Research Note: Evaluating the vertical transmission potential of Salmonella Reading in broiler breeders.

Salmonella Reading (S. Reading) recently emerged as a foodborne pathogen causing extensive human outbreaks in North America from consuming contaminated poultry products, mostly from turkeys. Understanding the transmission dynamics of this pathogen is crucial for preventing future outbreaks. This study investigated the ability of S. Reading to colonize the tissues and contaminate eggs of broiler breeders. We utilized 2 S. Reading strains, marked with bioluminescence gene: the outbreak strain RS330 and a reference strain RS326. We used 32 commercially sourced broiler breeder hens, 34 wk of age, randomly assigned to the 2 treatments (16 hens per strain). Each hen was intravaginally inoculated with 108 CFU of the respective strain on d 1 and was rechallenged on d 4. Eggs were collected daily postchallenge to recover bioluminescent S. Reading strains from the external eggshell surface and internal egg contents. On d 7 postchallenge, 10 hens from each treatment group were euthanized. Ovaries, oviducts, and ceca were aseptically collected to detect S. Reading colonization. Results showed that 70.5% (36 of 51) and 34.5% (19 of 55) of external eggshell surfaces, and 4.0% (2 of 50) and 1.8% (1 of 54) of the internal egg contents tested positive for the outbreak and nonoutbreak strains. Additionally, 40.0% of ovaries, 70.0% of oviduct, and 70.0% of ceca samples from the outbreak strain group, and 20.0% of ovaries, 70.0% of oviduct, and 80.0% of ceca samples from nonoutbreak strain group were positive. No significant difference (P = 0.05) was observed in all the findings among the strains except for the eggshell surface contamination. These findings suggest that S. Reading can effectively colonize reproductive tissues, translocate to the ceca, and contaminate the eggs of hens. Future research is needed to determine whether S. Reading can remain viable within the eggs throughout incubation and until hatching.

Tissue-specific gene expression of genome-wide significant loci associated with major depressive disorder subtypes.

Major depressive disorder (MDD) is a clinically and genetically heterogeneous disorder. To reduce heterogeneity, large-scale genome-wide association studies have recently identified genome-wide significant loci associated with seven MDD subtypes. However, it was unclear in which tissues the genes near those loci are specifically expressed. We investigated whether genes related to specific MDD subtypes would be preferably expressed in a specific tissue. At 14 novel subtype-specific loci related to seven MDD subtypes-(1) non-atypical-like features MDD, (2) early-onset MDD, (3) recurrent MDD, (4) MDD with suicidal thoughts, (5) MDD without suicidal thoughts, (6) MDD with moderate impairment, and (7) postpartum depression, we investigated whether 22 genome-wide significant genetic variant-mapped genes were tissue-specifically expressed in brain, female reproductive, male specific, cardiovascular, gastrointestinal, or urinary tissues in the Genotype-Tissue Expression (GTEx) subjects (n ≤ 948). To confirm the tissue-specific expression in the GTEx, we used independent Human Protein Atlas (HPA) RNA-seq subjects (n ≤ 95). Of 22 genes, nine and five genes were tissue-specifically expressed in brain and female reproductive tissues, respectively (p < 2.27 × 10-3). RTN1, ERBB4, and AMIGO1 related to early-onset MDD, recurrent MDD, or MDD with suicidal thoughts were highly expressed in brain tissues (d = 1.19-2.71), while OAS1, LRRC9, DHRS7, PSMA5, SYPL2, and GULP1 related to non-atypical-like features MDD, early-onset MDD, MDD with suicidal thoughts, or postpartum depression were expressed at low levels in brain tissues (d = -0.17--1.48). DFNA5, CTBP2, PCNX4, SDCCAG8, and GULP1, which are related to early-onset MDD, MDD with moderate impairment, or postpartum depression, were highly expressed in female reproductive tissues (d = 0.80-2.08). Brain and female reproductive tissue-specific expression was confirmed in the HPA RNA-seq subjects. Our findings suggest that brain and female reproductive tissue-specific expression might contribute to the pathogenesis of MDD subtypes.

Evidences of differential methylation in the genome during development in the cactophilic Drosophila species.

