Unlocking the "Black box": internal female genitalia in Sepsidae (Diptera) evolve fast and are species-specific

Background: The species-specificity of male genitalia has been well documented in many insect groups and sexual selection has been proposed as the evolutionary force driving the often rapid, morphological divergence. The internal female genitalia, in sharp contrast, remain poorly studied. Here, we present the first comparative study of the internal reproductive system of Sepsidae. We test the species-specificity of the female genitalia by comparing recently diverged sister taxa. We also compare the rate of change in female morphological characters with the rate of fast-evolving, molecular and behavioral characters. Results: We describe the ectodermal parts of the female reproductive tract for 41 species representing 21 of the 37 described genera and define 19 morphological characters with discontinuous variation found in eight structures that are part of the reproductive tract. Using a well-resolved molecular phylogeny based on 10 genes, we reconstruct the evolution of these characters across the family 120 steps; Consistency Index (CI): 0.41]. Two structures, in particular, evolve faster than the rest. The first is the ventral receptacle, which is a secondary sperm storage organ. It accounts for more than half of all the evolutionary changes observed (7 characters; 61 steps; CI: 0.46). It is morphologically diverse across genera, can be bi-lobed or multi-chambered (up to 80 chambers), and is strongly sclerotized in one clade. The second structure is the dorsal sclerite, which is present in all sepsids except Orygma luctuosum and Ortalischema albitarse. It is associated with the opening of the spermathecal ducts and is often distinct even among sister species (4 characters; 16 steps; CI: 0.56). Conclusions: We find the internal female genitalia are diverse in Sepsidae and diagnostic for all species. In particular, fast-evolving structures like the ventral receptacle and dorsal sclerite are likely involved in post-copulatory sexual selection. In comparison to behavioral and molecular data, the female structures are evolving 2/3 as fast as the non-constant third positions of the COI barcoding gene. They display less convergent evolution in characters (CI = 0.54) than the third positions or sepsid mating behavior (CICOI = 0.36; CIBEHAV = 0.45)

Bending for love: Losses and gains of sexual dimorphisms are strictly correlated with changes in the mounting position of sepsid flies (Sepsidae: Diptera)

10.1186/1471-2148-8-155BMC Evolutionary Biology81

Intraspecific mating system evolution and its effect on complex male secondary sexual traits: Does male–male competition increase selection on size or shape?

Sexual selection is generally held responsible for the exceptional diversity in secondary sexual traits in animals. Mating system evolution is therefore expected to profoundly affect the covariation between secondary sexual traits and mating success. Whereas there is such evidence at the interspecific level, data within species remain scarce. We here investigate sexual selection acting on the exaggerated male fore femur and the male wing in the common and widespread dung flies Sepsis punctum and S. neocynipsea (Diptera: Sepsidae). Both species exhibit intraspecific differences in mating systems and variation in sexual size dimorphism (SSD) across continents that correlates with the extent of male–male competition. We predicted that populations subject to increased male–male competition will experience stronger directional selection on the sexually dimorphic male foreleg. Our results suggest that fore femur size, width and shape were indeed positively associated with mating success in populations with male‐biased SSD in both species, which was not evident in conspecific populations with female‐biased SSD. However, this was also the case for wing size and shape, a trait often assumed to be primarily under natural selection. After correcting for selection on overall body size by accounting for allometric scaling, we found little evidence for independent selection on any of these size or shape traits in legs or wings, irrespective of the mating system. Sexual dimorphism and (foreleg) trait exaggeration is therefore unlikely to be driven by direct precopulatory sexual selection, but more so by selection on overall size or possibly selection on allometric scaling

Sexual selection accounts for the geographic reversal of sexual size dimorphism in the dung fly, Sepsis punctum (Diptera: Sepsidae)

Sexual size dimorphism (SSD) varies widely across and within species. The differential equilibrium model of SSD explains di- morphism as the evolutionary outcome of consistent differences in natural and sexual selection between the sexes. Here, we comprehensively examine a unique cross-continental reversal in SSD in the dung fly, Sepsis punctum. Using common garden labo- ratory experiments, we establish that SSD is male-biased in Europe and female-biased in North America. When estimating sexual (pairing success) and fecundity selection (clutch size of female partner) on males under three operational sex ratios (OSRs), we find that the intensity of sexual selection is significantly stronger in European versus North American populations, increasing with male body size and OSR in the former only. Fecundity selection on female body size also increases strongly with egg number and weakly with egg volume, however, equally on both continents. Finally, viability selection on body size in terms of intrinsic (physiological) adult life span in the laboratory is overall nil and does not vary significantly across all seven populations. Although it is impossible to prove causality, our results confirm the differential equilibrium model of SSD in that differences in sexual selection intensity account for the reversal in SSD in European versus North American populations, presumably mediating the ongoing speciation process in S. punctum

Size rather than complexity of sexual ornaments prolongs male metamorphosis and explains sexual size dimorphism in sepsid flies

