Follicular oocyte as a potential target for severe acute respiratory syndrome coronavirus 2 infection.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) was reported in December 2019 and rapidly became a pandemic as coronavirus disease 2019 (COVID-19). Apart from other organs, presence of specific receptor angiotensin-converting enzyme (ACE2) and corresponding proteases such as transmembrane serine protease 2, basigin and cysteine protease cathepsin L make follicular somatic cells as well as oocyte as potential targets for SARS-CoV-2 infection. The SARS-CoV-2 causes inflammation and hypoxia that generate reactive oxygen species (ROS) in critically ill patients. In addition, a large number of casualties and insecurity of life due to repeated waves of SARS-CoV-2 infection generate psychological stress and cortisol resulting in the further generation of ROS. The excess levels of ROS under physiological range cause meiotic instability, while high levels result in oxidative stress that trigger various death pathways and affect number as well as quality of follicular oocytes. Although, emerging evidence suggests that the SARS-CoV-2 utilises cellular machinery of ovarian follicular cells, generates ROS and impairs quality of follicular oocytes, the underlying mechanism of viral entry into host cell and its negative impact on the follicular oocyte remains poorly understood. Therefore, this review summarises emerging evidence on the presence of cellular machinery for SARS-CoV-2 in ovarian follicles and the potential negative impact of viral infection on the follicular oocytes that affect ovarian functions in critically ill and stressed women.

Virus analog decreases estradiol secretion in FSH-treated human ovarian granulosa cells.

The aim of this study was to evaluate the effect of virus infection on estradiol (E2) production in human ovarian granulosa cells. Polyriboinosinic polyribocytidylic acid [Poly (I: C)], a synthetic analog of viral double stranded RNA that can be recognized by Toll like receptor 3 (TLR3), was used to imitate virus infection. Granulosa cells (GCs) obtained from patients undergoing in vitro fertilization and embryo transfer (IVF-ET) were cultured in vitro and treated with Poly (I: C), FSH, or both. Concentration of E2 was assayed by electrochemiluminescence. The mRNA and protein expression of TLR3 and aromatase were determined by real-time quantitative PCR (qPCR) and Western blot, respectively. The results showed that expression of TLR3 mRNA was significantly increased after Poly (I: C) stimulation. Poly (I: C) decreased E2 synthesis in FSH-treated GCs. Poly (I: C) inhibited the expression of aromatase in FSH-treated GCs. This study demonstrated that Poly (I: C) inhibits the synthesis of estradiol by granulosa cells under the stimulation of FSH, which might contribute to disturbance of follicular development and ovulation.

The follicle promotes the evolution of variants of avian leukosis virus subgroup J in vertical transmission.

Avian leukosis virus (ALV) is one of the main causative agent of tumor development, which brings enormous economic losses to the poultry industry worldwide. ALV can be transmitted horizontally and vertically, and the latter often give rise to more adverse pathogenicity. However, the propagation and evolution of ALV underlying vertical transmission remain not-well understood. Herein, an animal model for the evolution of variants of ALV subgroup J (ALV-J) in the vertical transmission was built and different organs from infected hens and plasma from their ALV-positive progenies were collected, and then three segments in the hypervariable regions of ALV (gp85-A, gp85-B, LTR-C) were amplified and sequenced using conventional Sanger sequencing and MiSeq high-throughput sequencing, respectively. The results showed that the genomic diversity of ALV-J occurred in different organs from ALV-J infected hen, and that the dominant variants in different organs of parental hens, especially in follicle, changed significantly compared with original inoculum strain. Notably, the dominant variants in progenies exhibited higher homologies with variants in parental hens' follicle (88.9%-98.9%) than other organs (85.6%-91.1%), and most consistent mutations in the variants were observed between the progenies and parental hen's follicle. Furthermore, HyPhy analysis indicated that the global selection pressure value (ω) in the follicle is significantly higher than those in other organs. In summary, an animal model for vertical transmission was built and our findings revealed the evolution of variants of ALV in the process of vertical transmission, moreover, the variants were most likely to be taken to the next generation via follicle, which may be related to the higher selection pressure follicle underwent.

Bovine herpesvirus 1 can cross the intact zona pellucida of bovine oocytes after artificial infection.