DNA methylation with 5-methylcytosine (5mC) has been reported in the genome of several eukaryotes, with marked differences between vertebrates and invertebrates. DNA methylation is poorly understood as its role in evolution in insects. Drosophila gouveai (cluster Drosophila buzzatii) presents larvae that develop obligatorily in necrotic tissues of cacti in nature, with the distribution of populations in South America, and plasticity of phenotypes in insect-plant interaction. We characterize organisms at developmental stages and analyze variations at multiple methylation-sensitive loci in pupae, and adult flies using methylation sensitive amplification polymorphism. We obtained 326 loci with CCGG targets in the genome of D. gouveai. Genomic regions with molecular lengths from 100 to 700 pb were most informative about methylation states. Multiple loci show differences in methylation-sensitive sites (MSL) concerning developmental stages, such as in pupae (MSL = 40), female reproductive tissue (MSL = 76), and male reproductive tissues (MSL = 58). Our results are the first evidence of genome-wide methylation in D. gouveai organisms.

Expression Analysis of Lipocalin 2 (LCN2) in Reproductive and Non-Reproductive Tissues of Esr1-Deficient Mice.

Estrogen receptor alpha (ERα) is widely expressed in reproductive organs, but also in non-reproductive tissues of females and males. There is evidence that lipocalin 2 (LCN2), which has diverse immunological and metabolic functions, is regulated by ERα in adipose tissue. However, in many other tissues, the impact of ERα on LCN2 expression has not been studied yet. Therefore, we used an Esr1-deficient mouse strain and analyzed LCN2 expression in reproductive (ovary, testes) and non-reproductive tissues (kidney, spleen, liver, lung) of both sexes. Tissues collected from adult wild-type (WT) and Esr1-deficient animals were analyzed by immunohistochemistry, Western blot analysis, and RT-qPCR for Lcn2 expression. In non-reproductive tissues, only minor genotype- or sex-specific differences in LCN2 expression were detected. In contrast, significant differences in LCN2 expression were observed in reproductive tissues. Particularly, there was a strong increase in LCN2 in Esr1-deficient ovaries when compared to WTs. In summary, we found an inverse correlation between the presence of ERα and the expression of LCN2 in testes and ovaries. Our results provide an important basis to better understand LCN2 regulation in the context of hormones and in health and disease.

Transcriptome profiling of male and female Ascaris lumbricoides reproductive tissues.

BACKGROUND: Ascaris lumbricoides causes human ascariasis, the most prevalent helminth disease, infecting approximately 1 billion individuals globally. In 2019 the global disease burden was estimated to be 754,000 DALYs and resulted in 2090 deaths. In the absence of a vaccination strategy, treatment of ascariasis has relied on anthelminthic chemotherapy, but drug resistance is a concern. The propensity for reinfection is also a major challenge to disease control; female worms lay up to 200,000 eggs daily, which contaminate surrounding environments and remain viable for years, resulting in high transmission rates. Understanding the molecular mechanisms of reproductive processes, including control of egg production, spermatogenesis, oogenesis and embryogenesis, will drive the development of new drugs and/or vaccine targets for future ascariasis control. METHODS: Transcriptome profiles of discrete reproductive and somatic tissue samples were generated from adult male and female worms using Illumina HiSeq with 2 × 150 bp paired-end sequencing. Male tissues included: testis germinal zone, testis part of vas deferens, seminal vesicle and somatic tissue. Female tissues included: ovary germinal zone, ovary part of the oviduct, uterus and somatic tissue. Differentially expressed genes (DEGs) were identified from the fragments per kilobases per million reads (FPKM) profiles. Hierarchical analysis was performed to identify tissue-specific genes. Furthermore, Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses were employed to identify significant terms and pathways for the DEGs. RESULTS: DEGs involved in protein phosphorylation and adhesion molecules were indicated to play a crucial role in spermatogenesis and fertilization, respectively. Those genes associated with the G-protein-coupled receptor (GPCR) signaling pathway and small GTPase-mediated signal transduction pathway play an essential role in cytoskeleton organization during oogenesis. Additionally, DEGs associated with the SMA genes and TGF-ß signaling pathway are crucial in adult female embryogenesis. Some genes associated with particular biological processes and pathways that were identified in this study have been linked to defects in germline development, embryogenesis and reproductive behavior. In the enriched KEGG pathway analysis, Hippo signaling, oxytocin signaling and tight junction pathways were identified to play a role in Ascaris male and female reproductive systems. CONCLUSIONS: This study has provided comprehensive transcriptome profiles of discrete A. lumbricoides reproductive tissue samples, revealing the molecular basis of these functionally important tissues. The data generated from this study will provide fundamental knowledge on the reproductive biology of Ascaris and will inform future target identification for anti-ascariasis drugs and/or vaccines.

