Examination of the interior of sand fly (Diptera: Psychodidae) abdomen reveals novel cuticular structures involved in pheromone release: Discovering the manifold

The males of many species of New World Phlebotomines produce volatile terpenoid chemicals, shown in Lutzomyia longipalpis s.l. to be sex/aggregation pheromones. Pheromone is produced by secretory cells which surround a cuticular reservoir which collects the pheromone and passes it through a cuticular duct to the surface of the insect. The pheromone then passes through specialised cuticular structures on the abdominal surface prior to evaporation. The shape and distribution of the specialised structures are highly diverse and differ according to species. In this study we used SEM to examine the interior cuticular pheromone collection and transport structures of 3 members of the Lu. longipalpis s.l. species complex and Migonemyia migonei. We found a new structure which we have called the manifold which appears to be a substantial extension of the interior tergal cuticle connected in-line with the cuticular duct and reservoir. The manifold of the Campo Grande member of the complex is longer and wider than the Jacobina member whereas the manifold of the Sobral member was shorter than both other members of the complex. Overall, the secretory apparatus of the Sobral member was smaller than the other two. The manifold of M. migonei was very different to those found in Lu. longipalpis s.l. and was positioned in a pit-like structure within the tergal cuticle. The secretory reservoir was connected by a short duct to the manifold. Differences in the size and shape of the manifold may be related to the chemical structure of the pheromone and may have taxonomic value. Examination of the interior cuticle by SEM may help to locate the secretory apparatus of vector species where pheromonal activity has been inferred from behavioural studies but the external secretory structures or pheromones have not yet been found.

The Role of Filippi’s Glands in the Silk Moths Cocoon Construction

Filippi’s glands (FGs), formerly also called Lyonet’s glands, are accessory secretory structures of the labial (silk) glands of lepidopteran caterpillars, which were implicated to play an important role in the maturation of the silk material and the construction of the cocoon. In our previous study, we have identified several species of giant silk moths that completely lack the FGs. Interestingly, the absence of FGs in these species correlates with the construction of a loose cocoon architecture. We investigated the functions of FGs by their surgical extirpation in the last instar larvae of the silkworm, Bombyx mori. We found that the absence of FGs altered the structure of the resulting cocoon, in which the different layers of silk were separated. In further experiments, we found no effects of the absence of FGs on larval cocoon formation behavior or on changes in cocoon mass or lipid content. Differential proteomic analysis revealed no significant contribution of structural proteins from FGs to silk cocoon material, but we identified several low abundance proteins that may play a role in posttranslational modifications of some silk proteins. Proteomic analysis also revealed a difference in phosphorylation of the N-terminal sequence of fibroin-heavy chain molecule. Thus, FGs appear to affect silk stickiness during spinning by regulating posttranslational modifications. This could also explain the link that exists between the absence of these glands and the formation of loose cocoons in some giant silk moth species.

New Aspects of Secretory Structures in Five Alismataceae Species: Laticifers or Ducts?

The secretory structures of Alismataceae have been described as secretory ducts, laticifer ducts, laticifer canals or schizogenous ducts. However, these terms are not found in the specialized literature, and ontogenetic analyses for the exact classification of these structures are missing. Accordingly, more studies regarding the secretory structures of Alismataceae are necessary to establish homology in the family or in the order. Thus, the aim of this study was to describe the anatomy, ontogeny, distribution in the organs and exudate composition of the secretory structures present in five Alismataceae species in order to determine whether the family has laticifers or secretory ducts. Samples of leaves, flowers and floral apices were processed for anatomical and histochemical analyses by light microscopy. The analysis indicated the presence of anastomosing secretory ducts in all species, occurring in both leaves and flowers. The exudate contains lipids, alkaloids, proteins and polysaccharides, including mucilage. The secretory duct structure, distribution and exudate composition suggest a defense role against herbivory and in wound sealing. The presence of secretory ducts in all species analyzed indicates a probable synapomorphy for the family.

Pollination and Floral Biology of a Rare Morning Glory Species Endemic to Thailand, Argyreia siamensis

Argyreia siamensis is extremely rare, and very little is known about its reproduction. The species has colorful flowers that seem likely to attract pollinators, but population sizes are typically small (<30 individuals). To determine whether poor reproduction contributes to its rarity, we investigated its mating system and potential pollinators in two populations. We also examined the staminal trichomes and floral nectary to investigate their role in pollinator attraction. The mating system was assessed with a bagging experiment and pollinator visits were recorded with action cameras. Additionally, we tested the staminal trichomes and floral nectary for terpenes and flavonoids and examined floral nectary micromorphology via scanning electron microscope and compound light microscope. Our results reveal that A. siamensis is self-incompatible and dependent on pollinators; the western population was pollinated by bees (Meliponini and Amegilla), while the eastern population was mainly pollinated by skipper butterflies (Hesperiidae). Both staminal trichomes and the floral nectary appear to contribute to pollinator attraction through the presence of terpenes and flavonoids (in both secretory structures) and nectariferous tissue and nectarostomata (in the nectary). Our results indicate that A. siamensis has reliable and effective pollinators and that insufficient pollination is likely not a primary cause of its rarity.

