Impacts of Non-Lethal High-Temperature Stress on the Development and Reproductive Organs of Bradysia odoriphaga

Bradysia odoriphaga is an agricultural pest in China’s vegetable industry. In this study, pupae and adults were exposed to various non-lethal high-temperatures. The results demonstrated a decreased rate of eclosion once the pupae were exposed to temperatures exceeding 37 °C for 1 h. No effect on the lifespan of unmated female adults was observed after exposure to temperature stress, while unmated male adult lifespan decreased (>37 °C for 2 h). The size of the testis and ovaries for unmated male and female adults decreased, as did the fecundity and egg hatching rate for mated females. Compared with the control group (25 °C), the testis size of unmated male adults decreased after high-temperature stress followed by recovery at 25 °C for 1 h, though the size of the ovaries of female adults did not change. Additionally, the size of the testis and ovaries for unmated male and female adults decreased following high-temperature stress and 24 h of recovery at 25 °C. High temperatures affected males more than females; 37 °C is the critical temperature to control the population of B. odoriphaga. These results lay the foundation for the future development of environmentally friendly high-temperature prevention and pest-control strategies.

Hmga2 deficiency is associated with allometric growth retardation, infertility, and behavioral abnormalities in mice

The high mobility group AT-hook 2 (HMGA2) protein works as an architectural regulator by binding AT-rich DNA sequences to induce conformational changes affecting transcription. Genomic deletions disrupting HMGA2 coding sequences and flanking non-coding sequences cause dwarfism in mice and rabbits. Here, CRISPR/Cas9 was used in mice to generate the Hmga2 null allele that specifically disrupts only the coding sequence. The loss of one or both alleles of Hmga2 resulted in reduced body size of 20% and 60%, respectively, compared to wild-type littermates as well as an allometric reduction in skull length in Hmga2-/- mice. Both male and female Hmga2-/- mice are infertile, whereas Hmga2+/- mice are fertile. Examination of reproductive tissues of Hmga2-/- males revealed a significantly reduced size of testis, epididymis, and seminal vesicle compared to controls, and 70% of knock-out males showed externalized penis, but no cryptorchidism was observed. Sperm analyses revealed severe oligospermia in mutant males and slightly decreased sperm viability, increased DNA damage but normal sperm chromatin compaction. Testis histology surprisingly revealed a normal seminiferous epithelium, despite the significant reduction in testis size. In addition, Hmga2-/- mice showed a significantly reduced exploratory behavior. In summary, the phenotypic effects in mouse using targeted mutagenesis confirmed that Hmga2 is affecting prenatal and postnatal growth regulation, male reproductive tissue development, and presents the first indication that Hmga2 function is required for normal mouse behavior. No specific effect, despite an allometric reduction, on craniofacial development was noted in contrast to previous reports of an altered craniofacial development in mice and rabbits carrying deletions of both coding and non-coding sequences at the 5’part of Hmga2.

AXDND1, a novel testis-enriched gene, is required for spermiogenesis and male fertility

Spermiogenesis is a complex process depending on the sophisticated coordination of a myriad of testis-enriched gene regulations. The regulatory pathways that coordinate this process are not well understood, and we demonstrate here that AXDND1, as a novel testis-enriched gene is essential for spermiogenesis and male fertility. AXDND1 is exclusively expressed in the round and elongating spermatids in humans and mice. We identified two potentially deleterious mutations of AXDND1 unique to non‐obstructive azoospermia (NOA) patients through selected exonic sequencing. Importantly, Axdnd1 knockout males are sterile with reduced testis size caused by increased germ cell apoptosis and sloughing, exhibiting phenotypes consistent with oligoasthenoteratozoospermia. Axdnd1 mutated late spermatids showed head deformation, outer doublet microtubules deficiency in the axoneme, and loss of corresponding accessory structures, including outer dense fiber (ODF) and mitochondria sheath. These phenotypes were probably due to the perturbed behavior of the manchette, a dynamic structure where AXDND1 was localized. Our findings establish AXDND1 as a novel testis-enrich gene essential for spermiogenesis and male fertility probably by regulating the manchette dynamics, spermatid head shaping, sperm flagellum assembly.