Biomaterial strategies for the application of reproductive tissue engineering.

Human reproductive organs are of vital importance to the life of an individual and the reproduction of human populations. So far, traditional methods have a limited effect in recovering the function and fertility of reproductive organs and tissues. Thus, aim to replace and facilitate the regrowth of damaged or diseased tissue, various biomaterials are developed to offer hope to overcome these difficulties and help gain further research progress in reproductive tissue engineering. In this review, we focus on the biomaterials and their four main applications in reproductive tissue engineering: in vitro generation and culture of reproductive cells; development of reproductive organoids and models; in vivo transplantation of reproductive cells or tissues; and regeneration of reproductive tissue. In reproductive tissue engineering, designing biomaterials for different applications with different mechanical properties, structure, function, and microenvironment is challenging and important, and deserves more attention.

Reproductive tissue-specific translatome of a rice thermo-sensitive genic male sterile line.

Translational regulation, especially tissue- or cell type-specific gene regulation, plays essential roles in plant growth and development. Thermo-sensitive genic male sterile (TGMS) lines have been widely used for hybrid breeding in rice (Oryza sativa). However, little is known about translational regulation during reproductive stage in TGMS rice. Here, we use translating ribosome affinity purification (TRAP) combined with RNA sequencing to investigate the reproductive tissue-specific translatome of TGMS rice expressing FLAG-tagged ribosomal protein L18 (RPL18) from the germline-specific promoter MEIOSIS ARRESTED AT LEPTOTENE1 (MEL1). Differentially expressed genes at the transcriptional and translational levels are enriched in pollen and anther-related formation and development processes. These contain a number of genes reported to be involved in tapetum programmed cell death (PCD) and lipid metabolism during pollen development and anther dehiscence in rice, including several encoding transcription factors and key enzymes, as well as several long non-coding RNAs (lncRNAs) that potentially affect tapetum and pollen-related genes in male sterility. This study represents the comprehensive reproductive tissue-specific characterization of the translatome in TGMS rice. These results contribute to our understanding of the molecular basis of sterility in TGMS rice and will facilitate further genetic manipulation of TGMS rice in two-line breeding systems.

Integrated multiple transcriptomes in oviductal tissue across the porcine estrous cycle reveal functional roles in oocyte maturation and transport.

Understanding the changes in the swine female reproductive system is important for solving issues related to reproductive failure and litter size. Elucidating the regulatory mechanisms of the natural estrous cycle in the oviduct under non-fertilisation conditions can improve our understanding of its role in the reproductive system. Herein, whole transcriptome RNA sequencing of oviduct tissue samples was performed. The differentially expressed genes (DEGs) were identified for each time point relative to day 0 and classified into three clusters based on their expression patterns. Clusters 1 and 2 included genes involved in the physiological changes through the estrous cycle. Cluster 1 genes were mainly involved in PI3K-Akt signaling and steroid hormone biosynthesis pathways. Cluster 2 genes were involved in extracellular matrix-receptor interactions and protein digestion pathways. In Cluster 3, the DEGs were downregulated in the luteal phase; they were strongly associated with cell cycle, calcium signaling, and oocyte meiosis. The gene expression in the oviduct during the estrous cycle influenced oocyte transport and fertilization. Our findings provide a basis for successfully breeding pigs and elucidating the mechanisms underlying the changes in the pig oviduct during the estrous cycle.

