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Among adaptive traits under sexual selection, the length of spermatozoa shows high interspecific variation. In insects, extremes exist for both short and long sperm. The spermatozoa of the endoparasitic wasp Cotesia congregata (Say) are the shortest flagellated sperm described in animals, 6.6 µm in length. By comparison, the sperm of Drosophila bifurca are almost 6000 times longer. Thus, C. congregata has the potential to shed light on the selection pressures that drive variation in sperm length in relation to their production and use. The reproductive organs, sperm counts, controlled oviposition and sex ratios were investigated. The testes showed stratified differentiation stages of spermatogenesis, and sperm counts revealed continuous spermatogenesis in the late pupal stage. The small female spermatheca stored ~1000 sperm, resulting in an extremely high sperm concentration. The number of progeny per brood decreased over time until depletion of eggs. Females produced up to 370 daughters, corresponding to the effective use of 34% of the average sperm stock. Haploid males made up a greater proportion of broods in later parasitisms. Sperm miniaturization may be an adaptation to transfer increased quantities for the entire reproductive life of females in the absence of sperm competition but in the reduced space offered by the spermatheca.

Conférence en ligne aux Journées Francophones de Nutrition - JFN du 10 au 12/11/2021.; International audience; In urban and peri-urban areas, where waste management is more than ever a major issue, insect farming may be an ecological way to recycle bio-waste in a circular bioeconomy approach. Indeed, someinsect species such as Hermetia illucens can grow on many types of organic substrates and can convert them into valuable byproducts(1). There may be a wide variety of insect bioconversion products since larvae are a natural source of micronutrients, amino acids and antimicrobial compounds. They are also rich in proteins which can be used for animal feed, in fats used for the production of biodiesel, in chitin used in various industries, and they produce a type of manure called frass that can be used as biofertilizer(2). However, besides its ecological and nutritional benefits, risks of insectbased bioconversion should be identified and monitored. Indeed, some chemical contaminants may be found in insect farming environment, in their feeding substrates or can be produced during processing methods(3). Moreover, microbiological contaminants may be found in the farming insect chain(4). Therefore, it is necessary to assess the impact of these contaminants on insect survival and to determine their fate in the insect. It is also important to assess the environmental, social and economic impact of insect-based bioconversion and to determine the consumer acceptability of these new processes and products

Endogenous viruses form an important proportion of eukaryote genomes and a source of novel functions. How large DNA viruses integrated into a genome evolve when they confer a benefit to their host, however, remains unknown. Bracoviruses are essential for the parasitism success of parasitoid wasps, into whose genomes they integrated ~103 million years ago. Here we show, from the assembly of a parasitoid wasp genome at a chromosomal scale, that bracovirus genes colonized all ten chromosomes of Cotesia congregata. Most form clusters of genes involved in particle production or parasitism success. Genomic comparison with another wasp, Microplitis demolitor, revealed that these clusters were already established ~53 mya and thus belong to remarkably stable genomic structures, the architectures of which are evolutionary constrained. Transcriptomic analyses highlight temporal synchronization of viral gene expression without resulting in immune gene induction, suggesting that no conflicts remain between ancient symbiotic partners when benefits to them converge.

Endogenous viruses form an important proportion of eukaryote genomes and a source of novel functions. How large DNA viruses integrated into a genome evolve when they confer a benefit to their host, however, remains unknown. Bracoviruses are essential for the parasitism success of parasitoid wasps, into whose genomes they integrated ~103 million years ago. Here we show, from the assembly of a parasitoid wasp genome at a chromosomal scale, that bracovirus genes colonized all ten chromosomes of Cotesia congregata. Most form clusters of genes involved in particle production or parasitism success. Genomic comparison with another wasp, Microplitis demolitor, revealed that these clusters were already established ~53 mya and thus belong to remarkably stable genomic structures, the architectures of which are evolutionary constrained. Transcriptomic analyses highlight temporal synchronization of viral gene expression without resulting in immune gene induction, suggesting that no conflicts remain between ancient symbiotic partners when benefits to them converge., Jeremy Gauthier et al. present the chromosome scale assembly of the genome of the parasitic wasp C. congregata and show that bracovirus genes have colonized all ten chromosomes. Comparison with genome scaffolds of another wasp reveals a striking stability of these regions over ~53 million years, suggesting strong evolutionary constraints.

