Adverse effects on pollen exposed to Atta texana and other North American ants: implications for ant pollination

Summary The effects of the surface secretions of eight species of ants on three types of pollen were bioassayed by exposure to the integument of undisturbed, living individuals for 20 min. Ant species included Atta texana which cultures fungi by means of various types of secretions. The frequency of grains showing membrane dysfunction, and therefore reduced viability, was quantified by means of a fluorochromatic test. Comparisons of treated and control samples showed that in 46 out of 50 bioassays there was a reduction in pollen viability following exposure to ants, 38 being statistically significant. Variation in the outcome of bioassays showed differential potency among ant species and differential vulnerability among pollen types. Ant pollination may be uncommon because surface secretions, often from the metapleural glands, cause membrane dysfunction in pollen. Ant species without metapleural glands may be pollinators, but ant pollinated plants may have pollen resistant to the secretion.     

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     

Pseudocopulation of an orchid by male ants: a test of two hypotheses accounting for the rarity of ant pollination

Summary The orchid Leporella fimbriata is pollinated by pseudocopulation with winged males of the ant Myrmecia urens. This recently studied interaction provides a unique opportunity to examine the two current hypotheses concerning the apparent rarity of ant pollination systems worldwide. The first hypothesis requires a series of specialized growth forms and floral characteristics regarded as adaptations to ant pollination. L. fimbriata does not possess them. The second considers the pollenicidal effects of secretions from the metapleural gland of ants. These glands are absent in M. urens males and it may be that the occurrence of ant pollination requires the absence of metapleural glands in the vector.     

Weaving resistance: silk and disease resistance in the weaver ant <Emphasis Type="Italic">Polyrhachis dives</Emphasis>

Social insects are at risk from a diverse range of parasites. The antibiotic-producing metapleural gland is an ancestral trait in ants which is thought to be one of their primary mechanisms of resistance. However, the metapleural gland has been lost secondarily in three ant genera, which include weaver ants that are characterised by the remarkable construction of their nests using larval silk. Silken nests may have allowed reduced investment in costly disease resistance mechanisms like the metapleural gland if the silk has antimicrobial properties, as in other insects, or is a hygienic substrate. Here we examine this hypothesis in the weaver ant Polyrhachis dives. We found no evidence of a beneficial effect of silk. The presence of silk did not improve the already high resistance of ants to the entomopathogenic fungus Metarhizium, the ants only rarely interacted with the silk regardless of whether they were exposed to Metarhizium or not, and silk also did not inhibit the in vitro germination or growth of Metarhizium. Furthermore, silk was found in vitro to be heavily contaminated with the facultative entomopathogenic fungus Aspergillus flavus, and many more ants sporulated with this fungus when kept with silk in vivo than when they were kept without silk. Further work is needed to examine the effects of silk on other parasites and of silk from other weaver ants. However, the results in combination suggest that silk in P. dives is unlikely to provide protection against parasites and that it is also not a hygienic substrate. Alternative explanations may therefore be needed for the loss of the metapleural gland in weaver ants.     

WIND POLLINATION OF TAXUS CUSPIDATA

The air disturbance patterns created by and around the ovules of Taxus cuspidata are quantified for various orientations to the direction of ambient airflow, and are shown to largely dictate the motion (vectoral trajectories) and mode of deposition of windborne pollen on ovule surfaces. Perpendicular orientation to the direction of airflow results in two regions characterized by high densities of adhering pollen — one on the windward surface of the ovule, resulting from direct inertial collision, and another on the leeward surface resulting from non-inertial sedimentation. Parallel and inclined orientations of the ovule to the direction of airflow produce quantitative and qualitative variations in the pattern of adhering pollen resulting from inertial and non-inertial deposition. Direct collision of windborne pollen grains with the micropylar ends of ovules occurs for all orientations to wind direction. The aerodynamics of the ovulate shoot complex of Taxus cuspidata is related to that previously described for conifer ovulate cones, cycad megastrobili, and simulated wind tunnel analyses of archaic Paleozoic ovules based on scale models. Water transport of pollen (adhering to integument and bract surfaces) to micropyles quantitatively alters the distribution of adhering pollen grains on ovule surfaces. Although there is no evidence that pollen grains of this species are osmotically ruptured, observations do not preclude the possibility that water transport of pollen may reduce the number of viable pollen grains reaching the micropyle.     