Bovine herpesvirus 1 (BHV1) is an important bovine pathogen, responsible for respiratory diseases and reproductive problems. This study investigated the penetration capacity of BHV1 into oocytes after co-incubation for either 1 h or 24 h. Immunofluorescence assays in cumulus-oocyte complexes (COCs) and denuded oocytes (without the presence of cumulus cells) were performed and evaluated using confocal laser scanning microscopy. Blood samples and ovaries from BHV1 seronegative cows were used. The oocytes recovered were divided into two groups. Group I comprised COCs (n = 312) and denuded oocytes (n = 296), which were experimentally infected with BHV1 and incubated for 1 h at 38.5°C and 5% CO2. Group II comprised COCs (n = 425) and denuded oocytes (n = 405), which were co-incubated with BHV1 under the same conditions for 24 h. The negative control of these two groups was respectively subjected to the same protocol, except for exposure to BHV1. To our knowledge, this study provides the first evidence of BHV1 detection within COCs and denuded oocytes exhibiting intact zona pellucida when co-incubated with the virus for 24 h. Immunolocalization also confirmed the presence of BHV1 in the cytoplasm of the cumulus cells of all COCs exposed to the virus after both incubation periods. In conclusion, detection of BHV1 inside oocytes has a great meaning for the field of animal reproduction. The detection of BHV1 in different layers of cumulus cells also demonstrates that these cells are sources of viral infection.

Case report of Zika virus during controlled ovarian hyperstimulation: results from follicular fluid, cumulus cells and oocytes.

We describe a case of a 37-year-old female, indicated for in vitro fertilisation. She developed skin rash on her trunk and limbs, during the treatment. RT-PCR results were positive in her blood and negative in her husband's blood and semen. Oocyte aspiration was performed, retrieving 7 oocytes, follicular fluid, and cumulus cells. RT-PCR results for the follicular fluid and cumulus cells were negative for ZIKV, and positive for only 2 oocytes. This is the first report in the literature analysing ZIKV in the follicular fluid, cumulus cells, and oocytes, and will contribute to the understanding of ZIKV infection and transmission.

[Intrafollicular infection of mammals and human oocytes by the herpes simplex virus].

The goal of this work was to study the capacity of the herpes simplex virus (HSV) of infecting ovary with disease in case of the intravaginal experimental animals. The results of the study demonstrated that the ascending HSV infection in mice lead to modification of all the cells of the ovary, including follicular cells synthesizing estrogen and progesterone. The two hormones influence the development of the disease. Estrogens provide the protective effects against the virus. Progesterone does not modify the body sensitivity to HSV, but reduces the effectiveness of the antiviral immunity, resulting in increased mortality of animals. We demonstrated that infection of oocytes in ovarian follicles of female mice during infection with HSV modified the process in vitro and for the first time demonstrated the detection of viral antigens in mature oocytes in patient with infertility. During the intracytoplasmic sperm injection into the infected oocytes (ICSI), the failure of fertilization was observed. These results are of interest, because there is no available literature on whether HSV infection of oocytes can have a direct negative impact on the process of fertilization in humans.

Correlation between vertical transmission of hepatitis B virus and the expression of HBsAg in ovarian follicles and placenta.

BACKGROUND: The aim of this study was to investigate the correlation between the expression of hepatitis B surface antigen (HBsAg) in human ovary and placenta and the vertical transmission of hepatitis B virus (HBV). METHODOLOGY/PRINCIPAL FIDNINGS: Ovarian and placental tissue specimens of pregnant women infected with HBV were collected during cesarean section and immunostained for HBsAg. The sera of the corresponding newborns were tested for HBV markers and HBV DNA. HBsAg was detected in 15 out of 33 (45%) placental tissues and was further detected in capillary endothelial cells in 4 specimens (26%), of which 3 (75%) corresponding infants were infected with HBV in utero. Out of the 33 ovarian tissues, 7 (21%) were positive for HBsAg, of which 2 (28%) showed HBsAg in ovarian follicles and the 2 corresponding infants (100%) had intrauterine HBV infection. CONCLUSIONS/SIGNIFICANCE: HBsAg expression in cells of the ovarian follicle or placental capillary endothelium signal a higher risk for intrauterine HBV infection.

Viruses associated with ovarian degeneration in Apis mellifera L. queens.

