Genetic testing of sperm donors at a human sperm bank in China.

Background: In China, numerous human sperm banks only perform three-generation family history evaluation to exclude genetic diseases with clinical symptoms; therefore, many inherited risks cannot be detected before donor qualification even when a thorough genetic family history evaluation has been performed. Hence, the risk of recessive disease inheritance persists with the current eligibility guidelines in China regarding the donor selection process. Methods: Retrospective study that reviewed the genetic test analyses and clinical outcomes of young adult men who were qualified sperm donors at the Hunan Province Human Sperm Bank of China from January 1, 2018, to May 1, 2021. We included a total of 3231 qualified sperm donors: all donors underwent primary screening for thalassemia and glucose-6-phosphate dehydrogenase (G6PD) deficiency. Whereafter, 278 of donors underwent genetic testing for specific genes, and 43 donors underwent whole exome sequencing. Results: 2.4% of 3231 qualified sperm donors might have thalassemia and 1.4% might have G6PD deficiency. Sperm donors with thalassemia and G6PD deficiency would be eliminated. Specific gene testing identified 7 of the 278 donors (2.5%) as carriers of at least one pathogenic or likely pathogenic variant in a gene, including 1.9% of 154 donors (3/154) as carrier variants in α-Like or ß-Like globin genes, 17.6% of 17 donors (3/17) as carrier variants in GJB2, 12.5% of 8 donors (1/8) as carrier variants in SMN1. In addition, among the 43 sperm donors carrying the 111 pathogenic/likely pathogenic variants, eight (18.6%) were carriers of pathogenic variants of the GJB2 gene. The frequency, therefore, was approximately 1 in 5. Conclusions: The data suggest that used blood routine and RDT can make a preliminary screening of sperm donors, and special gene testing should be performed for sperm donors according to the regional incidence of specific genetic diseases. Meanwhile, whole exome sequencing can be used as a supplementary application in sperm donor genetic testing, and aid a successful and healthy pregnancy. However, industry guidelines must be modified to incorporate its use.

A novel homozygous frameshift mutation in MNS1 associated with severe oligoasthenoteratozoospermia in humans.

Oligoasthenoteratozoospermia (OAT) refers to the combination of various sperm abnormalities, including a decreased sperm count, reduced motility, and abnormal sperm morphology. Only a few genetic causes have been shown to be associated with OAT. Herein, we identified a novel homozygous frameshift mutation in meiosis-specific nuclear structural 1 (MNS1; NM_018365: c.603_604insG: p.Lys202Glufs*6) by whole-exome sequencing in an OAT proband from a consanguineous Chinese family. Subsequent variant screening identified four additional heterozygous MNS1 variants in 6/219 infertile individuals with oligoasthenospermia, but no MNS1 variants were observed among 223 fertile controls. Immunostaining analysis showed MNS1 to be normally located in the whole-sperm flagella, but was absent in the proband's sperm. Expression analysis by Western blot also confirmed that MNS1 was absent in the proband's sperm. Abnormal flagellum morphology and ultrastructural disturbances in outer doublet microtubules were observed in the proband's sperm. A total of three intracytoplasmic sperm injection cycles were carried out for the proband's wife, but they all failed to lead to a successful pregnancy. Overall, this is the first study to report a loss-of-function mutation in MNS1 causing OAT in a Han Chinese patient.

Improving native human sperm freezing protection by using a modified vitrification method.

Slow freezing is the most commonly used technique for the cryopreservation of spermatozoa in clinical practice. However, it has been shown to have a negative impact on sperm function and structure. Vitrification as a successful alternative method has been proved to have better protective effects on human embryos, but vitrification of spermatozoa is still subject to low recovery rates. In this study, a modified vitrification method for native spermatozoa was developed. A total of 28 semen samples were included; each sample was divided into three equal parts and assigned to fresh, slow freezing, and vitrification groups. Sperm vitality, motility, morphology, DNA integrity, and acrosome reaction were assessed for each of the groups. The results showed that vitrification achieves better results for several sperm protection parameters than slow freezing; vitrification achieves a higher recovery rate (P < 0.05), motility (P <0.05), morphology (P <0.05), and curve line velocity (P <0.05) than slow freezing. Furthermore, DNA fragmentation was decreased (P <0.05) and better acrosome protection (P <0.05) was exhibited in the spermatozoa after vitrification. Principal component analysis of all sperm parameters revealed that the vitrification cluster was closer to the fresh cluster, indicating that spermatozoa are better preserved through vitrification. In conclusion, while both slow freezing and vitrification have negative effects on sperm function and structure, the vitrification protocol described here had a relatively better recovery rate (65.8%) and showed improved preservation of several sperm quality parameters compared with slow freezing.