International audience; The population dynamics of annual social hymenoptera such as vespids depend largely on the fertility of the foundresses, which, in turn, is a key factor in the context of biological invasions. The native European hornet Vespa crabro (Vc) and the invasive Asian hornet Vespa velutina (Vv) have generally similar ecological traits, e.g. nesting and feeding habits, although they differ in colony size, which is higher in Vv. Furthermore, in contrast to Vc, Vv is more specialized in its predatory habits, intensively hunting honey bees at the hive. Comparing the morphological and reproductive traits of two closely related species occupying the same ecological niche, one of which is a native species and the other an alien, can help us to gain an understanding of the invasion process. To this end, we here compare reproductive (ovarian size and maturation, fat level, spermatheca size and sperm stock, fecundity) and morphological traits (head size, weight) of the foundresses of these two hornet species. We observed that ovarian maturation began approximately one month earlier in Vv than in Vc, and that the fat level in the former was lower. We found twice the number of sperm in the mated foundresses of Vv than in those of Vc (more than 100 x 10(3) and less than 50 x 10(3) sperm, respectively), in a 16% smaller spermatheca in Vc. Furthermore, the sperm of Vv was found to be 65% shorter than that of Vc. The precocity and higher potential fecundity of Vv queens may have favoured this species over Vc in terms of predatory behaviour, and thereby contributed to its invasiveness.

Understanding reproduction is essential for controlling pests and supporting beneficial insects. Among the many factors allowing optimal reproduction, sperm availability is key to sex ratio control in hymenopteran parasitoids since males are haploid and only females come from fertilization. Microplitisrufiventris (Hymenoptera; Braconidae) is a solitary endoparasitoid of some noctuids. This insect could be used for the control of the cotton leafworm Spodopteralittoralis. Under controlled conditions, sperm quantity was measured in virgin males at 1, 5, 10, and 15 days; it increases in adult males until the fifth day. Sperm stock of control males increased from 2500 at one day to 6700 at 15 days. With the control climatic condition being 25 °C, we tested the effects of a time-limited increase of temperature that can be found in Egypt (36 and 40 °C) during one day at the early pupal stage. Emerging males had 1500 and 420 sperm at 36 and 40 °C, respectively; both lived shorter than the control. The sperm potential of males is dependent on both age and temperature during the early pupal stage. It could have dramatic consequences on the sex ratio of M. rufiventris in natural and controlled populations.

Vespa velutina var nigrithorax (Lepelletier, 1835) is an invasive predator of bees accidentally introduced in France in 2004, and it is having a serious impact on apiculture and ecosystems. Studying the reproduction of an invasive species is key to assess its population dynamic. This study explores the sexual maturation of V. velutina males and the evolution of their fertility. The main studied parameters were physiologic (spermiogenesis, spermatogenesis) and anatomic (testes size and structure, head width). Two populations of males were described based on their emergence period: early males in early summer or classic males in autumn. Each testis has an average of 108 testicular follicles. Spermatogenesis is synchronous, with only 1 sperm production wave, and completed, on average, at 10.3 d after emergence with the degeneration of the testes. The sperm counts in seminal vesicles of mature males are 3 × 106 in October/November and 0.8 × 106 in June. In comparison, females store 0.1 × 106 sperm in their spermathecae. The early males emerged from colonies made by fertilized queens. The reproductive potential of these early males seemed limited, and their function in the colony is discussed. The sperm stock evolution in autumn males suggests the occurrence of a reproductive pattern of male competition for the access to females and a single copulation per male. The synchronicity of male and foundress emergences and sexual maturation is of primary importance for the mating success and the future colony development.

International audience; In insects, decreased reproduction is a sublethal consequence of high temperatures, with males being more sensitive to this in many species. In hymenoptera, arrhenotokous parthenogenesis means that female offspring are produced using sperm and are thus diploid, while males are haploid. Consequently, sperm stocks in males and females (after copulation) are a key regulator of the sex ratio. Anisopteromalus calandrae is a parasitoid wasp in which males can suffer from subfertility due to a drastic decrease in sperm count after exposure to high temperatures during a critical early pupal stage. However, in this species spermatogenesis continues during adulthood, therefore the heat sensitivity of adult males remains to be studied. Laboratory studies were conducted on virgin and previously mated young adult males under control (30°C) and heat shock (10 min at 48°C) conditions to exhaust their initial sperm stock. After heat shock, in both virgin and already mated males, the individual sperm potential was half that of controls. Both groups continuously produced sperm, but sperm stock of heat shocked males' never reached that of the controls. Heat shock reduced survival at 10 days only in previously experienced males but had no impact on the mating ability in competition for a female compared to controls. Despite a reduced sperm count, heat shocked males had fully fertile spermatozoa. Such a physiological response to heat shock in a species with continuous sperm production could be of major interest for both wild populations in a context of temperature variations and parasitoid wasps introduced for agronomical purposes.