Why ant pollination is rare: new evidence and implications of the antibiotic hypothesis

The antibiotic hypothesis proposes that ant pollination is rare at least in part because the cuticular antimicrobial secretions of ants are toxic to pollen grains. We tested this hypothesis by comparing the effects of ants and bees on pollen in two regions: a tropical rainforest in Amazonian Peru and temperate forests and old fields in Canada. We found support for three predictions that follow from the antibiotic hypothesis. (1) For all 10 ant and 11 plant species in our study, contact with ants significantly reduced pollen germination, confirming the generality of this effect. (2) Contact with two bee species did not have similar effects; pollen exposed to bees germinated as well as control pollen. (3) Consistent with the presumed greater abundance of entomopathogens in the tropics, which may have selected for stronger antibiotic secretions in tropical ants, tropical ants had more negative effects on pollen than temperate ants. We speculate that the antibiotic hypothesis contributes not only to the rarity but also to the biogeography of ant pollination, and we discuss whether the negative effects of ants on pollen have resulted in selection for floral defenses against ants.     

FLOWER VISITATION,POLLEN DEPOSITION,AND POLLEN-TUBE COMPETITION IN HIBISCUS MOSCHEUTOS (MALVACEAE)

The potential influence of pollen-tube competition on offspring “quality” has received considerable attention in recent years. Yet the prevalence of pollen competition in natural populations is largely unknown because few investigators have actually measured rates of pollen deposition on stigmas. In this study, we assess the potential for pollen-tube competition in natural populations of the self-compatible, pollinator-dependent herbaceous perennial, Hibiscus moscheutos. Individual flowers averaged two to four visits per 15 min by potential pollinators (Ptilothrix and Bombus), and about 34% of these visits involved contact with a stigma. The median number of pollen grains deposited on virgin stigmas per contact visit was 70 grains (values ranged from 0 to 889), and flowers averaged about four contact visits per hour. Approximately 360 pollen grains must reach stigmas for full seed set to occur in a typical flower (an average ovary has 139 ovules, and 2.6 pollen grains are required per seed). Within 2 and 3 hr exposure to pollinators, 65% and 97% of the flowers received excess pollen (>360 grains) and median stigmatic pollen loads exceeded the number of ovules by a factor of 4.0 and 5.4, respectively. Based on 3 yr observations, it is concluded that pollen competition may frequently occur in this species.     

Comparative morpho-physiology of the metapleural glands of two Atta leaf-cutting ant queens nesting in clayish and organic soils

Queens of leaf-cutting ants found their nests singly, each consisting of a vertical tunnel and a final horizontal chamber. Because of the claustral mode of nest founding, the queen and/or her initial fungus garden are exposed to threats imposed by several soil pathogens, and the antibiotic secretions produced by their metapleural glands are considered a main adaptation to deal with them. Nests of two Atta leaf-cutting ant species, Atta vollenweideri and Atta sexdens rubropilosa, occur in different soil types, alfisols and oxisols. Their queens are known to excavate the initial nest in different soil horizons, clayish and organic, respectively, which differ in their fertility and associated microbiota. The aim of the present study was to comparatively investigate the morpho-physiology of the metapleural glands in queens of A. vollenweideri and A. sexdens rubropilosa, addressing the question whether the distinct selective pressure imposed by the microbiota in the two different soil types led to morpho-physiological differences in the metapleural glands that were consistent with their antiseptic function. The results revealed that metapleural glands of A. sexdens rubropilosa have a larger number of secretory cells, and consequently a higher production of antibiotic secretions, which may have been selected to allow nest founding at the superficial horizon of oxisols rich in organic matter and microorganisms. Glands of A. vollenweideri, on the contrary, presented fewer secretory cells, suggesting less production of antibiotic secretions. We argue that the excavation of deep founding nests in A. vollenweideri was primarily selected for during evolution to avoid the risk posed by flooding, and further hypothesize that a reduced number of cells in their metapleural glands occurred because of a weak pathogen-driven selective pressure at the preferred soil depth.     

Effects of ovular secretions on pollen in Pseudotsuga menziesii (Pinaceae)

Effects of ovular secretions on pollen grains were examined in Pseudotsuga menziesii. The exine is cast off in the micropylar canal. A membranelike structure covers parts of pollen grains and appears to protect them. The outer intine consists of fibrous materials, but it also shows a thicker filamentous appearance in some ovules during pollen elongation. The inner intine is electron-dense. Its fibrous nature is occasionally visible. Dissolution of the outer intine varies in amount and manner in ovules from different trees. The plasma membrane near the pollen wall alternatively appears normal and distorted. These different morphologies of the outer intine and of the plasma membrane are considered to result from secretions from the ovule. The outer intine may contain electron-dense globules that are formed in the tube cell and traverse the inner intine. Pollen tube formation appears to be triggered by a secretion from the ovule. Cross-pollinated grains are less distorted compared with self-pollinated grains.     