Understanding the swine female reproductive system is important for solving issues related to reproductive failure and litter size. The oviduct is the site of fertilization. After fertilization, the fertilized egg moves to the uterus for implantation. Elucidating the regulatory mechanisms of the estrous cycle in the oviduct can improve our understanding of their roles. In this study, whole transcriptome RNA sequencing of oviduct tissue samples was performed throughout the estrous cycle to screen for differentially expressed genes (DEGs). The DEGs were classified into three clusters based on their expression patterns. Clusters 1 and 2 included genes involved in the physiological changes observed through the estrous cycle. The expression levels of Cluster 3 genes were downregulated specifically in the luteal phase; this was associated with calcium signalling and oocyte meiosis. In this study, we identified that the expression of genes in the oviduct influences oocyte transport and fertilization, which are the key functions of the oviduct. This study provides a basis for successful breeding in the pig industry and elucidating the mechanisms underlying the changes in the pig oviduct during the estrous cycle.

Advanced bioengineering of male germ stem cells to preserve fertility.

In modern life, several factors such as genetics, exposure to toxins, and aging have resulted in significant levels of male infertility, estimated to be approximately 18% worldwide. In response, substantial progress has been made to improve in vitro fertilization treatments (e.g. microsurgical testicular sperm extraction (m-TESE), intra-cytoplasmic sperm injection (ICSI), and round spermatid injection (ROSI)). Mimicking the structure of testicular natural extracellular matrices (ECM) outside of the body is one clear route toward complete in vitro spermatogenesis and male fertility preservation. Here, a new wave of technological innovations is underway applying regenerative medicine strategies to cell-tissue culture on natural or synthetic scaffolds supplemented with bioactive factors. The emergence of advanced bioengineered systems suggests new hope for male fertility preservation through development of functional male germ cells. To date, few studies aimed at in vitro spermatogenesis have resulted in relevant numbers of mature gametes. However, a substantial body of knowledge on conditions that are required to maintain and mature male germ cells in vitro is now in place. This review focuses on advanced bioengineering methods such as microfluidic systems, bio-fabricated scaffolds, and 3D organ culture applied to the germline for fertility preservation through in vitro spermatogenesis.

NLRP3 inflammasome: A joint, potential therapeutic target in management of COVID-19 and fertility problems.

To overcome COVID-19 long-term consequences, one possible approach is to control inflammasomes activation, because SARS-CoV-2 can induce humoral and cellular immune responses. In this opinion article we hypothesized that if it is proven with convincing and unmistakable evidence that firstly, SARS-CoV-2 can enter cells and damage them through its common receptors in the reproductive tissues, and secondly, inflammasome pathway activation is responsible for the damages caused, then the inflammasome inhibitors might be considered as suitable candidates in preventing the pathological effects on the germ cells and reproductive tissues and subsequent fertility.

No Evidence for Seed Transmission of Tomato Yellow Leaf Curl Sardinia Virus in Tomato.

Seed transmission is an important factor in the epidemiology of plant pathogens. Geminiviruses are serious pests spread in tropical and subtropical regions. They are transmitted by hemipteran insects, but a few cases of transmission through seeds were recently reported. Here, we investigated the tomato seed transmissibility of the begomovirus tomato yellow leaf curl Sardinia virus (TYLCSV), one of the agents inducing the tomato yellow leaf curl disease, heavily affecting tomato crops in the Mediterranean area. None of the 180 seedlings originating from TYLCSV-infected plants showed any phenotypic alteration typical of virus infection. Moreover, whole viral genomic molecules could not be detected in their cotyledons and true leaves, neither by membrane hybridization nor by rolling-circle amplification followed by PCR, indicating that TYLCSV is not a seed-transmissible pathogen for tomato. Examining the localization of TYLCSV DNA in progenitor plants, we detected the virus genome by PCR in all vegetative and reproductive tissues, but viral genomic and replicative forms were found only in leaves, flowers and fruit flesh, not in seeds and embryos. Closer investigations allowed us to discover for the first time that these embryos were superficially contaminated by TYLCSV DNA but whole genomic molecules were not detectable. Therefore, the inability of TYLCSV genomic molecules to colonize tomato embryos during infection justifies the lack of seed transmissibility observed in this host.