International audience; Insect reproduction is influenced by various external factors including temperature, a well‐studied constraint. We investigated to what extent different levels of sperm limitation of males exposed to different heat stresses (34 and 36 °C) affect females’ offspring production and sex allocation in Nasonia vitripennis. In this haplodiploid parasitoid wasp attacking different species of pest flies, we investigated the effect of the quantity of sperm females received and stored in their spermatheca on their sperm use decisions, hence sex allocation, over successive ovipositions. In particular, we compared the sex allocation of females presenting three levels of sperm limitation (i.e., mated with control, 34 °C heat‐stressed or 36 °C heat‐stressed males) on each host they parasitized. To disentangle the potential reduction of sperm quality after a heat stress exposure from that of sperm quantity, we also explored the clutch size and sex ratio produced by females that were partially sperm limited after copulating with multiply mated males. Independently of their sperm numbers, all types of females produced a similar total number of offspring, but the more limited ones had fewer daughters. Sperm limitation further affected the distribution of daughters’ production across time. In addition to constraints acting on female physiology, male fertility should therefore be considered in studies measuring reproductive outputs of insects submitted to heat stresses.

The spermatozoon of the parasitoid wasp Cotesia congregata is an extremely short gamete measuring less than 7 μm; it is as yet the shortest flagellated sperm to be identified. The mature sperm consists of an acrosome, surrounded by an extra cellular coat, a condensed nucleus, two uncoiled mitochondrial derivatives and a short axoneme. Testes of young adults contain a continuum of differentiation stages. Initially, the flagellum is approximately 5 μm long. It conserves its length in round, elongated and mature spermatids, but is reduced to less than 3 μm in mature spermatozoa. The nucleus is 2 μm in diameter when round, 10 μm long when it becomes a long boat-hull shaped filament, and then reduces to 3.6 μm. Thus, during development the gamete reaches a total length of 15 μm before finally reducing to less than half that length. Some traits of mature sperm anatomy are similar to related species of the Braconidae family, but others seem to be specific and could be due to the shortness of the cell. This uncommon elongation and subsequent shortening of such a tiny flagellated cell constitutes a model for both nucleus and cilium development.

Vespa velutina var nigrithorax (Lepelletier, 1835) is an invasive predator of bees accidentally introduced in France in 2004, and it is having a serious impact on apiculture and ecosystems. Studying the reproduction of an invasive species is key to assess its population dynamic. This study explores the sexual maturation of V. velutina males and the evolution of their fertility. The main studied parameters were physiologic (spermiogenesis, spermatogenesis) and anatomic (testes size and structure, head width). Two populations of males were described based on their emergence period: early males in early summer or classic males in autumn. Each testis has an average of 108 testicular follicles. Spermatogenesis is synchronous, with only 1 sperm production wave, and completed, on average, at 10.3 d after emergence with the degeneration of the testes. The sperm counts in seminal vesicles of mature males are 3 × 10

International audience; Male fitness depends on the number of lifetime progeny of their mates and could be constrained by the chance of finding a mate, lifespan and temporal patterns of sperm production and allocation. Here, we used the parasitic wasp Nasonia vitripennis with a two-week lifespan and a gregarious lifestyle, to analyze how the reproductive system is organized to allocate spermatozoa over consecutive matings. Results show that spermatogenesis is synchronized and completed one day before emergence so that males emerge with a full sperm complement. We also found a regulation of spermatozoa transfer between testis and seminal vesicles that allows males to partition small ejaculates over multiple matings. Overall, this study shows that for N. vitripennis, male fertilization potential is determined (1) at the pupal stage, when spermatogenesis takes place to generate a complete life-long stock, (2) on emergence, when transport of spermatozoa from testes to seminal vesicles is initiated and (3) in adulthood, during which spermatozoa are partitioned over successive copulations. Such life history-traits are consistent with the gregarious lifestyle of N. vitripennis.

The reproductive capacity of the parasitoïd of cowpea bruchids Anisopteromalus calandrae Howard (Hymenoptera: Pteromalidae) male depends on the number of spermatozoa produced and spilled in the seminal vesicle. The objective of this study is to determine the dimensional characteristics of the different parts of the male genital tract of A. calandrae depending on age. Photographs of the parts of the male genital tract of the nymph and the virgin adults of 0- 1- 5- 10- and 15-day old were taken by light microscopy. The dimensional characteristics of the organs of the genital tract of A. calandrae has shown that when the surface of the testes decreases, that of the anterior portion of the seminal vesicle increases and the ratio between the width of the light and the total width of the accessory gland increases from emergence to the fifteenth day. The dimension of the testes decreases in favor of that of the seminal vesicle and the accessory gland of A. calandrae male depending on age.

International audience; The effects of a herbicide – Paraquat – on male development and reproduction were tested on the parasitoid wasp Anisopteromalus calandrae (Hymenoptera, Pteromalidae) by injecting host larvae with different concentrations of this substance. Data measured were: (1) developmental success, (2) sperm stock in seminal vesicles, (3) ability to copulate and transfer sperm, and (4) offspring production. Both developmental success and sperm in seminal vesicles were reduced in the ‘‘Paraquat” groups. However, neither sperm stored in the females' spermatheca, nor offspring production (number of female offspring and sex ratio) differed from controls, whatever the host treatment. The decreased of male sperm reserve in ‘‘Paraquat” group is likely to reduce their reproductive success because they can succefully inseminated less female than control males. These males are thus hyporfertiles. Because Paraquat has non-negligible consequences on non-target parasitoid males, it is likely to affect natural and controlled insect populations.