Queen fecundity is a critical issue for the health of honeybee (Apis mellifera L.) colonies, as she is the only reproductive female in the colony and responsible for the constant renewal of the worker bee population. Any factor affecting the queen's fecundity will stagnate colony development, increasing its susceptibility to opportunistic pathogens. We discovered a pathology affecting the ovaries, characterized by a yellow discoloration concentrated in the apex of the ovaries resulting from degenerative lesions in the follicles. In extreme cases, marked by intense discoloration, the majority of the ovarioles were affected and these cases were universally associated with egg-laying deficiencies in the queens. Microscopic examination of the degenerated follicles showed extensive paracrystal lattices of 30 nm icosahedral viral particles. A cDNA library from degenerated ovaries contained a high frequency of deformed wing virus (DWV) and Varroa destructor virus 1 (VDV-1) sequences, two common and closely related honeybee Iflaviruses. These could also be identified by in situ hybridization in various parts of the ovary. A large-scale survey for 10 distinct honeybee viruses showed that DWV and VDV-1 were by far the most prevalent honeybee viruses in queen populations, with distinctly higher prevalence in mated queens (100% and 67%, respectively for DWV and VDV-1) than in virgin queens (37% and 0%, respectively). Since very high viral titres could be recorded in the ovaries and abdomens of both functional and deficient queens, no significant correlation could be made between viral titre and ovarian degeneration or egg-laying deficiency among the wider population of queens. Although our data suggest that DWV and VDV-1 have a role in extreme cases of ovarian degeneration, infection of the ovaries by these viruses does not necessarily result in ovarian degeneration, even at high titres, and additional factors are likely to be involved in this pathology.

Proviral DNA of caprine arthritis encephalitis virus (CAEV) is detected in cumulus oophorus cells but not in oocytes from naturally infected goats.

The aim of this study was to determine whether oocytes taken from ovarian follicles in 123 naturally infected goats were carrying the proviral CAEV genome. Examination of DNA isolated from 190 batches of oocytes with intact cumulus cells and 190 batches of oocytes whose cumulus cells had been removed, taken from follicles of the same ovaries, demonstrated that 42/190 batches of oocytes with intact cumulus cells had the proviral CAEV genome, whereas none of the 190 batches of oocytes without cumulus cells were positive for the provirus. To confirm that the proviral genome was present in the cumulus cells and not in the oocyte cells, 586 oocytes from 56 different ovaries, were separated from their cumulus cells. The DNA was then extracted from each fraction and examined. The purity of the oocyte fraction was verified by searching for granulosa cell-specific mRNA, using RT-PCR; this was negative in all the batches of oocytes in which the cumulus cells had been removed. PCR analysis demonstrated that none of the oocytes without cumulus cells were positive, whereas 22/56 of the batches with cumulus cells were found to be positive. This study clearly demonstrates that despite being surrounded by infected cumulus cells, the oocytes are not infected, and that the enzymatic and mechanical technique for removing the cells surrounding the oocyte, as used in this study, is effective, thus enabling CAEV-free oocytes to be obtained from infected goats.

Evidence for the localization of porcine reproductive and respiratory syndrome virus (PRRSV) antigen and RNA in ovarian follicles in gilts.

The pathogenesis of porcine reproductive and respiratory syndrome virus (PRRSV) infection in ovary was studied in sexually mature, cycling, nonsynchronized gilts infected with the PRRSV 16244B, a virulent field strain. Previous studies have shown that PRRSV can be isolated from ovaries and is transplacentally passed from gilts to the fetuses. The cause of infertility following PRRSV infection is not known. In this study, we identified the tropism of PRRSV in ovarian tissue from experimentally infected gilts in samples collected between 7 and 21 days postinfection (DPI). Tissues were collected and examined by virus isolation, in situ hybridization (ISH), immunohistochemistry (IHC), and double labeling to identify PRRSV-infected cell types. PRRSV was isolated in ovarian follicles at 7 days DPI. The IHC and ISH indicated that PRRSV-positive cells in ovaries were predominantly macrophages, which were numerous in atretic follicles. No evidence of infection and/or perpetuation of PRRSV in ova was observed, indicating that the female gonad is an unlikely site of persistence. No alteration of the normal ovarian architecture that would support a possible role of PRRSV infection in porcine female infertility was observed.

Bovine viral diarrhea virus replication in bovine follicular epithelial cells derived from persistently infected heifers.

Bovine follicle fluid and oocytes surrounded by follicular epithelial (FE) cells were collected from ovaries of two heifers persistently infected with bovine viral diarrhea virus (BVDV). BVDV was present in the follicle fluid at a higher titer than in serum. The oocytes were matured in vitro under culture conditions of 39 degrees C in humidified air containing 5% CO2. In vitro fertilization was performed after 24 hr in culture (the day of insemination was defined as day 1), and culture was continued through day 10. BVDV was present in the culture medium at titers of 10(2.25) to 10(3.25) TC(I)D50/0.1 ml. The virus was also detected in FE cells collected on day 10. Viral antigen was demonstrated in the cytoplasm of FE cells by the indirect immunofluorescence technique. However, no BVDV was detected in the embryos on day 10. These findings suggested that the oocytes or embryos were unlikely to be infected with BVDV, but that the FE cells were infected with BVDV and supported virus replication in cattle persistently infected with BVDV.