Expandable spermatheca influences sperm storage in the simultaneously hermaphroditic snail Arianta arbustorum

Summary

In many simultaneously hermaphroditic land snail species, the sperm storage organ (spermatheca) is highly structured, suggesting that the female function might be able to influence offspring paternity. Physical properties of the sperm storage organ, including its initial size and sperm storage capacity, may also affect fertilization patterns in multiply mated snails. We examined the structure, volume and tubule length of empty spermathecae in the land snail, Arianta arbustorum, and assessed differences in spermatheca size following a single copulation. The number of spermathecal tubules ranged from 2–7, but was not correlated with the volume of empty spermathecae. The volume of sperm stored in the spermatheca after a copulation was correlated with neither the number of spermathecal tubules nor copulation duration. Mean spermathecal volume more than doubled between two and thirty-six hours after sperm uptake, but the length of the spermathecal tubules did not change. Interestingly, the volume of sperm stored in the spermatheca seems not to be related to the size of the spermatophore and thus not to the number of sperm received (= allosperm). The amount of allosperm digested in the bursa copulatrix was highly variable and no significant relationship with the size of the spermatophore received was found. These findings suggest that numerical aspects of sperm transfer are less important in influencing fertilization success of sperm in A. arbustorum than properties of the female reproductive tract of the sperm receiver.     

The spermatheca in the land snail, Arianta arbustorum (Pulmonata: Stylommatophora): muscle system and potential role in sexual selection

Abstract. Previous studies have shown variable patterns of paternity after multiple mating, and also variation in sperm storage among individuals of Arianta arbustorum , which suggests that the spermatheca may influence paternity in this promiscuous land snail. To identify possible morphological correlates of sperm manipulation, we investigated arrangement and ultrastructure of the muscles of the spermatheca. The musculature surrounding the 2–9 spermathecal tubules is arranged in a complex three dimensional network. In addition, each tubule has a thin sheath in which longitudinally oriented cells make up the innermost layer. Usually, the smooth muscle cells are enclosed by connective tissue. Only occasionally is direct muscle-muscle contact established through dense plaques. The short thick filaments, their small diameter, the relatively weak development of the tubular system and sarcoplasmic reticulum, and the low density of mitochondria indicate that the muscle cells contract relatively fast but with little strength, that they recover slowly, and have low endurance. A single muscle cell may be innervated by several axons and one axon may contact several muscle cells. Combining evidence of the present paper and a foregoing investigation on the spermathecal epithelium, we suggest that the main function of the spermathecal muscles is to expel sperm stored for fertilization, while the ciliation of the common duct is probably responsible for the distribution of sperm among the tubules.     

Functional association between female sperm storage organs and male sperm removal organs in calopterygid damselflies

Female damselflies in the family Calopterygidae have two sperm storage organs: a spherical bursa copulatrix and a tubular spermatheca. Male flies have a peculiar aedeagus with a recurved head with which to remove bursal sperm, and lateral spiny processes to remove spermathecal sperm. The lateral processes differ among species and populations in terms of their width relative to the spermathecal duct: the narrower processes are physically able to access spermathecal sperm, while the wider ones are not. In the present study, sperm storage patterns and aedeagal structures were compared between two calopterygid species with different spermathecal structures –Calopteryx cornelia and Mnais pruinosa– with respect to not only sperm quantity (number) but also sperm quality (viability), by using a recently developed method based on live/dead dual fluorescence. Calopteryx cornelia is a typical spermathecal sperm remover. In this species, viability was similar between bursal and spermathecal sperm. In contrast, in M. pruinosa, the spermatheca was much smaller than the bursa and often contained no sperm. Even when the spermatheca of this species did contain sperm, a high percentage of it was dead. Although the spermatheca of M. pruinosa has such atrophic tendencies, males have nevertheless developed long and spiny lateral processes similar to those of C. cornelia, suggesting the processes have functions other than spermathecal sperm removal. They possibly function as stoppers or guides for manipulating the aedeagal head to remove the sperm mass from the bursa.     