International audience; Post-copulatory paternity biases after female multiple mating are major constraints on both male and female reproductive systems. The outcome of paternity in certain situations is only controlled directly by male sperm stock. This was tested experimentally in the parasitoid wasp Anisopteromalus calandrae (Howard) (Hymenoptera: Pteromalidae), in which sperm stocks are small (several hundred) and the fertilizing efficiency of stored sperm is high (the ratio of sperm stored/fertilized eggs is about 0.75). Sperm in seminal vesicles and paternity of males of different status (virgin young, virgin old, or young previously mated) were measured after female single and double mating. The amount of sperm in the seminal vesicle differed according to male status (increasing from previously mated males to old males), but there was no difference in sperm stored by females after a single mating. In double mating experiments with two males of different status, paternity increased linearly with the relative amount of sperm in seminal vesicles. Paternity distribution conforms to ‘a fair raffle' of sperm from both donors following complete mixing of sperm prior to fertilization. Thus, in a female multiple mating context, male fitness depends principally on their sperm stock, which in turn depends on life history parameters, such as age and previous mating.

Eupelmus vuilleti is a primary and solitary ectoparasitoid of the larval stages of Bruchids (Callosobruchus maculatus, Bruchidius atrolineatus). In a context of intense competition for healthy hosts, E. vuilleti displays ovicide and larvicide behaviours towards the Pteromalid D. basalis during its development (kleptoparasitism), and in an extreme expression of kleptoparasitism the E. vuilleti females hyperparasitize the final larval stage (L5 stage) of D. basalis. In this study, we compared the variability of reproductive success in males that had developed in the context of hyperparasitism to that in males that had developed on primary hosts. The adaptation capacity of the males when 24 h old was analysed in terms of their weight, of the quantity of spermatozoids stored in the seminal vesicles, of the quality of insemination determined from the quantity of spermatozoids stored in the spermatheca of the females after the first mating, and of the number of daughters produced. Adults of E. vuilleti, the larvae of which had developed as hyperparasitoids, are smaller than those that have developed on primary hosts, but they keep all the abilities required to parasite a population of primary hosts once the competitive pressure is reduced.

The number of spermatozoa that a male transfers to the female during copulation is a main component of its individual fitness, especially under the pressure of sperm competition. This paper presents experimental results on the direct relationship between the male's sperm investment and its paternity in the offspring of dual-mated females. An eye colour mutant (red-eyed) is used to study the differences in the mating and fertilization abilities of males through observation of single and dual matings of females in Anisopteromalus calandrae (Hymenoptera, Chalcidoidea, Pteromalidae). Experimentally, females accept dual matings only in the simultaneous presence of two males. Counts of spermatozoa in the seminal vesicles of virgin males show that red-eyed males have more sperm than wild-eyed ones (approximately 1.46-fold greater). Red- and wild-eyed males do not differ in their mating behaviour and females mate indifferently with both phenotypes. Compared with once-mated females, double-mated females increase neither sperm storage nor lifetime fecundity, and the offspring sex ratio is female-biased. Females mated with two males of different phenotypes produce offspring of both phenotypes throughout their reproductive life, whatever the order of males in the copulation sequence. Any mating pattern appears to produce more red- than wild-eyed offspring (between 1.45- and 1.88-fold greater). Thus, proportions of offspring of each male match the proportions of their sperm potential. With no preference of female for red-eye or wild-eye males being demonstrated at either behavioural or physiological levels, a male's investment in sperm quantity appears to determine its individual reproductive success, at least in these experimental conditions.

Sperm quantity and quality during storage may be constraints acting on female fecundity and hence fitness. In Hymenoptera, the importance of sperm quality has rarely been considered, despite its central role in reproductive strategies and especially in sex ratio control. In these insects, fertilized eggs develop into females and unfertilized eggs into males. Experiments were conducted on the female wasp, Dinarmus basalis, in the laboratory with and without egg-laying resources (hosts). The first point was to test if sperm age influenced sperm storage by measuring sperm count and viability using a sperm viability test (SYBR-14 : propidium iodide). The second point was the influence of prolonged storage in the female genital tract on the quantity, sex ratio and fitness of offspring produced. Results show that sperm viability in the spermatheca does not change significantly with maternal age, and that the sperm stock is not affected when females are deprived of hosts. Egg-laying is gradually restored after 21 days of host deprivation but remains at a low level after 115 days. The fitness of mated D. basalis females is therefore not constrained by sperm quantity or quality and seems to depend on host availability and female age.

