A new pollination system: brood-site pollination by flower bugs in Macaranga (Euphorbiaceae)

Background and Aims Macaranga: (Euphorbiaceae) is a large genus of dioecious trees with approx.260 species. To date, only one pollination study of the genushas reported brood-site pollination by thrips in M. hullettii.In this study, the pollination system of Macaranga tanariusis reported. Methods: The study was conducted on Okinawa and Amami Islands, Japan.Flower visitors on M. tanarius were collected and their pollenload and behaviour on the flowers examined, as well as inflorescencestructure and reward for the pollinators. Key Results: The most abundant flower visitors found on the male and femaleinflorescences were Orius atratus (Anthocoridae, Hemiptera),followed by Decomioides schneirlai (Miridae, Hemiptera). Pollenload on O. atratus from flowering pistillate inflorescenceswas detected as well as from staminate flowers. Orius atratusand D. schneirlai are likely to use the enclosed chambers formedby floral bracts as breeding sites before and during floweranthesis, and feed on nectar on the adaxial surface of flowerbracts. The extrafloral nectary has a ball-shaped structureand the contained nectar is not exposed; the hemipterans piercethe ball to suck out the nectar. Conclusions: The results indicate that the plant is pollinated by flowerbugs breeding on the inflorescences. This study may be the firstreport of pollination systems in which flower bugs are the mainpollinators. Similarity of pollination systems between M. hullettiiand M. tanarius indicates that the two brood-site pollinationsystems have the same origin. The pollinator species belongsto a predacious group, whose major prey includes thrips. Thepollination system might represent a unique example of evolutionfrom predatory flower visitors feeding on the pollinators (thrips)to the main pollinators.     

A chloroplast genealogy of myrmecophytic Macaranga species (Euphorbiaceae) in Southeast Asia reveals hybridization, vicariance and long-distance dispersals

Macaranga (Euphorbiaceae) includes about 280 species with a palaeotropic distribution. The genus not only comprises some of the most prominent pioneer tree species in Southeast Asian lowland dipterocarp forests, it also exhibits a substantial radiation of ant-plants (myrmecophytes). Obligate ant-plant mutualisms are formed by about 30 Macaranga species and 13 ant species of the genera Crematogaster or Camponotus. To improve our understanding of the co-evolution of the ants and their host plants, we aim at reconstructing comparative organellar phylogeographies of both partners across their distributional range. Preliminary evidence indicated that chloroplast DNA introgression among closely related Macaranga species might occur. We therefore constructed a comprehensive chloroplast genealogy based on DNA sequence data from the noncoding ccmp2, ccmp6, and atpB-rbcL regions for 144 individuals from 41 Macaranga species, covering all major evolutionary lineages within the three sections that contain myrmecophytes. A total of 88 chloroplast haplotypes were identified, and grouped into a statistical parsimony network that clearly distinguished sections and well-defined subsectional groups. Within these groups, the arrangement of haplotypes followed geographical rather than taxonomical criteria. Thus, up to six chloroplast haplotypes were found within single species, and up to seven species shared a single haplotype. The spatial distribution of the chloroplast types revealed several dispersals between the Malay Peninsula and Borneo, and a deep split between Sabah and the remainder of Borneo. Our large-scale chloroplast genealogy highlights the complex history of migration, hybridization, and speciation in the myrmecophytes of the genus Macaranga. It will serve as a guideline for adequate sampling and data interpretation in phylogeographic studies of individual Macaranga species and species groups.     

Slippery ant-plants and skilful climbers: selection and protection of specific ant partners by epicuticular wax blooms in Macaranga (Euphorbiaceae)

 In many ant-plant species of the genus Macaranga in South-East Asia, conspicuous blooms of epicuticular wax crystals cover the stem surface. We found that many ant species were unable to walk on these surfaces. Only the specific ant partners of glaucous Macaranga host plants were capable of moving on the slippery stems without difficulty. Therefore, the epicuticular coatings of Macaranga myrmecophytes appear to have a selective function and protect the associated ants against competitors. The epicuticular aggregates function as a physical barrier; no evidence of chemical repellence was found. The extent to which ”foreign” ant species are excluded from a tree strongly depends on inclination, diameter and length of the glaucous stem sections. The particular growth form of some glaucous Macaranga ant-plants enhances the influence of the wax barriers. The ant associates of glaucous and glossy Macaranga ant-plants (genera Crematogaster and Camponotus) differ strongly in their capacity to adhere to the glaucous stems. For this reason, the wax blooms in Macaranga can act as an ecological isolation mechanism for the sympiotic ants. Within the genus Macaranga, we find a high correspondence between the occurrence of glaucousness and obligatory ant association (50% in ant-plants; 6.7% in non-myrmecophytes). The genus Macaranga thus represents one of the few cases known so far where epicuticular wax crystals are likely to have evolved in relation to insects. Received: 2 January 1997 / Accepted: 9 June 1997     