Lack of patriline-specific differences in chemical composition of the metapleural gland secretion in <Emphasis Type="Italic">Acromyrmex octospinosus</Emphasis>

Summary. Multiple queen-mating (polyandry) in social insects increases the genetic variability among worker offspring, which may enhance colony survival, social productivity and defence against parasites. The unique and complex symbiosis of leaf-cutting ants with a clonal mutualistic fungus makes this social system particularly vulnerable to contamination by pathogenic and unwanted saprophytic fungi and bacteria. Proper defence against such threats requires effective and flexible chemical defence mechanisms. A prime candidate for providing such defences is the metapleural gland secretion, which is known to have broad antibiotic properties. Here we use the leaf-cutting ant Acromyrmex octospinosus to specifically test the hypothesis that genetically more diverse worker-offspring produce a more variable spectrum of metapleural gland compounds. We used DNA microsatellite markers to assign workers from two colonies to the six most common patrilines in each colony, and have analysed the degree to which the observed variance in the quantitative chemical composition of the metapleural gland secretion can be explained by genetic differences among patrilines. We found a marginally significant patriline-effect on the overall variability of metapleural gland compounds in one colony, but could not detect such effect in the other colony. We discuss a number of possible reasons why the genetic variance component for quantitative variation in metapleural gland secretion may be low.     

POLLEN-OVULE RATIOS AND BREEDING SYSTEM EVOLUTION IN SOLANUM SECTION BASARTHRUM (SOLANACEAE)

Pollen-ovule ratio, pollen grains per flower, pollen grain volume, pollen biomass, ovules per ovary, seeds per fruit, seed volume, seed biomass, and corolla size were studied to reveal patterns of variation within a monophyletic group. Solanum sect. Basarthrum includes 22 neotropical species of two clades, one having small, few-seeded red fruits, and the other having larger, many-seeded green fruits. The former includes self-incompatible species and a dioecious species, while the latter includes self-incompatible species, self-compatible (and autogamous) species, and a domesticate (the “pepino”). Although the pollen quantity and ovule quantity of the self-incompatible species are significantly higher in the green-fruited subgroup than in the red-fruited subgroup, the pollen-ovule ratios of the self-incompatible species are not significantly different between the two subgroups, suggesting parallel evolution of the pollen-ovule ratio. Furthermore, the pollen-ovule ratio tracks the breeding system: self-incompatible species have significantly higher pollen-ovule ratios than self-compatible species, resulting both from fewer pollen grains and more ovules of the latter. The pollen-ovule ratio of the dioecious species is among the highest of all, resulting only from fewer ovules. The pepino is self-compatible but has a pollen-ovule ratio like its wild self-incompatible progenitors and shows wide variation in seed production and pollen quality, but not in pollen quantity. Among all species, pollen size and quantity are inversely proportional, as are seed size and quantity.     

Communication in the migratory termite-hunting ant Pachycondyla (= Termitopone) marginata (Formicidae, Ponerinae)

The Neotropical species Pachycondyla marginata conducts well-organized predatory raids on the termite species Neocapritermes opacus and frequently emigrates to new nest sites. During both activities the ants employ chemical trail communication. The trail pheromone orginates from the pygidial gland. Among the substances identified in the pygidial gland secretions, only citronellal was effective as a trail pheromone. Isopulegol elicited an increase in locomotory activity in the ants and may function as a synergist recruitment signal. The chemical signal is enhanced by a shaking display performed by the recruiting ant.     

Differential resistance and the importance of antibiotic production in Acromyrmex echinatior leaf-cutting ant castes towards the entomopathogenic fungus Aspergillus nomius

Paired exocrine metapleural glands are present in almost all ants and produce compounds with antibiotic properties towards a variety of pathogenic fungi and bacteria. In Acromyrmex leaf-cutting ants, small workers have relatively large metapleural glands compared to large workers, and thus harbour approximately half the number of gland cells of large workers, despite being only one-fifteenth their body mass. Here we present results showing that when the two worker castes of A. echinatior are treated with spores of the pathogenic fungus Aspergillus nomius in doses that correspond to the difference in metapleural gland cell numbers they do not differ in survival. However, we also show, for the first time, that small workers survive significantly longer than large workers when both are challenged with a dose of spores that corresponds to their difference in body mass. Furthermore, the time until Aspergillus nomius hyphae and spores appear on the cadavers of workers dead from infection, is significantly increased in the small worker caste. In addition to supporting previous findings that the metapleural glands have an important defence function, the results of this study indicate that the relatively large glands in small workers makes this caste particularly well adapted to preventing pathogenic microorganisms from entering the colony. Received 23 January 2006; revised 7 April 2006; accepted 11 April 2006.     