The quality of a sperm population can be characterized physiologically and its fecundity predicted by its viable : non-viable sperm ratio. To improve the knowledge of reproductive strategies in two ectoparasitoid hymenopteran species, Eupelmus orientalis Crawford (Hymenoptera: Eupelmidae) and Dinarmus basalis Rondani (Hymenoptera: Pteromalidae), the assessment of sperm viability using the dual fluorescence staining procedure SYBR-14 : propidium iodide was developed. The aim of the study was to provide a comparative test in vitro applicable to both sexes to study the evolution of sperm quality at various stages of the reproductive processes. The reliability of propidium iodide to detect non-viable sperm (stained in red) was confirmed in both species on the basis of two stress tests (ethanol and Triton X-100) but our study also revealed that propidium iodide concentrations must be adequately adjusted for each single species. This experiment also demonstrated the physiological heterogeneity of sperm populations in E. orientalis and D. basalis males and females. In both species, 40% of the sperm in the seminal vesicles was found to be non-viable. By contrast with E. orientalis, the populations of non-viable sperm estimated from the seminal vesicles of D. basalis were found to be strongly different from those observed in the spermatheca. From the present results, the population of viable sperm detected in the spermatheca of females from both species proved a reliable predictor of fertilization achieved in ovipositing females.

Multiple mating and its effects on the sex ratio in Dinarmus basalis (Hymenoptera: Chalcidoidea, Pteromalidae), an ectoparasitoid of Callosobruchus maculatus (Coleoptera: Bruchidae), were investigated under controlled conditions. Once-mated females suffer a sperm depletion about 21 days after mating and thereafter are constrained to produce only haploid males. On the other hand, three-times-mated females store more sperm in their spermathecae (335 vs 147) and produce daughters during the major part of their reproductive life. Consequently, once-mated females showed a male-biased life time sex ratio (mean = 0.31) as opposed to three-times-mated females (mean = 0.63). Females can copulate only at the beginning of their reproductive life, and multiple mating must occur before egg-laying activity. This behavioral strategy could be an avoidance of consanguinity in a promiscuous environment. This ectoparasitoid species reveals a reproductive strategy which promotes polyandry and a very high sperm efficiency.

In Eupelmus orientalis and Dinarmus basalis, two ectoparasitoid species of Callosobruchus maculatus larvae, pre-imaginal development occurs within a leguminous seed, and is then impossible to observe directly. Offspring sex ratio is normally determined when adults emerge, and mortality during pre-imaginal stages remains unknown. By using translucent gelatine capsules containing a host larva to mimic the seed, we conducted an experimental study of offspring production and sex ratio with measurements of growth and pre-imaginal mortality. Mated females were allowed to lay eggs on hosts in seeds or in gelatine capsules. In the gelatine capsules the parasitoid was observed daily and measured to assess if the offspring sex ratio can be determined before emergence of adults. When seeds are replaced with gelatine caps, the number of egg-laying females decreases (48% in Eupelmus orientalis and 72% in Dinarmus basalis of females ovipositing on seeds lay eggs on gelatine capsules), and the offspring output changes qualitatively in Eupelmus orientalis (egg numbers constant, sex ratio diminishing from 0.75 to 0.46) and quantitatively in Dinarmus basalis (egg numbers diminishing, sex ratio maintained about 0.7). In Eupelmus orientalis, pre-imaginal mortality occurs principally in the first three clays, the critical stage is hatching, and the final mortality is 30.1%. In Dinarmus basalis, mortality of larvae increases gradually from the 2nd to the 7th day, the final mortality being 13.6%, (3) In both species sex of the offspring can be determined before emergence but only after most mortality has occurred.

Proofs of concept have shown that chromosomal gene clusters encoding ribosomal RNA (rRNA) constitute gene delivery integration loci that are optimal for transgene expression. However, because homologous recombination is efficient to integrate DNA segments into these genes in animals, new molecular tools are required to construct systems able to target molecules in the immediate vicinity of the rRNA genes. We investigated the properties of several DNA binding domains (DBDs) able to recognize specifically a motif within a 100-bp region of the rRNA genes that is 99-100% conserved among eukaryotes. Our findings demonstrate that two Myb-like DBDs originating from the endonucleases encoded by R2 non-LTR retrotransposons are promising candidates since they i) specifically recognize, with high affinity, a 20-bp binding site located within the expected genomic rDNA target, ii) act as monomers, iii) contain a nuclear localization signal, iv) remain functional when fused to another domain and, v) do not alter the functionality of the protein to which they are fused. However, results obtained in vivo with several R2DBD fusions reveal that two properties remain to be engineered before these DBDs can be integrated into a molecular targeting system directed into rRNA genes. The first concerns the ability of R2DBD to locate within the nucleolus, the organelle in which the rRNA genes reside. The second is the tendency of R2DBD to accumulate in certain parts of the nuclei, which limits its diffusion within nuclei. Solutions are discussed to circumvent these current limitations.Our results supply important information concerning the R2DBD properties and the targeting of plasmid DNA within nuclei. They will need to be further analyzed from three aspects; the unexploited advantages of the R2DBDs, the possibilities and limitations of fusion peptides for targeting integrations of non-viral vector, and the alternatives to fusion peptides for targeting vectors.