Aurora a kinase (AURKA) is required for male germline maintenance and regulates sperm motility in the mouse

Aurora A kinase (AURKA) is an important regulator of cell division and is required for assembly of the mitotic spindle. We recently reported the unusual finding that this mitotic kinase is also found on the sperm flagellum. To determine its requirement in spermatogenesis, we generated conditional knockout animals with deletion of the Aurka gene in either spermatogonia or spermatocytes to assess its role in mitotic and postmitotic cells, respectively. Deletion of Aurka in spermatogonia resulted in disappearance of all developing germ cells in the testis, as expected given its vital role in mitotic cell division. Deletion of Aurka in spermatocytes reduced testis size, sperm count, and fertility, indicating disruption of meiosis or an effect on spermiogenesis in developing mice. Interestingly, deletion of Aurka in spermatocytes increased apoptosis in spermatocytes along with an increase in the percentage of sperm with abnormal morphology. Despite the increase in abnormal sperm, sperm from spermatocyte Aurka knockout mice displayed increased progressive motility. In addition, sperm lysate prepared from Aurka knockout animals had decreased protein phosphatase 1 (PP1) activity. Together, our results show that AURKA plays multiple roles in spermatogenesis, from mitotic divisions of spermatogonia to sperm morphology and motility.

Changes in histology, protein expression, and autophagy in dairy goat testes during non-breeding season

Seasonal reproduction contributes to increased chances of offspring survival in some animals. Dairy goats are seasonal breeding mammals. In this study, adult male Guanzhong dairy goats (10–12 months old) were used. Testis size, semen quality, hormone level, apoptosis of germ cells, and autophagy of Sertoli cells were analyzed in dairy goats during the breeding (October) and non-breeding (April) seasons. We found that, during the non-breeding season for dairy goats, semen quality, follicle-stimulating hormone (FSH) levels, and testosterone levels were reduced, and the number of apoptotic germ cells increased. The proliferation with decrease activity of germ cells in dairy goat during the non-breeding season was significantly affected. However, the testis size did not change seasonally. Interestingly, Sertoli cell autophagy was more active during the non-breeding season. The expression levels of FSH receptor (FSHR), wilms tumor 1 (WT1), androgen binding protein (ABP), glial cell derived neurotrophic factor (GDNF), and stem cell factor (SCF) decreased in dairy goats during the non-breeding season. In summary, our results indicate that spermatogenesis in dairy goats during the non-breeding season was not completely arrested. In addition, germ cell apoptosis and the morphology of Sertoli cells considerably changed in dairy goats during the non-breeding season. Sertoli cell autophagy is involved in the seasonal regulation of spermatogenesis in dairy goats. These findings provide key insights into the fertility and spermatogenesis of seasonal breeding animals.

Genes underlying the evolution of tetrapod testes size

Background Testes vary widely in mass relative to body mass across species, but we know very little about which genes underlie and contribute to such variation. This is partly because evidence for which genes are implicated in testis size variation tends to come from investigations involving just one or a few species. Contemporary comparative phylogenetic methods provide an opportunity to test candidate genes for their role in phenotypic change at a macro-evolutionary scale—across species and over millions of years. Previous attempts to detect genotype-phenotype associations across species have been limited in that they can only detect where genes have driven directional selection (e.g. brain size increase). Results Here, we introduce an approach that uses rates of evolutionary change to overcome this limitation to test whether any of twelve candidate genes have driven testis size evolution across tetrapod vertebrates—regardless of directionality. We do this by seeking a relationship between the rates of genetic and phenotypic evolution. Our results reveal five genes (Alkbh5, Dmrtb1, Pld6, Nlrp3, Sp4) that each have played unique and complex roles in tetrapod testis size diversity. In all five genes, we find strong significant associations between the rate of protein-coding substitutions and the rate of testis size evolution. Such an association has never, to our knowledge, been tested before for any gene or phenotype. Conclusions We describe a new approach to tackle one of the most fundamental questions in biology: how do individual genes give rise to biological diversity? The ability to detect genotype-phenotype associations that have acted across species has the potential to build a picture of how natural selection has sculpted phenotypic change over millions of years.