Quantification of zearalenone and α-zearalenol in swine liver and reproductive tissues using GC-MS.

The mycotoxin zearalenone (ZEN) is a common contaminant of swine feed which has been related to a wide range of reproductive anomalies in swine, such as pelvic organ prolapse, anestrous, and pseudopregnancy. New information is needed to understand how ZEN and related metabolites accumulate in swine reproductive tissues. We conducted a feeding study to track ZEN and the metabolite α-zearalenol (α-ZEL) in swine liver and reproductive tissues. Thirty pubertal gilts were randomly assigned one of three treatments, with ten pigs in each treatment group: (1) base feed with solvent for 21 days, (2) ZEN-spiked feed for seven days followed by base feed with solvent for 14 days, and (3) ZEN-spiked feed for 21 days. At the end of the trial, liver, anterior vagina, posterior vagina, cervix, uterus, ovaries, and broad ligament were collected from pigs. ZEN was found in the anterior vagina, posterior vagina, cervix, and ovaries, with significantly higher concentrations in the cervix relative to other reproductive tissues. ZEN and α-ZEL were found in liver tissue from pigs in each treatment group. Our results show that ZEN accumulates more in the cervix than other reproductive tissues. The presence of ZEN in reproductive tissues may be indicative of ZEN-related reproductive symptoms. Future work could examine how ZEN concentrations vary in reproductive tissues as a factor of the pigs age, weight, sex, or parity, to establish parameters that make pig more sensitive to ZEN.

Evaluating risk, safety and efficacy of novel reproductive techniques and therapies through the EuroGTP II risk assessment tool.

STUDY QUESTION: Can risks associated with novelties in assisted reproduction technologies (ARTs) be assessed in a systematic and structured way? SUMMARY ANSWER: An ART-specific risk assessment tool has been developed to assess the risks associated with the development of novelties in ART (EuroGTP II-ART). WHAT IS KNOWN ALREADY: How to implement new technologies in ART is well-described in the literature. The successive steps should include testing in animal models, executing pre-clinical studies using supernumerary gametes or embryos, prospective clinical trials and finally, short- and long-term follow-up studies on the health of the offspring. A framework categorizing treatments from experimental through innovative to established according to the extent of the studies conducted has been devised. However, a systematic and standardized methodology to facilitate risk evaluation before innovations are performed in a clinical setting is lacking. STUDY DESIGN, SIZE, DURATION: The EuroGTP II-ART risk assessment tool was developed on the basis of a generic risk assessment algorithm developed for tissue and cell therapies and products (TCTPs) in the context of the project 'Good Practices for demonstrating safety and quality through recipient follow-up European Good Tissue and cells Practices II (EuroGTP II)'. For this purpose, a series of four meetings was held in which eight ART experts participated. In addition, several tests and simulations were undertaken to fine-tune the final tool. PARTICIPANTS/MATERIALS, SETTING, METHODS: The three steps comprising the EuroGTP II methodology were evaluated against its usefulness and applicability in ART. Ways to improve and adapt the methodology into ART risk assessment were agreed and implemented. MAIN RESULTS AND THE ROLE OF CHANCE: Assessment of the novelty (Step 1), consisting of seven questions, is the same as for other TCTPs. Practical examples were included for better understanding. Identification of potential risks and consequences (Step 2), consisting of a series of risks and risk consequences to consider during risk assessment, was adapted from the generic methodology, adding more potential risks for processes involving gonadic tissues. The algorithm to score risks was also adapted, giving a specific range of highest possible risk scores. A list of strategies for risk reduction and definition of extended studies required to ensure effectiveness and safety (Step 3) was also produced by the ART experts, based on generic EuroGTP II methodology. Several explanations and examples were provided for each of the steps for better understanding within this field. LIMITATIONS, REASONS FOR CAUTION: A multidisciplinary team is needed to perform risk assessment, to interpret results and to determine risk mitigation strategies and/or next steps required to ensure the safety in the clinical use of novelties. WIDER IMPLICATIONS OF THE FINDINGS: This is a dynamic tool whose value goes beyond assessment of risk before implementing a novel ART in clinical practice, to re-evaluate risks based on information collected during the process. STUDY FUNDING / COMPETING INTEREST(S): This study was called EUROGTP II and was funded by the European Commission (Grant agreement number 709567). The authors declare no competing interests concerning the results of this study.