Comparative chloroplast DNA phylogeography of two tropical pioneer trees, Macaranga gigantea and Macaranga pearsonii (Euphorbiaceae)

Macaranga (Euphorbiaceae) has received much ecological and evolutionary research attention as a genus that includes some of the most conspicuous pioneer trees of Southeast Asian tropical rainforests and because of its manifold associations with ants, including about 30 species that are obligate ant-plants (myrmecophytes). We used sequence data from three chloroplast DNA loci (ccmp5, ccmp6, atpB-rbcL) to assess phylogeographical patterns in species of Macaranga, section Pruinosae, sampled from various regions of Borneo and the Malay Peninsula. Forty-nine chloroplast DNA haplotypes (HT) were identified among 768 specimens from five species, Macaranga gigantea (N = 329; 23 HT), Macaranga pearsonii (N = 347; 21 HT), Macaranga puberula (N = 24; 4 HT), Macaranga hosei (N = 48; 6 HT), and Macaranga pruinosa (N = 20; 5 HT). Forty-one haplotypes were species-specific, whereas eight haplotypes were shared by two, three, or four species and occupied internal positions in a parsimony network. Population genetic parameters based on haplotype frequencies proved to be in a similar range in the non-myrmecophytic M. gigantea and in the ant-associated M. pearsonii, which have overlapping distributions in northern and eastern Borneo. A comparison of G _ST and N _ST values revealed a strong phylogeographic structure in both species, whereas colonization pathways suggested by the network topology were different. Both species exhibited similar levels of haplotypic diversity and moderate to high levels of population differentiation. There were no obvious indications for an influence of the symbiotic ant partners on the population structure of their host plants.     

Isolation and characterization of 11 new microsatellite markers for Macaranga (Euphorbiaceae)

We provide primer sequences for 11 new polymorphic microsatellite markers developed in the tropical ant-plant genus Macaranga (Euphorbiaceae), after enrichment cloning of Macaranga tanarius and Macaranga hypoleuca. Allele numbers per locus ranged from two to 16 among 20 accessions of M. tanarius, and from three to 10 among 22 accessions of M. hypoleuca. Observed and expected heterozygosities ranged from 0.150 to 0.900 and from 0.375 to 0.894 in M. tanarius, and from 0.545 to 1.000 and from 0.434 to 0.870 in M. hypoleuca, respectively. Six of the 11 primer pairs successfully cross-amplified polymorphic polymerase chain reaction products in Macaranga winkleri.     

The significance of ant and plant traits for ant pollination in Leporella fimbriata

Summary Ant metapleural glands secrete surface antibiotics that affect pollen as well as bacteria and fungi. This may be one reason why ant pollination is rare. It is predicted that pollination by ants is possible only in the presence of certain ant and/or plant traits. Two traits are investigated; first, absence of the metapleural glands, and second, the presence of stigmatic secretions that insulate pollen from the ant integument. The pollinator of the orchid Leporella fimbriata is the ant Myrmecia urens. Only one caste is involved, the winged males, and they differ significantly from the queen and worker castes in that they do not possess metapleural glands. This paper reports experiments which test for differential effects on pollen between the males and other castes and evaluates the importance of stigmatic secretions. The results show that the absence of metapleural glands makes no difference as all three castes have strong disruptive effect on pollen artificially applied to the integument. However, during pollination the orchid secures the pollen mass to the ant surface by stigmatic secretions and normal pollen function, fruit production and seed set occur. It appears that both ant and plant traits are pre-adaptive having evolved for functions other than ant pollination.This is contribution number 98 to the Research Unit for Biodiversity and Bioresources, Macquarie University     

Food bodies and their significance for obligate ant-association in the tree genus Macaranga (Euphorbiaceae)

