficus hirta , the fig wasp population can be maintained successfully within an individual plant, through the strong intra-tree asynchrony in flowering. By experimentally bagging trees, we showed that the pollinating wasps ( Blastophaga javana hilli ) could live for two generations, and the non-pollinating wasps ( Sycoscapter sp.) for up to three generations in a closed intra-tree system. However, there was a sharp decline in wasp abundance, deviating sex ratios and decreasing flower occupancy before their ultimate extinction, indicating that the wasp populations were not sustainable. This phenological strategy may enable dioecious figs, which are not constrained by the cost of selfing, to occupy a wider breadth of niches in both tropical and seasonal habitats.     

How an orchid harms its pollinator

Certain orchids produce flowers that mimic the sex pheromones and appearance of female insects in order to attract males by sexual deception for the purpose of pollination. In a series of field experiments, we found that the sexually deceptive orchid, Chiloglottis trapeziformis, can have a negative impact on its wasp pollinator Neozeleboria cryptoides. Male and female wasps, however, were affected differently by the orchid's deceit because of their different roles in the mimicry system. Male wasps could not discriminate between the chemical cues of orchids and female wasps, a vital signal in long-range attraction. Males, however, learn to avoid areas containing orchids. This strategy has implications for females attempting to attract mates in areas occupied by orchids. Compared with circumstances when females were on their own, females in the presence of orchids elicited fewer male approaches and no copulation attempts. Females in a large orchid patch also elicited fewer male approaches than females in a small patch. The nature of the orchid's impact on its wasp pollinator indicates an arms race evolutionary scenario in this interaction between plant and pollinator.     

A comparison of pollinator fig wasp development in figs of Ficus montana and its hybrids with Ficus asperifolia

Figs (Moraceae) and pollinator fig wasps (Hymenoptera: Agaonidae) have a highly specific mutualistic relationship but fig wasps occasionally enter atypical hosts, and this can lead to hybrid fig trees and the potential for gene flow between species. Many fig trees are dioecious, with fig wasp offspring developing in galled ovules inside figs on male trees, whereas seeds develop only in figs on female trees. We generated experimental hybrids between the Asian Ficus montana Blume and a closely related African species Ficus asperifolia Miquel. Male F1s were sterile if entered by Kradibia tentacularis (Grandi) (Agaonidae), the pollinator of F. montana, because its offspring always failed to develop, without ovule enlargement. As with the F1s, figs on most male backcross plants [F. montana × (F. montana × F. asperifolia)] also aborted shortly after pollinator entry, resulting in a higher turnover of figs than with F. montana, although the times taken for the figs to reach receptivity were similar. Pollinator larvae nonetheless consistently managed to develop inside the figs of one backcross plant and also occasionally in a few figs from another backcross individual. In these figs, galled ovules developed as normal, whereas in figs that aborted the galled ovules failed to enlarge. The sex ratio of K. tentacularis progeny in the backcross figs was female biased and did not differ from that in F. montana figs. Sycoscapter spec. (Hymenoptera: Pteromalidae), a parasitoid of K. tentacularis, was able to lay eggs and developed normally inside male backcross figs where its host was present.     

Interactions between pollinator and non‐pollinator fig wasps: correlations between their numbers can be misleading

Ficus and their species–specific pollinator fig wasps represent an obligate plant–insect mutualism, but figs also support a community of non‐pollinating fig wasps (NPFWs) that consist of phytophages and parasitoids or inquilines. We studied interactions between Kradibia tentacularis, the pollinator of a dioecious fig tree species Ficus montana, and an undescribed NPFW Sycoscapter sp. Members of Sycoscapter sp. oviposited 2–4 weeks after pollinator oviposition, when host larvae were present in the figs. No negative correlation was found between the numbers of the two wasp species emerging from figs in a semi‐natural population. However, in experiments where the numbers of pollinator foundresses entering a fig were controlled, Sycoscapter sp. significantly reduced the numbers of pollinator offspring. Consequently, it can be concluded that Sycoscapter sp. is a parasitoid of K. tentacularis (which may also feed on plant tissue). Sycoscapter females concentrate their oviposition in figs that contain more potential hosts, rendering invalid conclusions based on simple correlations of host and natural enemy numbers.     

