Introduction
In late 2019, an outbreak of pneumonia caused by a newly discovered coronavirus called Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) originated in Wuhan, China . This virus spread quickly worldwide and let to a severe acute respiratory syndrome known as ”COVID-19” as designated by the World Health Organization (WHO). The rapid transmission of this novel virus resulted in the WHO declaring it a pandemic, causing significant changes in social behavior. COVID-19 has had a profound impact on the health of affected individuals and has also greatly affected healthcare delivery for all patients, not just those with COVID-19.
Coronaviruses have a spherical shape awith spike proteins on their envelope, giving them a club-like appearance. The virus contains a single-stranded RNA genome with a positive sense, which is enclosed within a helical nucleocapsid. The entry of SARS-CoV-2 into cells is mediated by the viral glycoprotein Spike binding to the enzymatic domain of angiotensin-converting enzyme 2 (ACE-2) on the cell surface . The virus is then internalized through endocytosis by another membrane bound protease called transmembrane serine protease 2 (TMPRSS2) . Several testicular cells, including Sertoli, Leydig cells and spermatogonia cells, express the ACE-2 receptor on their surface along with proteins belonging to the angiotensin pathway (ATR1, ATR2, ACE1, and TMPRSS2) . There receptors and proteins are crucial for SARS-CoV-2 infection. Given that other RNA viruses such as Ebola, Marburg, and Zika, have been found to breach the blood-testis barrier and cause inflammation , it was hypothesized that the male reproductive system could also be a target for SARS-CoV-2. Additionally, numerous studies have reported that viral infections significantly contribute to long-term male infertility. Viruses like HPV (papillomavirus), HSV (herpes simplex virus), and HIV (immunodeficiency virus) can infect the male reproductive system, leading to severe consequences for reproductive health , possibly due to triggering an inflammatory response that result in the release of inflammatory cytokines and subsequent oxidative stress.
In light of these considerations, we conducted an investigation into the inflammatory status of the testis by evaluating the presence of inflammatory cytokines and immune cell populations in seminal fluid after viral clearance.
To date, there has been limited research on the mechanisms responsible for the observed effects of COVID-19 infection on semen quality. However, a recent study has provided valuable insights, confirming that COVID-19 infection can reduce semen volume, progressive motility, morphology, sperm count, and DNA integrity .
Preserving male reproductive cells, including semen, epididymal, and testicular spermatozoa, through cryopreservation, is a crucial approach for managing permanent or temporary infertility in young men and children. Cryopreservation technology, in particular, plays an important role in preserving fertility for these patients . It remains uncertain whether viral particles persist after cryopreservation, and this question is especially relevant for cryopreserved samples processed during the COVID outbreak. Therefore, we investigated the presence of SARS-CoV-2 genome in seminal fluid before and after cryopreservation using two methods: conventional slow freezing and vitrification. While slow freezing is a commonly employed technique in assisted reproductive technology, seminal fluid vitrification may offer a more efficient and faster approach for sperm vitrification. However, due to the risk of contamination from direct contact with liquid nitrogen, sperm vitrification is considered a hazardous procedure .13,271113–19
Our aim was to investigate the effect of COVID-19 on the male reproductive tract. We collected semen samples from 20 patients, with a median interval of 3 months from SARS-CoV-2 infection. Our findings demonstrated that both ACE2 and TMPRSS2 were detectable in both the plasma and sperm fractions even months after the viral infection. Additionally, we found viral RNA-dependent RNA polymerase (RdRp) in 5 out of 20 patients, indicating the potentional presence of the virus in the body three months after the initial infection. The study also revealed high levels of inflammatory cytokines and chemokines in the plasma seminal fraction of individuals exposed to SARS-CoV-2-, suggesting the persistence of an inflammatory condition that could impact spermatozoa vitality. We also identified immune populations in the semen fraction that are known to play a role in clearing viral infection, and these populations positively correlate with the expression of of SARS-CoV-2 receptors and inflammatory cytokines.
Furthermore, we investigated the potential implicaitons of SARS-CoV-2 in reproductive medicine, particularly in medically assisted reproduction and gamete cryopreservation. We examined the presence of SARS-CoV-2 genome in seminal fluid before and after cryopreservation using two methods: conventional slow freezing and vitrification. Our findings demonstrate that both conventional freezing and vitrification are safe procedures for preserving male fertility in assisted reproductive technology programs.