Functions of the spermathecal muscle of the boll weevil,Anthonomus grandis

In female boll weevils, Anthonomus grandis, spermathecal filling was not affected by severing the spermathecal muscles. Females whose spermathecal muscles were severed 2 to 4 weeks after mating laid infertile eggs and resumed virginal ovipositional behaviour indicating the importance of this muscle in supplying sperm for egg fertilization. The presence of normal active sperm within the spermatheca in no way influenced ovipositional behaviour. In females whose spermathecal muscles had been severed, 22 per cent sperm displacement occurred after a second mating compared with 66 per cent for normal females. The physical displacement of sperm was thus largely dependent on a functional spermathecal muscle.     

Spernathecal morphology,sperm transfer and a novel mechanism of sperm displacement in the rove beetle,Aleochara curtula (Coleoptera,Staphylinidae)

Summary The mechanism by which sperm are transferred from the male's spermatophore to the female's storing cage is described for the rove beetle Aleochara curtula, emphasizing a novel mechanism of sperm displacement by competing males. The cuticular, U-shaped spermatheca is equipped with a valve structure and two sclerotized teeth. The tube of the spermatophore extends into the spermathecal duct through the guidance of the flagellum of the male endophallus. Further elongation of the spermatophore tube, however, occurs only after separation of the pair. A primary tube bursts at its tip after passing through the valve. Within the lumen of the primary tube, a second tube passes through the valve and continues to extend up to the apical bulb of the spermatheca, doubles back on itself and swells to form a balloon filling most of the spermatheca. The balloon of the spermatophore is pierced within the spermatheca by tooth-like structures pressed against the spermatophore through contraction of the spermathecal muscle. The same process of spermatophore growing and swelling is also observed in mated females. Sperm from previous copulations are backflushed through the valve and the spermathecal duct, indicative of last-male sperm predominance.Abbreviations ad adhesive secretion covering the sperm - sac am amorphous secretion of the spermatophore - as ascending portion of the spermatophore - ds descending portion of the spermatophore - end parts of the male endophallus - ext extended tube - f flagellum - gs genital segment - lt large tooth - m muscle of the spermatheca - nsc non sclerotized cuticle - op opening of the spermathecal gland - pt primary tube - sc sclerotized cuticle - sd spermathecal duct - se secretion of the spermathecal gland - sf secretion flowing out of the primary tube - sg spermathecal gland - sm sperm - smt small tooth - sp spermatheca - ss sperm sac - st secondary tube - vm vaginal muscle     

Paternity analysis of wild‐caught females shows that sperm package size and placement influence fertilization success in the bushcricket Pholidoptera griseoaptera

In species where females store sperm, males may try to influence paternity by the strategic placement of sperm within the female's sperm storage organ. Sperm may be mixed or layered in storage organs, and this can influence sperm use beyond a ‘fair raffle’. In some insects, sperm from different matings is packaged into discrete packets (spermatodoses), which retain their integrity in the female's sperm storage organ (spermatheca), but little is known about how these may influence patterns of sperm use under natural mating conditions in wild populations. We examined the effect of the size and position of spermatodoses within the spermatheca and number of competing ejaculates on sperm use in female dark bushcrickets (Pholidoptera griseoaptera) that had mated under unmanipulated field conditions. Females were collected near the end of the mating season, and seven hypervariable microsatellite loci were used to assign paternity of eggs laid in the laboratory. Females contained a median of three spermatodoses (range 1–6), and only six of the 36 females contained more than one spermatodose of the same genotype. Both the size and relative placement of the spermatodoses within the spermatheca had a significant effect on paternity, with a bias against smaller spermatodoses and those further from the single entrance/exit of the spermatheca. A higher number of competing males reduced the chances of siring offspring for each male. Hence, both spermatodose size and relative placement in the spermatheca influence paternity success.     