Coevolution of reproductive characteristics in three dioecious fig species and their pollinator wasps

This study investigates dioecious fig species using a pollinator introduction experiment. Our aims were to determine: (1) whether there was a significant difference in foundress distribution between sexes per fig species; (2) whether fig size and foundress number affect reproductive success of dioecious figs; and (3) who is the ‘controlling partner’ in the fig/pollinator mutualism. Three dioecious fig species: Ficus semicordata, Ficus hispida and Ficus tinctoria from Xishuangbanna, China, were selected for this experiment. We found that there was no significant difference of the foundress number in female and male figs of F. semicordata, F. hispida and F. tinctoria. Also, the foundress number did not depend on the fig diameter. The numbers and the proportions of fig seeds and female wasp offspring significantly increased with more foundresses; and fig seed number was significantly higher than female wasp offspring in F. semicordata and F. hispida, but not in F. tinctoria. Our results indicate that figs are generally the ‘controlling partner’ in fig-wasp mutualisms in species with large figs, but not with small figs. Compared with published studies of reproductive success in monoecious figs, the dioecious figs seem to be more efficient in producing both seeds and wasp offspring when there is a high number of foundress.     

Costs of an induced immune response on sexual display and longevity in field crickets

Immune system activation may benefit hosts by generating resistance to parasites. However, natural resources are usually limited, causing a trade-off between the investment in immunity and that in other life-history or sexually selected traits. Despite its importance for the evolution of host defense, state-dependent fitness costs of immunity received little attention under natural conditions. In a field experiment we manipulated the nutritional condition of male field crickets Gryllus campestris and subsequently investigated the effect of an induced immune response through inoculation of bacterial lipopolysaccharides. Immune system activation caused a condition-dependent reduction in body condition, which was proportional to the condition-gain during the preceding food-supplementation period. Independent of nutritional condition, the immune insult induced an enduring reduction in daily calling rate, whereas control-injected males fully regained their baseline level of sexual signaling following a temporary decline. Since daily calling rate affects female mate choice under natural conditions, this suggests a decline in male mating success as a cost of induced immunity. Food supplementation enhanced male life span, whereas the immune insult reduced longevity, independent of nutritional status. Thus, immune system activation ultimately curtails male fitness due to a combined decline in sexual display and life span. Our field study thus indicates a key role for fitness costs of induced immunity in the evolution of host defense. In particular, costs expressed in sexually selected traits might warrant the honest advertisement of male health status, thus representing an important mechanism in parasite-mediated sexual selection.     

Selection to attract pollinators and to confuse antagonists specializes fig–pollinator chemical communications

Chemical communication is critical in establishing angiosperm–pollinator mutualisms. However, our understanding of how chemical communication shapes coevolution remains limited. Here, we integrated information theory to model three coevolutionary scenarios (I‒III), where the pollinator fitness is always optimized by the highest certainty of chemical information provided by plants, but plant fitness is determined by (I) the certainty of chemical information attracting pollinators, (II) the uncertainty of chemical information confusing antagonists, or (III) both aspects. We found that the statistical properties of empirical plant volatiles from 45 pairs of fig–pollinator mutualisms were best explained by the selection from both pollinators and antagonists (scenario III). Under this scenario, plant–pollinator mutualisms evolve to be specialized and as few as two volatile chemicals could supply sufficient information for pollinators’ host identification. Our study provides new insights into plant–pollinator coevolution and will facilitate further studies on the evolution and diversification in specialized plant–pollinator–herbivore systems.     

Distribution of nuclear mitochondrial pseudogenes in three pollinator fig wasps associated with Ficus pumila

Nuclear mitochondrial pseudogenes (NUMTs) are nuclear sequences transferred from mitochondrial genomes. Although widespread, their distribution patterns among populations or closely related species are rarely documented. We amplified and sequenced the mitochondrial cytochrome b (Cytb) gene to check for NUMTs in three fig wasp species that pollinate Ficus pumila (Wiebesia sp. 1, 2 and 3) in Southeastern China using direct and cloned sequencing. Unambiguous sequences (332) of 487 bp in length belonging to 33 haplotypes were found by direct sequencing. Their distribution was highly concordant with those of cytochrome c oxidase subunit I (COI). Obvious signs of co-amplification of NUMTs were indicated by their uneven distribution. NUMTs were observed in all individuals of 12 populations of Wiebesia sp. 3, and 13 individuals of three northern populations of Wiebesia sp. 1. Sequencing clones of potential co-amplification products confirmed that they were NUMTs. These NUMTs either clustered as NUMT clades basal to mtDNA Cytb clades (basal NUMTs), or together with Cytb haplotypes. Basal NUMTs had either stop codons or frame-shifting mutations resulting from deletion of a 106 bp fragment. In addition, no third codon or synonymous substitutions were detected within each NUMT clade. The phylogenetic tree indicated that basal NUMTs had been inserted into nuclei before divergence of the three species. No significant pairwise differences were detected in their ratios of third codon substitutions, suggesting that these NUMTs originated from one transfer event, with duplication in the nuclear genome resulting in the coexistence of the 381 bp copy. No significant substitution differences were detected between Cytb haplotypes and NUMTs that clustered with Cytb haplotypes. However, these NUMTs coexisted with Cytb haplotypes in multiple populations, suggesting that these NUMT haplotypes were recently inserted into the nuclear genome. Both basal and recently inserted NUMTs were rare events, and were absent in most populations of Wiebesia sp. 1 and 2. Further studies are needed to distinguish between mechanisms potentially generating this rarity, such as purifying selection, genetic drift or amplification failure.     