Many studies have reported an increase in male reproductive disorders due to the environment. Despite their ecological importance invertebrates have been poorly studied; they are yet affected in the same way as vertebrates by changes in the environment. Different stresses made on parasitic wasps are able to induce a male subfertility. These insects are easy to rear in the laboratory, used to control insect pests, and have other benefits presented in this article. Parasitic wasps are good models for understanding the mechanisms inducing subfertility and may provide new areas for research in vertebrates.

National audience; In insects, reproductive success and survival are affected by temperature. Reproduction is more sensitive than other physiological traits. While the effects of heat stress on females are well known, the effects on males are less clear. Hymenopteran parasitoids are valuable for studying the consequences of heat stress on male reproduction. In these species, through arrhenotoquous parthenogenesis, the sex ratio of the offspring is directly dependent on the sperm stock acquired by females during copulation. In the lab, heat temperature treatments (32–44 °C) were applied for 3 days in the pupal stage of Anisopteromalus caland-rae males, and development was completed at 30 °C. Three different effects were observed depending on the temperature: mortality above 42 °C, sterility of emerging males at 40 °C, and sub-fertility at 38 °C. This sub-fertility is characterized by a dramatic decrease in male sperm supplies, of up to 7% compared to control males. In the course of ageing, the sperm stock of sub-fertile males increases but never reaches the level of control males. Survival was significantly higher in control (30 °C) males than those treated at 38 °C. Male mating ability was similar whatever the treatment (control and 38 °C), but females mated with 38 °C-treated males stored 100 times less sperm on average than those mated with control males. The offspring sex ratio of females mated with 38 °C-treated males was strongly male biased. The physiological mechanisms are as yet unknown. The relationship between temperature, sperm stock and sex ratio should be taken into account in the management of parasitoids for integrated pest management.

International audience; De nombreuses études ont rapporté une augmentation des troubles de la reproduction mâle dus à l'environnement. Malgré leur importance, les invertébrés ont fait l'objet de peu de travaux ; ils sont pourtant affectés, au même titre que les vertébrés, par les changements de l'environnement. Différents stress appliqués à des hyménoptères parasitoïdes sont capables d'induire une hypofertilité mâle. Ces insectes, faciles à élever en laboratoire et utilisés en agronomie dans le contrôle des populations d'insectes ravageurs de cultures ou de récoltes, présentent également d'autres avantages, décrits dans cet article. Ce sont de bons modèles pour comprendre les mécanismes induisant l'hypofertilité et ouvrir peut-être de nouvelles pistes de recherche chez les vertébrés

Drosophilidae (Diptera) are well known for tremendous variability in sperm length as great as that found among vertebrates and invertebrates together. Paradoxically, the great length (> 1 mm) of Drosophila sperm creates difficulties for measuring them. Evidence is provided that the length of giant sperm can be indirectly, albeit reliably, estimated using the regression lines between sperm and testis or receptacle lengths, established from 26 species with short- and mid-length sperm. Using this method, the sperm length in Drosophila hydei was estimated to be about 17 mm, which would be the longest in the animal kingdom. The phylogenetic relationships between species are discussed.

Mariner-like elements (MLEs) are widespread transposable elements in animal genomes. They have been divided into at least five sub-families with differing host ranges. We investigated whether the ability of transposases encoded by Mos1, Himar1 and Mcmar1 to be actively imported into nuclei varies between host belonging to different eukaryotic taxa. Our findings demonstrate that nuclear importation could restrict the host range of some MLEs in certain eukaryotic lineages, depending on their expression level. We then focused on the nuclear localization signal (NLS) in these proteins, and showed that the first 175 N-terminal residues in the three transposases were required for nuclear importation. We found that two components are involved in the nuclear importation of the Mos1 transposase: an SV40 NLS-like motif (position: aa 168 to 174), and a dimerization sub-domain located within the first 80 residues. Sequence analyses revealed that the dimerization moiety is conserved among MLE transposases, but the Himar1 and Mcmar1 transposases do not contain any conserved NLS motif. This suggests that other NLS-like motifs must intervene in these proteins. Finally, we showed that the over-expression of the Mos1 transposase prevents its nuclear importation in HeLa cells, due to the assembly of transposase aggregates in the cytoplasm.