Ultrasound vibro-elastography for assessing mechanical properties of porcine reproductive tissues in an ex vivo model.

BACKGROUND: The aim of this study was to use ultrasound vibro-elastography (UVE) for measuring surface wave speed and assessing mechanical properties of ex vivo porcine reproductive tissues, including the uterus, bladder, cornua and cervix. METHODS: In UVE, a 0.1-s harmonic vibration at low frequency was generated on the tissue surface with a handheld shaker. A linear-array ultrasound probe was used to measure the resulting surface wave propagation. Surface wave speeds of tissues were measured in the frequency range of 100-300 Hz. Mechanical properties of the tissue were calculated based on wave speed dispersion with frequency. FINDINGS: The obtained results showed that the surface wave speeds of porcine bladder, cervix, cornua and uterus increased with frequency. There were no statistically significant differences in the wave speeds or mechanical properties among the porcine bladder, cervix, cornua and uterus. INTERPRETATION: Experimental data obtained in this study may be used as a reference to study in vivo surface wave speed or mechanical properties for porcine or human reproductive tissues.

Vitamin D Effects on the Immune System from Periconception through Pregnancy.

Vitamin D is a well-known secosteroid and guardian of bone health and calcium homeostasis. Studies on its role in immunomodulatory functions have expanded its field in recent years. In addition to its impact on human physiology, vitamin D influences the differentiation and proliferation of immune system modulators, interleukin expression and antimicrobial responses. Furthermore, it has been shown that vitamin D is synthesized in female reproductive tissues and, by modulating the immune system, affects the periconception period and reproductive outcomes. B cells, T cells, macrophages and dendritic cells can all synthesize active vitamin D and are involved in processes which occur from fertilization, implantation and maintenance of pregnancy. Components of vitamin D synthesis are expressed in the ovary, decidua, endometrium and placenta. An inadequate vitamin D level has been associated with recurrent implantation failure and pregnancy loss and is associated with pregnancy-related disorders like preeclampsia. This paper reviews the most important data on immunomodulatory vitamin D effects in relation to the immune system from periconception to pregnancy and provides an insight into the possible consequences of vitamin D deficiency before and during pregnancy.

Development of a new reproductive tissue cryopreservation clinical service for children: the Oxford programme.

PURPOSE: This article describes the development of a new reproductive tissue cryopreservation clinical service for children at high risk of infertility in the NHS during times of severe financial constraints in the health service. METHOD: A development plan with two phases was drawn up. Phase 1 restricted the service to childhood cancer patients referred to the Oxford Paediatric Oncology and Haematology Principle Treatment Centre. It was estimated that there would be 10 patients/year and used existing staff and facilities from paediatric oncology, surgery, anaesthetics radiology, pathology, psychology, teenage-young adult gynaecology, and an existing Human Tissue Authority tissue bank with a licence for storage of tissue under a Human Sector Licence. Phase 2 extended the service to include children and young adults across England, Wales and Ireland-patients from Scotland having access to a research programme in Edinburgh. The main challenge in phase 2 being resources and the need for patients to be able to be treated as close to home as safely as possible. RESULTS: The Oxford team developed information resources and eligibility criteria based on published best practice, referral and treatment pathways, multidisciplinary team meetings, a network of third party sites, and a dedicated case management and database. As the programme expanded, the Oxford team was able to justify to management the need for a dedicated theatre list. Patient feedback through questionnaires, qualitative work conducted as part of a Ph.D. thesis as well as direct patient stories and interviews in TV, and radio features underpins the positive impact the programme has on patients and their families. CONCLUSION: The Oxford Reproductive Cryopreservation programme delivers fertility preservation treatment to children and young adults at high risk of infertility safely, effectively and as close to home as possible. The onward view is to apply for national funding for this programme for recognition and sustainability.

Biomechanical properties of female dolphin reproductive tissue.