FIALA, B. & MASCHWITZ, U., 1992. Food bodies and their significance for obligate ant-association in the tree genus Macaranga (Euphorbiaceae). The production of extrafloral nectar and food bodies plays an important role in many tropical ant-plant mutualisms. In Malaysia, a close association exists between ants and some species of the pioneer tree genus Macaranga (Euphorbiaceae). Macaranga is a very diverse genus which exhibits all stages of interaction with ants, from facultative to obligatory associations. The ants nest inside the hollow internodes and feed mainly on food bodies provided by the plants. Food body production had previously been reported only in myrmecophytic Macaranga species, where it is usually concentrated on protected parts of the plants such as recurved stipules. We found that non-myrmecophytic Macaranga species also produce food bodies on leaves and stems, where they are collected by a variety of ants. Levels of food body production differ between facultatively and obligatorily ant-associated species but also among the various non-myrmecophytes. This may be related to the degree of interaction with ants. Food body production starts at a younger age in the myrmecophytic species than in the transitional or non-myrmeccophytic Macaranga. Although food bodies of the non-inhabited Macaranga species are collected by a variety of ants, there is no evidence of association with specific ant species. Our observations suggest that food bodies enhance the evolution of ant-plant interactions. Production of food bodies alone, however, does not appear to be the most important factor for the development of obligate myrmecophytism in Macaranga.     

Development and characterization of chloroplast microsatellite markers in Macaranga (Euphorbiaceae).

As part of our study on the phylogeography of the ant-plant genus Macaranga, we have screened for polymorphic regions in the chloroplast genome. Initially, ten universal PCR primer pairs targeted at chloroplast microsatellite loci were applied to a small set of specimens, covering various taxonomic levels from intrafamilial to intraspecific. Eight primer pairs produced PCR fragments that behaved as single and discrete bands on agarose gels. The five most promising candidate pairs were further analysed with an extended set of DNA templates, and PCR products were separated on sequencing gels. The number of size variants per locus varied from two to eight, combining into 17 haplotypes among 29 Macaranga accessions from 10 species. Comparative sequencing demonstrated that microsatellites were responsible for the observed size variation at three of five loci, whereas variation at the other loci was caused by larger insertions and (or) deletions (indels). In addition to poly(A) and poly(T) repeats, which are typically found in chloroplast DNA, we also identified a variable (CT)n repeat, with n = 4 to n = 8. Sequencing revealed three examples of size homoplasy, one of which was caused by a single base substitution that raised the actual number of haplotypes to 18. Relationships between haplotypes were assessed by phenetic analyses of size variants and by constructing a parsimony network based on sequence variation. For both types of analysis, the distribution of haplotypes correlated with geographically circumscribed regions rather than with taxonomic boundaries.     

山地五月茶的蝇类传粉研究

观测了山地五月茶Antidesma montanum的开花物候、开花动态、访花者种类和访花行为,并对其繁育系统、花粉组织化学、花粉胚珠比（P／O）、花粉活力进行了检测。结果表明：山地五月茶是雌雄异株植物;其总状花序花期可长达7天;雄花／雌花花序数目为140．33±27．79／208．33±33．65（n=6）,雌雄花的颜色为很淡的黄绿色小型花;雄花单花花期为2天,花药开裂当天有活力,花粉为非淀粉型,花粉胚珠比为3333．33±607．18;单花花蜜量可达0．34±0．03μL,花蜜含糖量3．69％±0．30％。花的结构和开花式样适合蝇类传粉。芳香的气味是吸引蝇类的直接物质：花蜜是传粉者的报酬。主要的传粉昆虫为双翅目Diptera丽蝇科Calliphoridae的Chrysomya megacephala、Chrysomyasp．、寄蝇科Tachinidae的Drino sp．和蝇科Muscidae的Spilogona sp．、Mitroplatia sp．。套网不授粉的处理不结实,表明山地五月茶不存在无融合生殖,人工辅助授粉的坐果率（39．1％）略高于自然坐果率（36．7％）,二者无显著差异,表明其坐果率主要受自身资源分配限制。还讨论了蝇类传粉与雌雄异株性系统以及蝇类传粉与热带林中小型黄绿色花植物的相关性。     

A revision of Ptychopyxis (Euphorbiaceae) in southeast Asia

Ptychopyxis is a genus comprised of 11 species. Nine are found in southeast Asia mainland and west Malesia and two species are endemic to New Guinea. One of the endemic New Guinean species is newly described here, viz. P. esdras Stoops sp. nov.     

Pedilanthus diazlunanus (Euphorbiaceae): pollination by hymenopterans in a bird-pollinated genus

Pollination in the genus Pedilanthus is commonly effected by hummingbirds. Pollination by vespid wasps in Pedilanthus is documented for the first time based on observations of Pachodynerus nasidens and Eumenes americanus pollinating Pedilanthus diazlunanus. An hypothesis concerning the mechanism by which hummingbird pollination was replaced by insect pollination in Pedilanthus is advanced based on observations of insects on P. bracteatus.     