Embryology of the dioecious Woonyoungia septentrionalis (Magnoliaceae)

The embryological characteristics and ovular integument development of the dioecious species Woonyoungia septentrionalis (Dandy) Law (Magnoliaceae), which are poorly understood, were investigated under laser scanning confocal microscope (LSCM) and light microscope (LM). The embryological characteristics conform to most of the previously studied species in Magnoliaceae. The anther has 4 microsporangia, and the anther wall develops according to the dicotyledonous type. Cytokinesis at meiosis of the microspore mother cells follows a modified simultaneous type, giving rise to isobilateral or decussate tetrads, and a cell plate is absent, but a membrane was observed. Mature pollen grains are 2‐cellular and have high germination rates. The ovule is anatropous, crassinucellate and bitegmic, and meiotic result in linear tetrads of megaspores, the one at the chalazal end functions directly as an embryo‐sac cell. The development of the embryo sac is of the Polygonum‐type and endosperm formation is of the nuclear type. The outer integument of the ovule differentiates into an outer fleshy and an inner stony layer while the inner integument is reduced to a tanniniferous layer. The normal embryological development, high germination rates of pollen and high seed set indicate that the primary reason for the decline of the species is not to be found in these developmental processes.     

Sexual Calling by Workers Using the Metatibial Glands in the Ant, Diacamma sp., from Japan (Hymenoptera: Formicidae)

Several species of the ant genus Diacamma reproduce through mated workers (gamergates). Such gamergates have no wings and therefore are unable to conduct nuptial flight. Instead they perform a sexual calling behavior by standing outside the nest and rubbing the tibiae of their hindleg over the surface of the arched gaster. In a series of exclusion experiments we demonstrate that secretions from the metatibial gland are the most important component in making the virgin female attractive to the males.     

The embryology ofStegnospermataceae,with a discussion on its status,affinities and systematic position

The embryology ofStegnosperma halimifolium andS. watsonii has been studied in detail. The tapetum is of the secretory type and its cells become multinucleate. Simultaneous cytokinesis in the pollen mother cells follows meiosis. The ripe pollen grains are 3-celled. The ovule is crassinucellate, bitegmic and amphitropous, with the micropyle formed by the inner integument alone. The female archesporium is one celled, and the parietal tissue 3–5 layered. The embryo sac development conforms to thePolygonum type. A central strand, 6 or 7 cells thick, differentiates inside the nucellus and extends from the base of the embryo sac to the chalazal region. The endosperm is nuclear. The embryogeny conforms to the Caryophyllad type. The seed coat is formed by the outer epidermis of the outer integument and the inner epidermis of the inner integument. Based on this evidence and other data, the status of the genus as an independent family,Stegnospermataceae (Stegnospermaceae) is confirmed. Apparently, it forms a connecting link betweenPhytolaccaceae andCaryophyllaceae.     

COMPARATIVE POLLEN MORPHOLOGY AND TAXONOMIC AFFINITIES IN CYCADALES

Scanning and transmission electron microscopy of pollen grains of 29 species, representing the ten extant genera of Cycadales, has provided valuable insight into their relationships. Pollen grains of these taxa are boat-shaped, monosulcate, and bilaterally symmetrical. They range from narrowly to widely elliptical or subcircular when viewed distally, and have an exine surface of psilate, foveolate, or fossulate. Pollen wall ultrastructure of Cycadales is typically tectate with alveolate∗∗∗spongy exine. The nexine is laminated in all genera. Nexine 1 (footlayer) is present in most species as a thin and often discontinuous layer. There is consistent variation in thickness of the sporoderm layers among the genera but relative uniformity within them. Pollen characteristics are well correlated with macro- and micromorphological features, chromosome numbers, geographical distribution, and postulated pollination mode. A close affinity between Encephalartos, Lepidozamia, and Macrozamia is recognized. Pollen characteristics of the genus Bowenia show some similarity with those of the latter group. Except for two species of Macrozamia which are narrowly elliptic, all of the genera have widely elliptic pollen and share a psilate exine surface and the thinnest sexine with nearly identical arrangement of alveoli. Pollen grains of the species in the genus Dioon exhibit a unique morphology but are more similar to Stangeria than they are to those of taxa in Zamiaceae. The circular outline of the grains and the foveolate exine surface are characters shared by these two genera, but several morphological features distinguish Dioon from Stangeria. Ceratozamia and Zamia share a widely elliptic shape, foveolate exine surface and nearly identical sexine, as well as morphological features and chromosome numbers. They differ from Microcycas in sexine thickness, gross morphology and chromosome numbers. The pollen grains of Cycas circinalis and C. revoluta differ in size and structure of the sexine from all other genera and from each other, substantiating their distinct subgeneric delimitations.     