Any hitherto available technique used to detect the presence of Wolbachia, a rickettsia-like endosymbiont of Drosophila simulans, i.e., crossing with tester strains, electron microscope observations, DAPI-staining of embryo, or amplification of specific sequences by PCR are notably time-consuming techniques. Staining sperm cysts with the fluorochrome DAPI is here shown to be a fast and reliable technique to detect the symbiont and evaluate the level of infection of individual males. Wolbachia cells are abundant in sperm cysts and make up a mass which is eliminated in the waste bag during spermatogenesis. Such a mass was absent in all those strains known to be aposymbiotic. In infected strains, tetracycline treatment led to the disappearance of the symbionts within sperm cysts. In aging males, a decrease in the proportion of infected cysts were observed. There was however no apparent change in the number of symbionts in the cysts which remained infected. The reduction of reproductive incompatibility, which is observed in older males, may be explained by this process, i.e., a decrease in the frequency of infected cysts.

In order to test the validity of the prediction of the mating pattern of females from the sperm length distribution in males, three species of Drosophila were analysed. Males in the three species are equally polygynous but females differ in the level of polyandry. A 'low recurrence polyandry' is observed in the sperm dimorphic species D. affinis while a 'high recurrence polyandry' is observed in the sperm monomorphic species D. latifasciaeformis and D. littoralis. These results are consistent with the hypothesis proposed previously that sperm dimorphism in males can only be maintained by a selective alternative in females (i.e. facultative female polygamy), whereas a stricter mating system (e.g. 'obligatory' polyandry) should only result in sperm monomorphism irrespective of the absolute value of sperm length.

International audience; Hymenoptera are haplodiploid insects, consequently sex ratio depends on female's sperm management which itself arises from the reproductive capacity of neighbouring males. To study the influence of ageing on male reproductive potential, laboratory experiments were conducted on Dinarmus basalis (Hymenoptera, Pteromalidae) males, a tropical wasp in which sperm counts are known to constrain sex ratio. Two groups of virgin males were compared: 1-day and 30-days old. Parameters recorded were sperm quantity and viability in seminal vesicles, shape of testis, mating ability in both individual and competitive situations and sperm stored by females after male multiple mating. Older males had twice as much sperm as young males, but their reproductive capacities did not differ. They were able to copulate with 20 successive virgin females in a short period. Sperm stored in spermathecae decreased with female mating order. In competition, old and young males had the same access to females. The difference between old and young males was visible at the level of reproductive tract: young males have functional testis and old males have empty non-functional testis. Spermatozoa are kept viable in male seminal vesicles for long periods. In this species, the reproductive potential of males is not altered by ageing. At the population level this may represent an adaptation for maintaining continuous reserves of sperm at the disposal of females.

International audience; In recent years, several studies have shown a decline in reproductive success in males in both humans and wildlife. Research on male fertility has largely focused on vertebrates, although invertebrates constitute the vast majority of terrestrial biodiversity. The reduction of their reproductive capacities due to environmental stresses can have strong negative ecological impacts, and also dramatic consequences on world food production if it affects the reproductive success of biological control agents, such as parasitic wasps used to control crop pests. Here Nasonia vitripennis, a parasitic wasp of various fly species, was studied to test the effects of 24h-heat stress applied during the first pupal stage on male fertility. Results showed that only primary spermatocytes were present at the first pupal stage in all cysts of the testes. Heat stress caused a delay in spermatogenesis during development and a significant decrease in sperm stock at emergence. Females mated with these heat-stressed males showed a reduce sperm count stored in their spermatheca. Females did not appear to distinguish heat-stressed from control males and did not remate more frequently to compensate for the lack of sperm transferred. As a result, females mated with heat-stressed males produced a suboptimal lifetime offspring sex ratio compared to those mated with control males. This could further impact the population dynamics of this species. N. vitripennis appears to be an interesting biological model to study the mechanisms of subfer-tility and its consequence on female reproductive strategies and provides new research perspectives in both invertebrates and vertebrates.

The effect of host size on male fitness was tested in the parasitoid wasp Dinarmus basalis (Hymenoptera, Pteromalidae) using hosts of different fresh weight. Fitness was measured as the sperm stock in seminal vesicles, and the ability to access females in single or competition situations. Both body size and sperm in seminal vesicles increased with host fresh weight. Males from small hosts had a reduced size and sperm stock compared to those from larger hosts. In single situations, males from both small and large hosts had similar reproductive capacities, whereas in multiple mating or competition situations, males from small hosts were at a disadvantage, inseminating fewer females and copulating less frequently. However, females did not appear to choose between males, and no effect on sperm stored in the spermatheca was observed. Being small does not prevent a D. basalis male mating and producing progeny in single situations, although more offspring could be expected from larger males because of their better competitive abilities.