Whales, dolphins, and porpoises have unusual vaginal folds of unknown function(s) that are hypothesized to play an important role in sexual selection. The potential function of vaginal folds was assessed by testing the mechanical properties of common bottlenose dolphin (Tursiops truncatus) reproductive tract tissues in 6 different regions and across age classes in post-mortem specimens. We assessed the regional (local) and overall effective elastic modulus of tissues using indentation and tensile tests, respectively. We explore the non-linear mechanical response of biological tissues, which are not often quantified. Indentation tests demonstrated that sexual maturity state, tissue region, force history, and force magnitude values significantly affected the measured effective elastic modulus. Tissue was stiffest in the vaginal fold region and overall stiffer in sexually immature compared to mature animals, likely reflecting biomechanical adaptations associated with copulation and parturition. Tensile tests showed that only tissue region significantly affected the effective modulus. Our data support the hypothesis that vaginal folds function as mechanical barriers to the penis and may provide females with mechanisms to reduce copulatory forces on other reproductive tissue. STATEMENT OF SIGNIFICANCE: Cetaceans have unusual folds of vaginal wall tissue that appear to evolve under sexual selection mechanisms and present physical barriers to the penis during copulation. We explore the biomaterial properties of vaginal fold tissue, how it varies from other reproductive tract tissues, and ontogenetic patterns. We demonstrate that vaginal folds can withstand higher mechanical forces and respond in a manner conducive to dissipating copulatory forces to other reproductive tissues. This study yields exciting insights on how female genital tissue may function during copulation, and is the first to do so in any vertebrate species. Additionally, we provide an example for testing biological tissues, non-linear properties, and materials with uneven surface structure and uneven thickness.

Comprehensive assessment of cryogenic storage risk and quality management concerns: best practice guidelines for ART labs.

Recent publicized events of cryogenic storage tank failures have created nationwide concern among infertility patients and patients storing embryos and gametes for future use. To assure patient confidence, quality management (QM) plans applied by in vitro fertilization (IVF) laboratories need to include a more comprehensive focus on the cryostorage of reproductive specimens. The purpose of this review is to provide best practice guidelines for the cryogenic storage of sperm, oocytes, embryos, and other reproductive tissues (e.g., testicular and ovarian tissue, cord blood cells, and stem cells) and recommend a strategy of thorough and appropriate quality and risk management procedures aimed to alleviate or minimize the consequences from catastrophic events.

In vivo genome editing targeted towards the female reproductive system.

The discovery of sequence specific nucleases such as ZFNs, TALENs, and CRISPR/Cas9 has revolutionized genome editing. The CRISPR/Cas9 system has particularly emerged as a highly simple and efficient approach towards generating genome-edited animal models of most of the experimental species. The limitation of these novel genome editing tools is that, till date, they depend on traditional pronuclear injection (PI)-based transgenic technologies developed over the last three decades. PI requires expensive micromanipulator systems and the equipment operators must possess a high level of skill. Therefore, since the establishment of PI-based transgenesis, various research groups worldwide have attempted to develop alternative and simple gene delivery methods. However, owing to the failure of chromosomal integration of the transgene, none of these methods gained the level of confidence as that by the PI method in order to be adapted as a routine approach. The recently developed genome editing systems do not require complicated techniques. Therefore, presently, attention is being focused on non-PI-based gene delivery into germ cells for simple and rapid production of genetically engineered animals. For example, a few reports during the previous 1-2 years demonstrated the use of electroporation (EP) in isolated zygotes that helped to overcome the absolute dependency on PI techniques. Recently, another breakthrough technology called genome editing via oviductal nucleic acids delivery (GONAD) that directly delivers nucleic acids into zygotes within the oviducts in situ was developed. This technology completely relieves the bottlenecks of animal transgenesis as it does not require PI and ex vivo handling of embryos. This review discusses in detail the in vivo gene delivery methods targeted towards female reproductive tissues as these methods that have been developed over the past 2-3 decades can now be re-evaluated for their suitability to deliver the CRISPR/Cas9 components to produce transgenic animals. This review also provides an overview of the latest advances in CRISPR-enabled delivery technologies that have caused paradigm shifts in animal transgenesis methodologies.