Pupal Cues Increase Sperm Production but Not Testis Size in an Insect

Theoretic and empirical studies show that social surroundings experienced by male insects during their larval or adult stage can influence their testicular investment in diverse ways. Although insect pupae do not feed and crawl, they can communicate using sex-specific and/or non-sex specific cues. Yet, it is unknown, in any insect, whether and how male pupae can fine-tune their resource allocation to sperm production and testis size in response to socio-sexual environments. We investigated this question using a moth, Ephestia kuehniella, which produces fertile eupyrene sperm and unfertile apyrene sperm. We held male pupae individually or in groups with different sex ratios, and dissected adults upon eclosion, measured their testis size, and counted both types of sperm. We demonstrated that after exposure to conspecific pupal cues regardless of sex, male pupae increased production of eupyrenes and apyrenes at the same rate but kept testis size unchanged. We suggest that testis size is fixed after pupation because most morphological traits are formed during the larval stage, allowing little room for pupae to adjust testis size. Like adults, male pupae with fully grown testes have sufficient resources to produce more sperm of both types according to the perceived increase in sperm competition risk.

Effects of a Recombinant Gonadotropin-Releasing Hormone Vaccine on Reproductive Function in Adult Male ICR Mice

Gonadotropin-releasing hormone (GnRH) regulates the reproductive endocrine system in mammals. The GnRH immunocontraception vaccine can aid animal population control and management. We evaluated a recombinant GnRH fusion protein with the adjuvant MONTANIDE ISA 206 VG as a GnRH vaccine in adult male ICR mice by evaluating anti-GnRH antibodies; concentrations of follicle-stimulating hormone (FSH), luteinizing hormone (LH), and testosterone; testis size and histomorphology; and semen quality. Response was assessed after intramuscular administration of the vaccine to mice in weeks 0, 4, and 8. The vaccine induced specific antibody response by week 5, with peak of antibody levels observed by week 13 and a declining level thereafter until the end of the study at week 24. Furthermore, it reduced serum FSH, LH, and testosterone concentrations. The vaccinated mice exhibited testicular atrophy and reduced sperm quality, concentration, morphology, and viability compared to control males. The outcomes of pairings of treated males with untreated females revealed reduced mating, pregnancy rates and number of litters compared to control pairings. Assessment of this GnRH vaccine in different species could assist its development for future applications.

Pupal Cues Increase Sperm Production but Not Testis Size in an Insect

Theoretic and empirical studies show that social surroundings experienced by male insects during their larval or adult stage can influence their testicular investment in diverse ways. Although insect pupae do not feed and crawl, they can communicate using sex-specific and/or non-sex specific cues. Yet, it is unknown, in any insect, whether and how male pupae can fine-tune their resource allocation to sperm production and testis size in response to socio-sexual environment. We investigated this question using a moth, Ephestia kuehniella, which produces fertile eupyrene sperm and unfertile apyrene sperm. We held male pupae individually or in groups with different sex ratios, and dissected adults upon eclosion, measured their testis size, and counted both types of sperm. We demonstrated that after exposure to conspecific pupal cues regardless of sex, male pupae increased production of eupyrenes and apyrenes at the same rate but kept testis size unchanged. We suggest that testis size is fixed after pupation because most morphological traits are formed during the larval stage, allowing little room for pupae to adjust testis size. Like adults, male pupae with fully grown testes have sufficient resources to produce more sperm of both types according to the perceived increase of sperm competition risk.