Histochemical observations in Piper malgassicum (Piperaceae) with a special focus on the epidermis

This is the first contribution about the histochemistry of vegetative and reproductive aerial organs in the genus Piper L. Piper malgassicum accumulates alkaloids and terpenes in the epidermis and the underlying layers of parenchyma, both in the leaves, in the stems and in anthers. Some idioblasts appear to contain a large amount of secondary metabolites. The micro-anatomical analysis showed peculiar secretory structures both in the leaves, in the anthers and in the ovary. Several lipid aggregates, alkaloid droplets and calcium oxalate crystals were observed in leaves and stems, indicating their role in defence strategies, mechanical support, and pollinators attraction. In the anthers, we observed elaioplasts whose content suggest an alternative and indirect function in pollination and defence against micro-organisms. Besides, some lipid aggregates surrounded by microtubules, detected in the anthers, were recognized as lipotubuloids. The tapetum was of secretory type. Alkaloids and terpenes were widely distributed in the plant confirming the important biological role of this type of biomolecules and its functional range. In the anthers, terpene and polyphenol inclusions appeared particularly abundant in the epidermal layer, whereas calcium oxalate crystals were observed close to the ovule in the ovary at maturity.

Interior of sand fly (Diptera: Psychodidae) abdomen reveals novel structures involved in pheromone release: discovering the Manifold.

The males of many species of New World Phlebotomines produce volatile terpenoid chemicals which have been shown in Lutzomyia longipalpis s.l. and L. cruciata to be sex/aggregation pheromones which attract female and male conspecifics. Pheromone is produced in secretory cells surrounding a cuticular reservoir which collects the pheromone and passes it through a cuticular duct to the surface of the insect. On the surface the pheromone passes through a specialised structure prior to evaporation. The shape and distribution of the structures are highly diverse and differ according to species. They range in appearance from slightly raised domes (papules) to almost spherical apple shaped structures to slight depressions with central spikes and all with a central pore. They can occur either singly or in many hundreds distributed on most abdominal tergites or grouped on one. The pheromone secreting apparatus in sand flies and other insects have historically been examined from the exterior using scanning electron microscopy (SEM) and from the interior using transmission electron microscopy. In this study we used SEM to examine the interior cuticular structure of 3 members of the Lutzomyia longipalpis s.l. species complex and Migonemyia migonei and found a new structure associated with pheromone release which we have called the Manifold. The Manifold is a substantial structure siting in-line between the cuticular duct and the underside of the tergite. Differences in the size and shape of the Manifold may be related to the chemical structure of the pheromone. In addition to the importance of this hitherto unknown structure in the production, dissemination and ecology of the pheromone, as well as its potential taxonomic value, examination of the interior cuticle by SEM may help locate the secretory apparatus in important vector species where pheromonal activity has been inferred from behavioural studies but the external secretory structures or potential pheromones have not been found.

Colleters, Extrafloral Nectaries, and Resin Glands Protect Buds and Young Leaves of Ouratea castaneifolia (DC.) Engl. (Ochnaceae)

Buds usually possess mechanical or chemical protection and may also have secretory structures. We discovered an intricate secretory system in Ouratea castaneifolia (Ochnaceae) related to the protection of buds and young leaves. We studied this system, focusing on the distribution, morphology, histochemistry, and ultrastructure of glands during sprouting. Samples of buds and leaves were processed following the usual procedures for light and electron microscopy. Overlapping bud scales protect dormant buds, and each young leaf is covered with a pair of stipules. Stipules and scales possess a resin gland, while the former also possess an extrafloral nectary. Despite their distinct secretions, these glands are similar and comprise secreting palisade epidermis. Young leaves also possess marginal colleters. All the studied glands shared some structural traits, including palisade secretory epidermis and the absence of stomata. Secretory activity is carried out by epidermal cells. Functionally, the activity of these glands is synchronous with the young and vulnerable stage of vegetative organs. This is the first report of colleters and resin glands for O. castaneifolia. We found evidence that these glands are correlated with protection against herbivores and/or abiotic agents during a developmental stage that precedes the establishment of mechanical defenses.