Novel homozygous truncating variants in ZMYND15 causing severe oligozoospermia and their implications for male infertility.

Sequence variants of ZMYND15 cause azoospermia in humans, but they have not yet been reported in infertile men with severe oligozoospermia (SO). We performed whole-exome and Sanger sequencing to identify suspected causative variants in 414 idiopathic participating infertile men with SO or azoospermia. Three novel homozygous truncating variants in ZMYND15 were identified in three of the 219 (1.37%) unrelated patients with SO, including c.1209T>A(p.Tyr403*), c.1650delC (p.Glu551Lysfs*75), and c.1622_1636delinsCCAC (p.Leu541Profs*39). In silico bioinformatic analyses as well as in vivo and in vitro experiments showed that the ZMYND15 variants carried by the affected subjects might be the underlying cause for their infertility. One patient accepted intracytoplasmic sperm injection therapy, using his ejaculated sperm, and his wife successfully became pregnant. Our findings expand the disease phenotype spectrum by indicating that ZMYND15 variants cause SO and male infertility and suggest a possible correlation between the severity of male infertility caused by ZMYND15 variants and male age.

[Trigger effect of hMG and hCG in the treatment of unexplainable non-obstructive azoospermia].

OBJECTIVE: To investigate whether the trigger effect of human menopausal gonadotropins (hMG) and human chorionic gonadotropins (hCG) attributes to the treatment of unexplainable non-obstructive azoospermia (NOA). METHODS: We retrospectively analyzed the clinical data about 282 cases of unexplainable NOA treated in the Maternity and Child Health Hospital of Guizhou Province from January 2010 to May 2017. All the patients underwent trigger treatment by intramuscular injection of hMG at 75 IU 3 times a week for 2 weeks, followed by hCG at 2 000 IU twice a week for another 2 weeks, and meanwhile took vitamin E, Levocarnitine and Tamoxifen as an adjunctive therapy. The treatment lasted 3－12 months. RESULTS: Fifty-eight of the 255 patients that completed the treatment were found with sperm in the semen after treatment, all with severe oligoasthenospermia. Forty-seven of the 58 cases received assisted reproductive technology (ART), of which 18 achieved clinical pregnancy. Semen centrifugation revealed no sperm in the other cases, of which 6 were found with epididymal sperm at epididymal and testicular biopsy after treatment and 3 of them achieved clinical pregnancy after ART. Sperm was found in the semen or at epididymal or testicular biopsy in 64 of the patients after treatment, with an effectiveness rate of 25.1%. CONCLUSIONS: Trigger treatment by injection of hMG and hCG combined with adjunctive oral medication has a certain effect on unexplainable NOA.

[Karyotype analysis of qualified sperm donors on preliminary screening].

OBJECTIVE: To explore the significance of karyotype analysis in screening sperm donors. METHODS: From January 1, 2004 to December 31, 2008, a total of 2537 potential sperm donors passed our preliminary screening, and all were routinely karyo-typed via peripheral blood. Follow-ups were conducted on the pregnancy outcome and congenital malformation after artificial insemination with the sperm from the qualified donors. RESULTS: Among the 2537 qualified sperm donors, 2362 were of the normal karyotype 46, XY and 135 showed polymorphism. Abnormal karyotype was found in 6 cases, and controversial abnormal karyotype in 34. CONCLUSION: Karyotype analysis can reduce the risk of chromosomal disease in neonates from artificial insemination, and genetic counseling for abnormal karyotype sperm donors may help them solve their future reproductive problems.