Congenital Zika virus infection impacts on male mouse offspring's reproductive biology.

In brief: Congenital ZIKV infection promotes alarming effects on male offspring's reproductive biology. This study showed the presence of the ZIKV antigen in the testis parenchyma, decreased testosterone levels, and sperm abnormalities in male offspring born to infected mothers. Abstract: Infection with ZIKV during pregnancy is associated with fetal developmental problems. Although neurological issues are being explored more in experimental studies, limited research has focused on the reproductive health consequences for offspring born to infected mothers. In this context, this study aimed to assess the impact of ZIKV infection during pregnancy on the testes and sperm of adult male offspring. Female mice were intraperitoneally inoculated with a Brazil strain of ZIKV during the 5.5th day of embryonic gestation. The offspring were evaluated 12 weeks after birth to analyze cellular and molecular changes in the testes and sperm. A novel approach combining variable-angle spectroscopic ellipsometry and machine learning modeling was also introduced for sperm sample analysis. The study revealed the presence of ZIKV protein in the testis parenchyma of adult male offspring born to infected mothers. It was shown that the testes exhibited altered steroidogenesis and inflammatory mediators, in addition to significant issues with spermiogenesis that resulted in sperm with DNA fragmentation, head defects, and protamination failure. Additionally, sperm dielectric properties and artificial intelligence showed potential for rapid identification and classification of sperm samples from infected mice. These findings provide crucial insights into the reproductive risks for men born from ZIKV-infected pregnant women.

Avian iPSC Derivation to Recover Threatened Wild Species: A Comprehensive Review in Light of Well-Established Protocols.

Induced pluripotent stem cells (iPSCs) were first generated by Yamanaka in 2006, revolutionizing research by overcoming limitations imposed by the use of embryonic stem cells. In terms of the conservation of endangered species, iPSC technology presents itself as a viable alternative for the manipulation of target genetics without compromising specimens. Although iPSCs have been successfully generated for various species, their application in nonmammalian species, particularly avian species, requires further in-depth investigation to cover the diversity of wild species at risk and their different protocol requirements. This study aims to provide an overview of the workflow for iPSC induction, comparing well-established protocols in humans and mice with the limited information available for avian species. Here, we discuss the somatic cell sources to be reprogrammed, genetic factors, delivery methods, enhancers, a brief history of achievements in avian iPSC derivation, the main approaches for iPSC characterization, and the future perspectives and challenges for the field. By examining the current protocols and state-of-the-art techniques employed in iPSC generation, we seek to contribute to the development of efficient and species-specific iPSC methodologies for at-risk avian species. The advancement of iPSC technology holds great promise for achieving in vitro germline competency and, consequently, addressing reproductive challenges in endangered species, providing valuable tools for basic research, bird genetic preservation and rescue, and the establishment of cryobanks for future conservation efforts.

From in vivo to in vitro: exploring the key molecular and cellular aspects of human female gametogenesis.

Human oogenesis is a highly complex and not yet fully understood process due to ethical and technological barriers that limit studies in the field. In this context, replicating female gametogenesis in vitro would not only provide a solution for some infertility problems, but also be an excellent study model to better understand the biological mechanisms that determine the formation of the female germline. In this review, we explore the main cellular and molecular aspects involved in human oogenesis and folliculogenesis in vivo, from the specification of primordial germ cells (PGCs) to the formation of the mature oocyte. We also sought to describe the important bidirectional relationship between the germ cell and the follicular somatic cells. Finally, we address the main advances and different methodologies used in the search for obtaining cells of the female germline in vitro.

Ecogenotoxicity of environmentally relevant atrazine concentrations: A threat to aquatic bioindicators.

Atrazine (ATZ) is a herbicide that is frequently present in surface waters and may result in damage to the health of various organisms, including humans. However, most scientific literature reports injuries caused by ATZ at high concentrations, which are not found in the environment. Therefore, the scope of this study was to investigate the impacts of realistic concentrations of ATZ found in surface waters (1, 2, 5, 10, 15 and 20 µg/L) using the bioindicators Allium cepa, Daphnia magna and zebrafish (Danio rerio). ATZ elicited a genotoxic effect in A. cepa, manifested by the induction of chromosomal aberrations, and a mutagenic effect with increased incidence of micronuclei formation, promotion of cell death and reduction in nuclear size revealed by flow cytometry analysis. D. magna exposed to 10, 15 and 20 µg/L of ATZ showed significant reduction in body size after 21 days, delayed first-brood release, decreased egg production and total offspring, as well as swimming behavioral changes. ATZ exposure promoted physiological and developmental alterations in zebrafish embryos, including an increased spontaneous movement rate, which led to premature hatching at all concentrations investigated. Increase in total body length, decrease of the yolk sac area, pericardial edema and higher heart rate were also detected in ATZ-treated zebrafish. In summary, environmentally relevant concentrations of ATZ can induce substantial alterations in the three bioindicators investigated. This study evidences the deleterious effects of ATZ on three aquatic bioindicators employing established and current techniques, and may contribute to elucidate the risks caused by this widely used herbicide even at low concentrations and short-to-medium-term exposure.