Male-biased sex ratio and promiscuous pollination in the dioecious island treeLaurus azorica (Lauraceae)

Pollination ofLaurus azorica (Lauraceae), a dioecious Macaronesian tree, was studied. Male and female trees had the same size distribution. The population had 2.5 times as many male trees as females. In addition, males produced more flowers, and their inflorescences lasted longer. Individual flower lifetime and length of flowering season were the same in both sexes. Between the years of observation, one tree changed sex. Pollinators wereHalictinae bees and the flyTachina canariensis. The bees collected pollen and nectar and the fly collected nectar from both sexes. Both species visited other plants as well. The evolution of breeding systems inLauraceae is discussed.     

Thrips pollination of androdioecious Castilla elastica (Moraceae) in a seasonal tropical forest

Androdioecy is a rare sexual system in nature, as predicted theoretically. Among the androecious species reported so far, Castilla elastica (Moraceae) is unique in that flowers are unisexual and staminate and pistillate flowers on cosexual plants are produced on different inflorescences. In addition, inflorescence structure of staminate inflorescences on males and staminate and pistillate inflorescences on cosexes is markedly different. Staminate inflorescences on males are bivalvate, while staminate inflorescences on cosexes are "fig-like" and urceolate. Pistillate inflorescences are discoidal. The difference may reflect different roles and requirements of the three inflorescences in pollination and protection from herbivores. This study reports thrips pollination of C. elastica, demonstrated by a pollinator introduction experiment. Thrips pollination of the species may be an example of mutualism originating from plant-herbivore interactions. In the Moraceae, shifts from simple herbivores on flowers to pollinators might have occurred many times, evolving into diverse pollination systems including the fig-fig wasp mutualism. The family, of which little is known about pollination systems, provides interesting and unique opportunities to study evolution of pollination systems and roles of nonpollinating associates of inflorescences.     

AFLP analysis of phylogenetic relationships among myrmecophytic species of Macaranga (Euphorbiaceae) and their allies

The palaeotropic pioneer tree genus Macaranga Thouars (Euphorbiaceae) is characterized by various types of mutualistic interactions with specific ant partners (mainly Crematogaster spp.). About 30 species are obligate ant-plants (myrmecophytes). We used amplified fragment length polymorphism (AFLP) markers to assess phylogenetic relationships among 108 Macaranga specimens from 43 species, including all available taxa from the three sections known to contain myrmecophytes. Eight primer combinations produced 426 bands that were scored as presence/absence characters. Banding patterns were analyzed phenetically, cladistically and by principal coordinates analysis. Monophyly of section Pruinosae is clearly supported. There is also good evidence for a monophyletic section Pachystemon that includes the puncticulata group. The monophyly of section Winklerianae and relationships between the three sections remain ambiguous. Section Pachystemon is subdivided into four well-supported monophyletic subclades that presumably correspond to taxonomic entities.We acknowledge the support by the Deutsche Forschungsgemeinschaft (DFG Fi606/4-1, DFG We1830/2-1, 4-1 and 4-2), which in part was granted in the frame of the DFG-SPP 1127 Radiations: origins of biological diversity. Part of the plant material was kindly supplied by Dr. H. Feldhaar (University of Würzburg), Dr. U. Moog (University of Kassel) and Dr. F. Slik (Leiden University Branch, Nationaal Herbarium Nederland). We thank the University of Malaysia (Dr. Rosli b. Hashim) and Taman Taman Sabah (Datuk Lamri Ali; Dr. J. Nais) for permits and logistic support, and EPU for permission to conduct research in Malaysia.     

Molecular phylogeny of Crematogaster subgenus Decacrema ants (Hymenoptera: Formicidae) and the colonization of Macaranga (Euphorbiaceae) trees