A light and electron microscopical study of the spermathecae and ventral receptacle of Anastrepha suspensa (Diptera: Tephritidae) and implications in female influence of sperm storage

Female insects with multiple sperm storage organs may potentially influence patterns of paternity by differential storage of sperm from competing males. The Caribbean Fruit Fly, Anastrepha suspensa, stores sperm differentially with respect to its three spermathecae. To understand the mechanisms and processes responsible for patterns of sperm storage and use in A. suspensa, details of the fine structure of female sperm storage organs were resolved by UV-light microscopy, confocal microscopy, tissue sectioning, and scanning and transmission electron microscopy. Structures not previously described for this species include a ventral receptacle for sperm storage and osmoregulation, a conical-shaped valve at the junction between the spermathecal capsules and their ducts, laminar and granular secretions, secretions from the signum, hemocytes surrounding the spermathecae, and spermathecae with sclerotized, hollow projections that terminate in single glandular cells. The independent organization of sperm storage organs, spermathecal ducts, associated musculature, gland cells, and innervation offer possible mechanisms by which sperm movement may be influenced by females. The implications of these structures for insemination and fertilization events are discussed.     

Ultrastructure of the spermatheca and its associated gland in the ant Crematogaster opuntiae (Hymenoptera,Formicidae)

Summary Sperm storage by females has reached an extreme degree of development in ants. Ant queens, which are unusually long-lived insects, typically store and maintain an unreplenished supply of viable sperm for ten or more years. The spermatheca of Crematogaster opuntiae includes a receptacle and a discrete pair of accessory, or spermathecal, glands, structures commonly found in sperm storage organs of insects. The bean-shaped receptacle consists of a layer of simple epithelium externally and a cuticular layer internally. In the hilar region, the epithelium is highly columnar and exhibits ultrastructural features characteristic of transport epithelia, such as infolded basal membranes, abundant polymorphic mitochondria, and apical microvilli. The spermathecal glands contain cells that have long, dense microvilli that project into a central lumen, abundant mitochondria, and large fields of glycogen. The valve and pump region of the spermatheca provide a mechanism to conserve sperm by controlling the rate of sperm release. The columnar epithelium may function as excretory tissue that serves to maintain an environment in which sperm can remain viable for many years.     

Sperm aggregations in the spermatheca of the red back salamander (Plethodon cinereus)

The spermatheca of Plethodon cinereus is a compound tubular gland that stores sperm from mating in early spring (March–April) to oviposition in summer (June–July). The seasonal variation of sperm storage in this species has previously been studied by light and transmission electron microscopy. In this paper, sperm aggregations, interaction of sperm with the spermathecal epithelium, and spermathecal secretions are studied using scanning electron microscopy. Within spermathecal tubules, relatively small groups of sperm are aligned along their entire lengths in parallel arrays. This pattern is similar to other plethdontids with complex spermathecae. Lumina of spermathecal tubules are filled with secretory material in April prior to the arrival of sperm, and after sperm appear, a coating of secretory material persists on the apices of the spermathecal epithelium. Sperm peripheral to the central luminal mass can become embedded in the secretory matrix or pushed deeper into the spermathecal epithelium. The spermathecal secretions may serve to attract and prolong the viability of sperm, but sperm that become enmeshed in the secretions or epithelium are phagocytized. Sperm and spermathecal secretions are largely absent after ovulation and in summer months, and new secretory vacuoles are formed in fall, although mating does not occur until spring.     

Morphology of the female reproductive system of European pea crabs (Crustacea,Decapoda, Brachyura,Pinnotheridae)