Eupelmus vuilleti is a primary and solitary ectoparasitoid of the larval stages of Bruchids (Callosobruchus maculatus, Bruchidius atrolineatus). In a context of intense competition for healthy hosts, E. vuilleti displays ovicide and larvicide behaviours towards the Pteromalid D. basalis during its development (kleptoparasitism), and in an extreme expression of kleptoparasitism the E. vuilleti females hyperparasitize the final larval stage (L5 stage) of D. basalis. In this study, we compared the variability of reproductive success in males that had developed in the context of hyperparasitism to that in males that had developed on primary hosts. The adaptation capacity of the males when 24 h old was analysed in terms of their weight, of the quantity of spermatozoids stored in the seminal vesicles, of the quality of insemination determined from the quantity of spermatozoids stored in the spermatheca of the females after the first mating, and of the number of daughters produced. Adults of E. vuilleti, the larvae of which had developed as hyperparasitoids, are smaller than those that have developed on primary hosts, but they keep all the abilities required to parasite a population of primary hosts once the competitive pressure is reduced.

The male reproductive tracts of Drosophila display considerable variation in the relative size of their components, notably of the testes, but there are few structural differences between species. Here we report a remarkable coiled structure separating the testes from the seminal vesicles in the giant sperm species Drosophila bifurca. This evolutionary novelty, known as the 'sperm roller', seems to be an exaggeration in the size of the testicular duct as revealed by light and electron microscopic observations. It consists of a tubular monocellular epithelium lying on the basal laminae and muscle and conjunctive cells. The lumen of the roller contains crypts. The apical membrane of the epithelial cells presents numerous long microvilli protruding into the lumen. The sperm roller structure is probably involved in managing sperm during their transit through the male genital tract, because sperm are seen in bundles at the base of the testis, whereas they are singly rolled up by the time they enter the seminal vesicles. The hypercoiling of the individual spermatozoon within the roller probably occurs as the result of an osmotic process produced by features of the epithelial wall and the dramatically increased exchange surface. This is the first report of a specialized device of this type in Drosophila or, more generally, in insects.

Sperm stocks in both males and females of the parthenogenetic wasp Eupelmus orientalis were investigated at various points during reproduction and compared to the progeny of females in controlled conditions. One day-old virgin males had approximately 5500 sperm, and from a total of about 1697 sperm transferred per copulation, 21% are stored in the spermathecae by females 24 hours after mating. At the end of the egg-laying period (at least 42 days), 2/5 of the initial amount of sperm remained in this storage organ. This decrease (from approximately 350 to 150) occurred essentially during the first 21 days of egg-laying activity, indicating that the majority of sperm stored were used during this period. Between 21 days and the end of fertile life, the number of sperm remained constant. The mean offspring production throughout reproductive life after one mating was 153, with 56.5% of the daughters laid at the beginning of the laying activity. Sex ratio was entirely female biased during the first 15 days (mean=0.65), then it decreased and became nearly equal after 20 days. Present results propose that females maximize the production of daughters i.e. of inseminated eggs until the 20th day and after this time lay as many daughters and sons despite their still having stored sperm. Physiological constraints due to ageing are proposed to explain this phenomenon.

It is generally assumed that sexes in animals have arisen from a productivity versus provisioning conflict; males are those individuals producing gametes necessarily small, in excess, and individually bereft of all paternity assurance. A 1- to 2-cm sperm, 5-10 times as long as the male body, might therefore appear an evolutionary paradox. As a matter of fact, species of Drosophila of the Drosophila subgenus differ from those of other subgenera by producing exclusively sperm of that sort. We report counts of such giant costly sperm in Drosophila littoralis and Drosophila hydei females, indicating that they are offered in exceedingly small amounts, tending to a one-to-one gamete ratio after a single mating. As a result, most of them are successfully involved in a fertilization. Hence, the concept of "paternity assurance of individual sperm" arises. Evidence is further provided here that almost the entire sperm is incorporated into the egg during fertilization. Labeling with specific antibodies in fertilized eggs reveals intact axonemes up to late gastrulation. The question, then, is why selection has favored such an unusual strategy. Explanations related to some prefertilization functions are ruled out. It is therefore tentatively proposed that virtually every giant sperm constitutes a "direct paternal legacy to the embryo," which, in contrast to any male-derived nuptial gift, cannot be minimized by female remating. We suggest that dramatic shortage of giant sperm with a high prospect of fusion and increased zygote provisioning is merely another way of being anisogamous.

All species of the Drosophila obscura group exhibit within-ejaculate sperm length dimorphism. The present work is a contribution to the understanding of sperm competition through a comparative study of sperm kinetic parameters in four of these species. Videomicrographic observations at 200 frames per second of sperm from males and females, out of the storage organ, prior or after storage were made. Drosophila sperm display both major and minor waves. The former is analysed by measuring coiling diameter (micron) and the latter by recording both beat frequency (s-1) and wave propagation velocity (micron.s-1). Results show that the 'behaviour' of short and long spermatozoa noticeably differ: short sperm kinetics remains unaltered after storage while both major and minor waves of long spermatozoa are markedly modified. Thus, evidence is provided here of a sort of "differential activation" which is assumed to result in different survival abilities of short and long sperm within the storage organ of females.