Male sexual ornaments often evolve rapidly and are thought to be costly, thus contributing to sexual size dimorphism. However, little is known about their developmental costs, and even less about costs associated with structural complexity. Here, we quantified the size and complexity of three morphologically elaborate sexually dimorphic male ornaments that starkly differ across sepsid fly species (Diptera: Sepsidae): (i) male forelegs range from being unmodified, like in most females, to being adorned with spines and large cuticular protrusions; (ii) the fourth abdominal sternites are either unmodified or are converted into complex de novo appendages; and (iii) male genital claspers range from small and simple to large and complex (e.g. bifurcated). We tracked the development of 18 sepsid species from egg to adult to determine larval feeding and pupal metamorphosis times of both sexes. We then statistically explored whether pupal and adult body size, ornament size and/or ornament complexity are correlated with sex-specific development times. Larval growth and foraging periods of male and female larvae did not differ, but the time spent in the pupal stage was ca 5% longer for sepsid males despite emerging 9% smaller than females on average. Surprisingly, we found no evidence that sexual trait complexity prolongs pupal development beyond some effects of trait size. Evolving more complex traits thus does not incur developmental costs at least in this system

Metabarcoding mosquitoes: MinION sequencing of bulk samples gives accurate species profiles for vector surveillance (Culicidae)

Mosquitoes (Family: Culicidae) are dominant vectors of pathogens, and their surveillance has been incorporated into major disease control programs worldwide. However, routine, species-level identification of mosquitoes is often a bottleneck for management, and Next Generation Sequencing (NGS) platforms and DNA metabarcoding can revolutionize this process. MinION nanopore technologies promise on-site sequencing and rapid sample processing rates ideal for time-sensitive biosurveillance. Here, we benchmark the results of DNA metabarcoding on the MinION against the Illumina MiSeq platform, which is known for its higher sequencing accuracy. We used metazoan COI mini-barcode primers to carry out DNA metabarcoding of mosquito bulk samples caught during a real vector survey, then compared the mosquito species profiles recovered on each sequencing platform. We also tested the influence of using different trap lures, storage methods, and pooling different specimen body parts on the number of species recovered. We report that mosquito species-level identifications were highly congruent between MinION and Illumina (93% overlap). We also find that CO2 gas cylinders outperformed biogenic CO2 sources significantly, by two-fold. Notably, we demonstrated the feasibility of detecting zoonotic reservoirs and pathogen signals from mosquito bulk samples. We present the first use of DNA metabarcoding on the MinION for vector surveillance and discuss future applications

Biomagnifcation and body distribution of ivermectin in dung beetles

We thank the staf of Doñana Biological Reserve (DBR-ICTS), Doñana National Park, and Los Alcornocales Natural Park, especially D. Paz, F. Ibáñez, P. Bayón, M. Malla and D. Ruiz for logistic facilities for the field work and permissions (2019107300000904/IRM/MDCG/mes) to collect cattle dung and dung beetles. We are grateful to J. Castro and A. Rascón for technical assistance. We also thank A. V. Giménez-Gómez for her technical assistance in the laboratory work. We thank also F.-T Krell and the two anonymous reviewers for their constructive comments. Financial support was provided by the project CGL2015-68207-R of the Secretaría de Estado de Investigación–Ministerio de Economía y Competitividad.A terrestrial test system to investigate the biomagnifcation potential and tissue-specifc distribution of ivermectin, a widely used parasiticide, in the non-target dung beetle Thorectes lusitanicus (Jekel) was developed and validated. Biomagnifcation kinetics of ivermectin in T. lusitanicus was investigated by following uptake, elimination, and distribution of the compound in dung beetles feeding on contaminated faeces. Results showed that ivermectin was biomagnifed in adults of T. lusitanicus when exposed to non-lethal doses via food uptake. Ivermectin was quickly transferred from the gut to the haemolymph, generating a biomagnifcation factor (BMFk) three times higher in the haemolymph than in the gut after an uptake period of 12 days. The fat body appeared to exert a major role on the biomagnifcation of ivermectin in the insect body, showing a BMFk 1.6 times higher than in the haemolymph. The results of this study highlight that the biomagnifcation of ivermectin should be investigated from a global dung-based food web perspective and that the use of these antiparasitic substances should be monitored and controlled on a precautionary basis. Thus, we suggest that an additional efort be made in the development of standardised regulatory recommendations to guide biomagnifcation studies in terrestrial organisms, but also that it is necessary to adapt existing methods to assess the efects of such veterinary medical products

First assessment of the comparative toxicity of ivermectin and moxidectin in adult dung beetles: Sub-lethal symptoms and pre-lethal consequences

Among macrocyclic lactones (ML), ivermectin (IVM) and moxidectin (MOX) potentially affect all Ecdysozoan species, with dung beetles being particularly sensitive. The comparative effects of IVM and MOX on adult dung beetles were assessed for the first time to determine both the physiological sub-lethal symptoms and pre-lethal consequences. Inhibition of antennal response and ataxia were tested as two intuitive and ecologically relevant parameters by obtaining the lowest observed effect concentration (LOEC) values and interpolating other relevant toxicity thresholds derived from concentration-response curves (IC50, as the concentration of each ML where the antennal response is inhibited by half; and pLC50, as the quantity of ingested ML where partial paralysis was observed by half of treated individuals) from concentration-response curves. Both sub-lethal and pre-lethal symptoms obtained in this study coincided in that IVM was six times more toxic than MOX for adult dung beetles. Values of LOEC, IC50 and pLC50 obtained for IVM and MOX evaluated in an environmental context indicate that MOX, despite needing more time for its elimination in the faeces, would be half as harmful to dung beetles as IVM. This approach will be valuable to clarify the real impact of MLs on dung beetle health and to avoid the subsequent environmental consequences

Reappraising Sexual Coevolution and the Sex Roles

Over recent decades, new ideas have radically altered how we see sex and reproduction. The implications of these ideas are still being explored, yielding intriguing discoveries