Commensal pea crabs inhabiting bivalves have a high reproductive output due to the extension andfecundity of the ovary. We studied the underlying morphology of the female reproductive system in the Pinnotheridae Pinnotheres pisum, Pinnotheres pectunculi and Nepinnotheres pinnotheres using light microscopy and transmission electron microscopy (TEM). Eubrachyura have internal fertilization: the paired vaginas enlarge into storage structures, the spermathecae, which are connected to the ovaries by oviducts. Sperm is stored inside the spermathecae until the oocytes are mature. The oocytes are transported by oviducts into the spermathecae where fertilization takes place. In the investigated pinnotherids, the vagina is of the “concave pattern” (sensu Hartnoll 1968 ): musculature is attached alongside flexible parts of the vagina wall that controls the dimension of its lumen. The genital opening is closed by a muscular mobile operculum. The spermatheca can be divided into two distinct regions by function and morphology. The ventral part includes the connection with vagina and oviduct and is regarded as the zone where fertilization takes place. It is lined with cuticle except where the oviduct enters the spermatheca by the “holocrine transfer tissue.” At ovulation, the oocytes have to pass through this multilayered glandular epithelium performing holocrine secretion. The dorsal part of the spermatheca is considered as the main sperm storage area. It is lined by a highly secretory apocrine glandular epithelium. Thus, two different forms of secretion occur in the spermathecae of pinnotherids. The definite role of secretion in sperm storage and fertilization is not yet resolved, but it is notable that structure and function of spermathecal secretion are more complex in pinnotherids, and probably more efficient, than in other brachyuran crabs. J. Morphol., 2011. © 2010 Wiley‐Liss, Inc.     

Function of multiple sperm-storage organs in female damselflies (Ischnura senegalensis): difference in amount of ejaculate stored, sperm loss, and priority in fertilization

We studied changes in the number of sperm within two kinds of female sperm-storage organ in the damselfly Ischnura senegalensis (Odonata: Coenagrionidae): the bursa copulatrix and the spermatheca. We counted the number of sperm within each storage organ and tested their viability after a single copulation in female damselflies kept for seven days with and without oviposition. We also counted sperm and tested their viability in females that underwent an interrupted second copulation after the sperm-removal stage, and after subsequent oviposition. Our results showed that the bursa copulatrix and spermatheca have different sperm storage roles. Immediately after copulation, most eggs appear to have been fertilized with bursal sperm, which were positioned near the fertilization point. By seven days after copulation, a greater proportion of spermathecal sperm were used for fertilization, as the number of bursal sperm had decreased. We hypothesize that female damselflies use the spermatheca for long-term storage and the bursa copulatrix for short-term storage: bursal sperm are more likely to be used for fertilization but may have a higher risk of mortality due to sperm removal by a competing male and/or sperm expelling by the female, whereas spermathecal sperm are safer but will be used for fertilization only after their release from the spermatheca.     

Sperm numbers,their storage and usage in the fly Dryomyza anilis

In the fly Dryomyza anilis females have two kinds of sperm storage organs: one bursa copulatrix and three spermathecae (two spermathecae with a common duct form the doublet, and the third is a singlet spermathecal unit). At the beginning of a mating the male deposits his sperm in the bursa copulatrix. After sperm transfer the male taps the female''s abdomen with his claspers. This behaviour has been shown to increase the male''s fertilization success. After mating, the female discharges large quantities of sperm before oviposition. To find out where the sperm remaining in the female are stored, I counted the number of sperm in the droplet and in the female''s sperm storage organs after different types of mating. I carried out three mating experiments. In experiment 1, virgin females were mated with one male and the matings were interrupted either immediately after sperm transfer or after several tapping sequences. The results show that during male tapping more sperm moved into the singlet spermatheca. In addition, the total number of sperm correlated with sperm numbers in all sperm storage organs, and male size was positively related to the number of sperm remaining in the bursa. In experiment 2, females mated with several males. The number of sperm increased with increasing number of matings only in the doublet spermatheca. No increase in the number of sperm in the singlet spermatheca during consecutive matings suggests that sperm were replaced or did not reach this sperm storage organ. In experiment 3, virgin females were mated with a single male and half of them were allowed to lay eggs. The experiment showed that during egglaying, females primarily used sperm from their singlet spermatheca. The results from the three experiments suggest that sperm stored in the singlet spermatheca is central for male fertilization success and male tapping is related to sperm storage in the singlet spermatheca. The different female''s sperm storage organs in D. anilis may have separate functions during sperm storage as well as during sperm usage.     

