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Distal part of the phallus of Cirrhia icteritia (Noctuidae) macerated with 10% KOH solution stained with chlorazol black and vesica inflated with absolute ethanol (light microscopy). The vesica of this species has three types of fixed cornuti attached to different areas. The black conical object is the tip of an insect pin. a Dorsal view. b Right view. c Ventral view. d Left view. 1 small cornutus; 2 large cornutus; 3 group of small cornuti (spinules). Scale bar 1 mm
Source publication
Existing techniques for examining the everted vesica (endophallus) of Lepidoptera are based primarily on cuticular preparations macerated with a caustic solution for taxonomic study. These techniques destroy muscles and other soft tissue, thus studies of the functional anatomy of the skeletomuscular apparatus of the phallus are not possible. Inject...
Citations
... The first ejaculated material (semi-liquid) is transported into the bursa during this time. In all subsequently fixed stages (30,40,50,60, 70 and 80 min), the vesica is everted, but not inflated by haemolymph. Instead, its volume is occupied by the expanded cuticular simplex filled with denser, jelly-like ejaculate, particularly in the 80 min, when the collum (the most rigid of all materials) of the spermatophore is transferred. ...
... 90 min under these conditions, and the moths were fixed eventually. One pair was fixed at every 10 ± 2 min after the onset of copulation, i.e., one couple at ca. 10,20,30,40,50,60,70, and 80 min. At the desired moment, 10 ml of freshly prepared and cooled (− 18 °C) Duboscq-Brasil fixative was poured on the copulating pair. ...
... The visualization of the muscles via confocal laser scanning microscopy (CLSM) follows Zlatkov et al. [50] with some modifications. Initially, the samples were fixed in Bouin's fluid and after washing, stored in ethanol. ...
Background
The process of copulation in Lepidoptera is understudied and poorly understood from a functional perspective. The purpose of the present paper is to study the interaction of the male and female genitalia of Tortrix viridana Linnaeus, 1758 via three-dimensional models of pairs fixed during copulation. Other techniques (confocal laser scanning microscopy, scanning electron microscopy and histology) were used to clarify the role of the organs involved in the process.
Results
Three-dimensional models based on micro-CT scanned copulating pairs were generated allowing visualisation of the position of the male and female counterparts, spatial changes during copulation, and the skeleto-muscular apparatus involved in the process. The male genitalia and their musculature are simplified in comparison with other lineages of the family, but the opposite is true for the female genitalia. The attachment of the couple is achieved only through flexion of the valvae, clasping the large and sclerotised sternite 7 of the female. The anal cone and socii of the male are in contact with certain parts of the anal papillae and the sterigma of the female. The long tubular vesica is inserted into the narrow posterior part of the ductus bursae. Its eversion is achieved by an increase in haemolymph pressure. A possible mechanism of stimulation of the female via pulsations of the diverticulum of the vesica was discovered. A compressed sclerotised area of the ductus bursae putatively serves as a valve controlling the transfer of ejaculated materials. Copulation progresses through two phases: in the first the vesica and its diverticulum are inflated by haemolymph, and in the second the diverticulum is not inflated, and the vesica is occupied by viscous ejaculated material. The formation of the multilayered spermatophore was observed, and we discovered that sperm is transferred very late in the copulation process.
Conclusions
Copulation process in Lepidoptera is studied for the first time with three-dimensional reconstructions of couples of Tortrix viridana, used as a model species. The internal genitalia is the scenario of multiple interactions between male and female, but the external remain static. A possible mechanism of stimulation of the female internal copulation organs is proposed.