To elucidate the evolution of one of the most species-rich ant-plant symbiotic systems, the association between Crematogaster (Myrmicinae) and Macaranga (Euphorbiaceae) in South-East Asia, we conducted a phylogenetic analysis of the ant partners. For the phylogenetic analysis partial mitochondrial cytochrome oxidase I and II were sequenced and Maximum Parsimony analysis was performed. The analyzed Crematogaster of the subgenus Decacrema fell into three distinct clades which are also characterized by specific morphological and ecological traits (queen morphology, host-plants, and colony structure). Our results supported the validity of our currently used morphospecies concept for Peninsula Malaysia. However, on a wider geographic range (including North and North-East Borneo) some morphospecies turned out to be species complexes with genetically quite distinct taxa. Our phylogenetic analysis and host association studies do not indicate strict cocladogenesis between the subgenus Decacrema and their Macaranga host-plants because multiple ant taxa occur on quite distinct host-plants belonging to different clades within in the genus Macaranga. These results support the view that host-shifting or host-expansion is common in the ants colonizing Macaranga. Additionally, the considerable geographic substructuring found in the phylogenetic trees of the ants suggests that allopatric speciation has also played a role in the diversification and the current distribution of the Decacrema ants.     

Microsatellite markers for the palaeotropic pioneer tree genus Macaranga (Euphorbiaceae) and their cross‐species transferability

We developed primer sequences for five polymorphic microsatellite loci in the tropical ant‐plant genus Macaranga (Euphorbiaceae). Population genetic parameters were determined on the basis of 30 individuals from each of two Macaranga species in Borneo. Allele numbers per locus ranged from three to 13. Expected and observed heterozygosities ranged from 0.160 to 0.850 and from 0.130 to 0.700, respectively. Four of the five primer pairs cross‐amplify polymorphic PCR products in a wide range of Macaranga species.     

Biased sex ratios in the dioecious annual Croton texensis (Euphorbiaceae) are not due to environmental sex determination

At Arapaho Prairie, in the sandhills of western Nebraska, the dioecious annual Croton texensis (Euphorbiaceae) exhibits biased sex ratios. Moreover, the direction of bias changes from year to year: in 1994 the study population was significantly female biased, in 1995 and 1996 it was significantly male biased, and in 1997 and 1998 the sex ratio did not differ from 1 : 1. Such variation in the observed sex ratio in plants is frequently attributed to environmental sex determination (ESD), which is favored by natural selection if the rate of fitness gain across an environmental gradient is greater for one sex than the other. We performed experiments to determine: (1) whether variation in the sex ratio is correlated with environmental conditions, as would be expected if ESD is operating, and (2) whether ESD, if present, would be favored by natural selection. In a common garden experiment in which water and fertilizer were manipulated the sex ratio was marginally male biased in treatments in which water was added, but not different from 1 : 1 in other treatments. In field plots into which seeds were planted none of several soil characteristics, nor overall plot quality for C. texensis (measured as average plant biomass) were correlated with plot sex ratio. However, plots in which a large number of planted seeds emerged tended to be female biased. These results provide very weak evidence for sex ratio bias across an environmental gradient, and thus provide little evidence for ESD. Moreover, sex-by-environment interactions for fitness, which are required for the evolution of ESD, were absent for all measured variables. Thus, ESD does not appear to be favored by natural selection in this population. Instead, these biases may have been caused by differences between the sexes in germination and/or early mortality.     

Reproductive ecology of dioecious Siparuna (Monimiaceae) in Ecuador–a case of gall midge pollination

The reproductive ecology of the dioecious genus Siparuna , belonging to the primitive family Monimiaceae, was investigated in Ecuador. Thirteen species were studied, and species growing at high altitudes (above 500 m) were compared with species growing at lower altitudes. The genus has specialized flowers pollinated by Cecidomyiidae (Asynapta sp. and other genera) that lay eggs chiefly in male flowers since these are more easily accessible than female flowers due to their morphology. Several traits ensure that pollen transfer is accomplished: 1. long flowering of individuals and single flowers; 2. specific floral morphology (unisexuality, landing platform, restricted pore) allowing the flowers to be visited by Cecidomyiidae seeking egg-laying sites, and 3. a low number of ovules that have to be pollinated in each flower. The low number of ovules per flower results in a high pollen-ovule (P/O) ratio even though pollen production in male flowers is only moderate.
High altitude species had larger flowers, higher pollen-ovule ratios, fewer ovules per female flower, and longer periods of anthesis. Gender differences were seen in the longer functionality of female flowers; the larger numbers of male flowers produced; and the average diameter at breast height (DBH) biased for one of the two sexes depending on the species investigated, with no clear trend in the size distributions of individuals of both sexes. Sex ratios departed significantly from unity both to male and female bias but usually were in favour of males. There were no general differences in the ratio of functional male to female flowers at the time of observation nor in the sizes of male and female flowers. The evolution of unisexual flowers in Siparuna can be explained as a result of the differential predation by larvae: unimportant in male flowers, destructive if occurring in female flowers.