Ultrastructural analysis of the morphology and function of the spermatheca of the pulmonate snail,Sonorella santaritana

The ultrastructure of the spermatheca of the reproductive tract in the pulmonate snail, Sonorella santaritana, was investigated. This organ has a debris-filled lumen and an outer wall which can be divided into three distinct layers. The cell layer adjacent to the lumen is comprised of two cell types, tall columnar epithelial cells with microvilli and cells lacking microvilli. The next layer also has two cell types, muscle cells and apparent pigment cells. The most distant layer is an adventitia of large glycogen-containing cells. The lumen of the spermatheca contains a core of partially digested sperm and related materials. The luminal contents and the cellular morphology of this organ suggest that the spermathecal functions are both digestive and absorptive. It is proposed that excess sperm and related materials are transported to the spermatheca, digested, and the usable products are reabsorbed.     

A newly discovered sperm transport system in the female of Lygaeidae bugs

In the female reproductive system of the relatively large hemipteran, the western conifer seed bug Leptoglossus occidentalis (Heidemann), a cuticle‐lined tube extends medially along the surface of the vagina from the proximal end of the spermathecal complex anteriorly to the base of the common oviduct. This medial tube houses the proximal end of the spermathecal duct, thereby enabling the transport of material from the spermatheca at the distal end of the spermathecal complex, past the vagina (or bursa copulatrix) and directly to the common oviduct. The proximal portion of the spermathecal complex also contains an insemination duct that is separate from the spermathecal duct. The insemination duct allows the male intromittent organ to extend from the vagina to the spermatheca without navigating through the spermathecal duct. The reproductive systems of two previously studied Hemiptera, the milkweed bug Oncopeltus fasciatus (Dallas) and the box elder bug Leptocoris trivittatus (Say), possess a similar cuticle‐lined medial tube housing the spermathecal duct. This new information provides a clearer understanding of sperm transport in the female reproductive system of Lygaeidae bugs, and helps to clarify the path of the male organ during copulation, as well as the movement of sperm during egg laying.     

Variation in spermathecal morphology is independent of sperm competition intensity in populations of the simultaneously hermaphroditic land snail Cornu aspersum

The complexity of the sperm-storing organ (spermatheca) has been hypothesized to reflect sperm competition intensity in several gastropod species. Furthermore, considerable variation in spermathecal morphology has been detected among populations of the same species. The morphological variation of the fertilization pouch was studied in five populations of the simultaneously hermaphroditic land snail Cornu aspersum (formerly, Helix aspersa). The populations studied differed in snail density and habitat humidity regimes, thus in sperm competition intensity. The study was conducted on wild adult snails and their progeny, which was reared in the laboratory for two successive generations. Finally, the morphology of the spermatheca was correlated to behavioral mating traits of the snails. The fertilization pouch consisted of a simple fertilization chamber and 4-19 blind tubules. The five studied populations did not differ in either mean number of spermathecal tubules, length of the fertilization chamber, length of the main tubule, or cumulative length of all tubules, while they differed in copulation frequency and mating propensity. No correlation was found between snail size and number of tubules, or length of any spermathecal structure measured. Additionally, no correlation was found between any behavioral trait and the morphological characteristics of the spermatheca. Strong correlations were found only among measurements of some of the spermathecal structures. Our results suggest that the complexity of the spermatheca is not related to sperm competition intensity and its structure is thus genetically determined.     

Beyond the mouse model: Using Drosophila as a model for sperm interaction with the female reproductive tract

Although the fruit fly, Drosophila melanogaster, has emerged as a model system for human disease, its potential as a model for mammalian reproductive biology has not been fully exploited. Here we describe how Drosophila can be used to study the interactions between sperm and the female reproductive tract. Like many insects, Drosophila has two types of sperm storage organs, the spermatheca and seminal receptacle, whose ducts arise from the uterine wall. The spermatheca duct ends in a capsule-like structure surrounded by a layer of gland cells. In contrast, the seminal receptacle is a slender, blind-ended tubule. Recent studies suggest that the spermatheca is specialized for long-term storage, as well as sperm maturation, whereas the receptacle functions in short-term sperm storage. Here we discuss recent molecular and morphological analyses that highlight possible themes of gamete interaction with the female reproductive tract and draw comparison of sperm storage organ design in Drosophila and other animals, particularly mammals. Furthermore, we discuss how the study of multiple sperm storage organ types in Drosophila may help us identify factors essential for sperm viability and, moreover, factors that promote long-term sperm survivorship.     