The metapleural gland of ants

The metapleural gland (MG) is a complex glandular structure unique to ants, suggesting a critical role in their origin and ecological success. We synthesize the current understanding of the adaptive function, morphology, evolutionary history, and chemical properties of the MG. Two functions of the MG, sanitation and chemical defence, have received the strongest empirical support; two additional possible functions, recognition odour and territorial marking, are less well supported. The design of the MG is unusual for insects; glandular secretions are stored in a rigid, non‐compressible invagination of the integument and the secretion is thought to ooze out passively through the non‐closable opening of the MG or is groomed off by the legs and applied to target surfaces. MG loss has occurred repeatedly among the ants, particularly in the subfamilies Formicinae and Myrmicinae, and the MG is more commonly absent in males than in workers. MG chemistry has been characterized mostly in derived ant lineages with unique biologies (e.g. leafcutter ants, fire ants), currently precluding any inferences about MG chemistry at the origin of the ants. A synthetic approach integrating functional morphology, phylogenetic transitions and chemical ecology of the MGs of both the derived and the unstudied early‐branching (basal) ant lineages is needed to elucidate the evolutionary origin and diversification of the MG of ants.     

Pollen size in Carex: The effect of different chemical treatments and mounting media

Pollen size and pollen aperture size for ten species of the genus Carex L., native to Estonia, have been measured using light microscopy. The species selected represent different sections of the genus, a range of habitats and different chromosome numbers. The effects of two basic chemical treatments, two mounting media and the effect of chemically induced dehydration with tertiary butyl alcohol (TBA) on the size of pollen grains were then recorded.

In general pollen size and pollen aperture size of the species examined is highly variable at both intraspecific and interspecific levels. Carex hirta has notably larger pollen grains than any of the other species investigated and, although correlations between size and chromosome number in the species examined are limited, it also has the highest chromosome number. Statistically significant size differences resulted from variations in chemical treatment, mounting media and tertiary butyl alcohol (TBA) induced dehydration. Acetolysed pollen grains are larger than potassium hydroxide (KOH) treated pollen grains. Pollen grains dehydrated after chemical treatment with TBA are larger than pollen grains not dehydrated. Pollen grains mounted in silicon oil are smaller than grains mounted in glycerine. But considering the great size variation of Carex pollen grains, the size changes caused by preparation procedures fall within the size variation range of the species examined.

All the samples contained a high number of deformed pollen grains and pollen grains with hardly distinguishable or no lateral apertures.     

Ant–hemipteran association decreases parasitism of Phenacoccus solenopsis by endoparasitoid Aenasius bambawalei

1. Mutualism between ants and honeydew-excreting hemipterans is ubiquitous in the ecosystem. It is widely accepted that ant tending facilitates the colony growth of hemipterans by protecting them from predators and parasitoids. However, few studies have explored how ant tending helps defend against natural enemies. 2. Ghost ant Tapinoma melanocephalum and the invasive mealybug Phenacoccus solenopsis have a close mutual relationship. Previous studies have shown that ghost ant tending can definitely reduce parasitism and visit frequency of Aenasius bambawalei, the dominant endoparasitoid of P. solenopsis. However, the ghost ant workers seldom attack the parasitoids. It is still unclear how the ghost ant adversely affects parasitoids. This study explored the mechanism underlying the impacts of ants on natural enemies of the mealybugs. 3. Honeydew produced by P. solenopsis was an attractant to A. bambawalei. Parasitoids exhibited less searching activity, shorter longevity and lower parasitism when supplied with less honeydew. Aenasius bambawalei showed significant avoidance of pygidial gland secretions and visual cues of ghost ants. Parasitism in plants treated with 6-methyl-5-hepten-2-one, actinidine, and gland extracts was significantly lower than that in plants treated only with solvents (paraffin oil or double-distilled water). 4. It is concluded that honeydew consumption by ghost ants could negatively influence the performance of parasitoids. The pygidial gland secretions and visual cues of ghost ants also significantly inhibit the parasitism. These results may contribute to a better understanding of the regulation mechanism in ant–hemipteran–enemy interactions.