GATA-1 mutation alters the spermatogonial phase and steroidogenesis in adult mouse testis.

GATA-1 is a transcription factor from the GATA family, which features zinc fingers for DNA binding. This protein was initially identified as a crucial regulator of blood cell differentiation, but it is currently known that the Gata-1 gene expression is not limited to this system. Although the testis is also a site of significant GATA-1 expression, its role in testicular cells remains considerably unexplored. In the present study, we evaluated the testicular morphophysiology of adult ΔdblGATA mice with a mutation in the GATA-1 protein. Regarding testicular histology, GATA-1 mutant mice exhibited few changes in the seminiferous tubules, particularly in germ cells. A high proportion of differentiated spermatogonia, an increased number of apoptotic pre-leptotene spermatocytes (Caspase-3-positive), and a high frequency of sperm head defects were observed in ΔdblGATA mice. The main differences were observed in the intertubular compartment, as ΔdblGATA mice showed several morphofunctional changes in Leydig cells. Reduced volume, increased number and down-regulation of steroidogenic enzymes were observed in ΔdblGATA Leydig cells. Moreover, the mutant animal showed lower serum testosterone concentration and high LH levels. These results are consistent with the phenotypic and biometric data of mutant mice, i.e., shorter anogenital index and reduced accessory sexual gland weight. In conclusion, our findings suggest that GATA-1 protein is an important factor for germ cell differentiation as well as for the steroidogenic activity in the testis.

Gene delivery to Nile tilapia cells for transgenesis and the role of PI3K-c2α in angiogenesis.

Microinjection is commonly performed to achieve fish transgenesis; however, due to difficulties associated with this technique, new strategies are being developed. Here we evaluate the potential of lentiviral particles to genetically modify Nile tilapia cells to achieve transgenesis using three different approaches: spermatogonial stem cell (SSC) genetic modification and transplantation (SC), in vivo transduction of gametes (GT), and fertilised egg transduction (ET). The SC protocol using larvae generates animals with sustained production of modified sperm (80% of animals with 77% maximum sperm fluorescence [MSF]), but is a time-consuming protocol (sexual maturity in Nile tilapia is achieved at 6 months of age). GT is a faster technique, but the modified gamete production is temporary (70% of animals with 52% MSF). ET is an easier way to obtain mosaic transgenic animals compared to microinjection of eggs, but non-site-directed integration in the fish genome can be a problem. In this study, PI3Kc2α gene disruption impaired development during the embryo stage and caused premature death. The manipulator should choose a technique based on the time available for transgenic obtainment and if this generation is required to be continuous or not.

Biology and identity of fish spermatogonial stem cell.

Although present at relatively low number in the testis, spermatogonial stem cells (SSCs) are crucial for the establishment and maintenance of spermatogenesis in eukaryotes and, until recently, those cells were investigated in fish using morphological criteria. The isolation and characterization of these cells in fish have been so far limited by the lack of specific molecular markers, hampering the high SSCs biotechnological potential for aquaculture. However, some highly conserved vertebrate molecular markers, such as Gfra1 and Pou5f1/Oct4, are now available representing important candidates for studies evaluating the regulation of SSCs in fish and even functional investigations using germ cells transplantation. A technique already used to demonstrate that, different from mammals, fish germ stem cells (spermatogonia and oogonia) present high sexual plasticity that is determined by the somatic microenvironment. As relatively well established in mammals, and demonstrated in zebrafish and dogfish, this somatic environment is very important for the preferential location and regulation of SSCs. Importantly, a long-term in vitro culture system for SSCs has been now established for some fish species. Therefore, besides the aforementioned possibilities, such culture system would allow the development of strategies to in vitro investigate key regulatory and functional aspects of germline stem cells (ex: self-renewal and/or differentiation) or to amplify SSCs of rare, endangered, or commercially valuable fish species, representing an important tool for transgenesis and the development of new biotechnologies in fish production.