Sperm transfer, sperm storage, and sperm digestion in the hermaphroditic land snail Succinea putris (Gastropoda, Pulmonata)

Abstract. Many hermaphroditic species are promiscuous, have a sperm digesting organ and an allosperm storage organ (i.e., spermatheca) with multiple compartments (i.e., spermathecal tubules) providing opportunities for sperm competition. The relative paternity of a sperm donor drives the evolution of mating behaviors that allow manipulation of the sperm receiver's reproductive behavior or physiology. We studied the relationship between sperm transfer, sperm storage, sperm digestion, and copulation duration in the hermaphroditic land snail Succinea putris , in which an active individual mates on top of a passive individual. Specifically, we examined (i) whether the entire copulation duration was required to complete reciprocal sperm transfer, (ii) sperm transfer patterns and their relationship with activity role, and (iii) the timing of sperm storage and sperm digestion. We found that reciprocal sperm transfer was completed within the first 5 h of copulation, which is ∼2–3 h before the end of copulation. Sperm transfer was mainly sequential, meaning that one individual donated all his ejaculate before its partner started to reciprocate. The initiation of sperm transfer did not depend on the activity role. The presence of allosperm in the spermatheca before sperm transfer suggests that individuals remate before they are allosperm depleted. No sperm was digested during copulation but sperm digestion took place 0–72 h after copulation. Our results suggest that contact mate guarding is a likely manipulation strategy in S. putris , because partners cannot immediately remate. In addition, staying in copula after sperm transfer is completed seems to prevent the immediate digestion of sperm and therefore may promote sperm displacement and allosperm storage.     

Post-mating effects in the grasshopper, Gomphocerus rufus L. mediated by the spermatheca

In the female grasshopper Gomphocerus rufus mating replaces copulatory readiness with immediate and long-lasting `secondary defense', during which further mating attempts are efficiently repelled. The behavioral change is caused by secretions from the male accessory glands' white secretory tubule 1 which is injected with the spermatophore material into the female's spermathecal duct. A bristle field of contact chemoreceptors at the entry of the spermathecal duct into the endbulb is assumed to be stimulated by the secretion. Ablation of the bristle field, interruption of the nervous pathway between the spermatheca and the ventral nervecord, or severance of the latter sustains sexual receptivity after mating. Both the secretion from white secretary tubule 1 and the spermatophore contained in the spermatheca of a mated female are digested by proteolytic enzymes from spermathecal gland cells. Dissolved material is resorbed by similar glandular-like cells. The intersexual conflicts of interest and their evolutionary consequences are discussed. Accepted: 4 December 1998     

Histological study of the spermatheca in three thelytokous parthenogenetic ant species,Pristomyrmex punctatus,Pyramica membranifera and Monomorium triviale (Hymenoptera: Formicidae)

Gotoh, A., Billen, J., Tsuji, K., Sasaki, T. and Ito, F. 2011. Histological study of the spermatheca in three thelytokous parthenogenetic ant species, Pristomyrmex punctatus, Pyramica membranifera and Monomorium triviale (Hymenoptera: Formicidae). —Acta Zoologica (Stockholm) 00 :1–8. The evolution of obligate parthenogenesis may induce the degeneration of female mating ability and subsequently affect the morphology of the female reproductive organs related to mating and/or sperm storage. Here, we investigated the size and structure of the sperm storage organ, the spermatheca, in three thelytokous parthenogenetic myrmicine ant species, Pristomyrmex punctatus, Pyramica membranifera and Monomorium triviale, and compared it with that of their related sexually reproducing species. So far, mated individuals have never been found in these three species, which appears to be in line with their parthenogenetic status. Although the spermatheca appears to be useless in these species, we could not find any evidence on the degeneration in size and morphology of their spermathecae. The spermathecal reservoir still has the columnar hilar epithelium, which is one of the major features for a functional spermatheca in ants.