Genetic characterization of fig tree mutants with molecular markers

The fig (Ficus carica L.) is a fruit tree of great world importance and, therefore, the genetic improvement becomes an important field of research for better crops, being necessary to gather information on this species, mainly regarding its genetic variability so that appropriate propagation projects and management are made. The improvement programs of fig trees using conventional procedures in order to obtain new cultivars are rare in many countries, such as Brazil, especially due to the little genetic variability and to the difficulties in obtaining plants from gamete fusion once the wasp Blastophaga psenes, responsible for the natural pollinating, is not found in Brazil. In this way, the mutagenic genetic improvement becomes a solution of it. For this reason, in an experiment conducted earlier, fig plants formed by cuttings treated with gamma ray were selected based on their agronomic characteristics of interest. We determined the genetic variability in these fig tree selections, using RAPD and AFLP molecular markers, comparing them to each other and to the Roxo-de-Valinhos, used as the standard. For the reactions of DNA amplification, 140 RAPD primers and 12 primer combinations for AFLP analysis were used. The selections did not differ genetically between themselves and between them and the Roxo-de-Valinhos cultivar. Techniques that can detect polymorphism between treatments, such as DNA sequencing, must be tested. The phenotypic variation of plants may be due to epigenetic variation, necessitating the use of techniques with methylation-sensitive restriction enzymes.     

Body size in a pollinating fig wasp and implications for stability in a fig‐pollinator mutualism

The fig–fig pollinator association is a classic case of an obligate mutualism. Fig‐pollinating wasps often have to fly long distances from their natal syconia to a receptive syconium and then must enter the narrow ostiole of the syconium to reproduce. Large wasps are expected to have a greater chance of reaching a receptive syconium. In this study, we tested this hypothesis and then examined whether the ostiole selectively prevented larger pollinators from entering the syconial cavity. In Xishuangbanna, China, Ceratosolen solmsi marchali Mayr (Hymenoptera: Agaonidae) pollinates the dioecious syconia of Ficus hispida L. (Moraceae). The body size of newly emerged wasps and wasps arriving at receptive syconia were compared. Wasps arriving at receptive syconia were significantly larger than newly emerged wasps. We also compared the size of wasps trapped in the ostiole with those in the cavity. Wasps trapped in the ostiole were significantly larger than those in the syconial cavity. Thus, in the case of F. hispida, large wasps were more likely to reach receptive syconia, but the ostiole limited maximum fig wasp size. This indicates that the ostiole, as a selective filter to pollinators, stabilizes pollinator size. Hence, it helps to maintain stability in the fig–fig pollinator mutualism.     

The non-pollinating fig wasps associated with Ficus guianensis: Community structure and impact of the large species on the fig/pollinator mutualism

Understanding the ecology of non-pollinating fig-wasp communities depends on a good knowledge of larval feeding habits of the species involved, which can be gall inducers, kleptoparasites, parasitoids or seed eaters. However, larval feeding habits are poorly known and most community ecology studies on NPFW are based on hypothetical feeding habits or data analyzed independently of feeding habit. Here we take advantage of the particular situation in Ficus guianensis whose community is dominated by large NPFW, i.e. species that are obviously larger than pollinators, to establish the community structure and feeding habits of the most frequent wasps. We provide the first non-ambiguous negative correlation between the number of NPFW and the production of pollinators and seeds. Each developing large NPFW represents a disproportionate cost to the mutualism as it is responsible for the loss of about ten seeds plus pollinators, i.e. about 10% of the production of a fig.     

Development and characterization of microsatellite markers for Liporrhopalum tentacularis Grandi,the pollinator fig wasp of Ficus montana Blume

Microsatellite markers for the pollinator fig wasp Liporrhopalum tentacularis were developed using genomic libraries enriched for di‐, tri‐ and tetranucleotide repeats. A subset of 31 positive clones was sequenced and primers were designed. Eleven primer pairs produced polymorphic amplification products in L. tentacularis. Eight markers gave unambiguously scorable patterns and were further characterized on 29 individuals collected from different fruits of the dioecious host fig Ficus montana in Indonesia. Three to 19 alleles per locus were detected in this set of samples. The observed heterozygosity ranged between 0.10 and 0.55.