Semen parameters of a semen donor before and after infection with human immunodeficiency virus type 1: Case report

E. van Leeuwen1,4, M. Cornelissen2, J.W. de Vries1, S.H. Lowe3, S. Jurriaans2, S. Repping1 and F. van der Veen1

1 Center for Reproductive Medicine, Department of Obstetrics and Gynecology, 2 Department of Human Retrovirology and 3 Department of Internal Medicine, Division of Infectious Diseases, Tropical Medicine and AIDS, Academic Medical Center, Amsterdam, The Netherlands

4 To whom correspondence should be addressed at. Academic Medical Center, Obstetrics and Gynecology, H4-205, Postbus 22660, 1100 DD Amsterdam, The Netherlands. Email: e.vanleeuwen{at}amc.uva.nl


    Abstract
 Top
 Abstract
 Introduction
 Case report
 Discussion
 Acknowledgements
 References
 
Semen samples from a donor who seroconverted for human immunodeficiency virus type 1 (HIV-1) during the period that he was donating at our clinic were stored before and after infection. Semen analysis was done on all of these samples before cryopreservation. Retrospectively, both qualitative and quantitative HIV-1 testing was performed on the cryopreserved semen samples to determine the time of primary HIV-1 infection. After HIV-1 infection, semen volume, sperm motility and the percentage of spermatozoa with normal morphology were reduced compared with the same parameters before HIV-1 infection. HIV-1 RNA was intermittently detectable in semen. HIV-1 infection led to a reduction in semen volume, sperm motility and normal sperm morphology in this donor. However, the clinical significance of these findings is unclear. A longitudinal cohort study on the effects of HIV-1 infection on semen quality is necessary to confirm these findings.

Key words: HIV-1/HIV RNA/semen donor/semen quality/seroconversion


    Introduction
 Top
 Abstract
 Introduction
 Case report
 Discussion
 Acknowledgements
 References
 
During the last decade, the effect of a human immunodeficiency virus type 1 (HIV-1) infection on semen quality has been evaluated in several studies (Krieger et al., 1991Go; Crittenden et al., 1992Go; Politch et al., 1994Go; Dondero et al., 1996Go; Dulioust et al., 2002Go). Most data suggest that semen quality is not altered in early HIV-1 infection (Krieger et al., 1991Go; Politch et al., 1994Go; Dondero et al., 1996Go). A drawback of these studies, however, is that all men were already HIV-1 infected when the first semen analysis was performed. Therefore, no comparison can be made between semen parameters before and after infection with HIV-1.

Here we report a unique case of one man who participated in a semen donation programme and became infected with HIV-1, thereby allowing for, what we believe to be, the first time the comparison of semen parameters before and after infection in the same individual.


    Case report
 Top
 Abstract
 Introduction
 Case report
 Discussion
 Acknowledgements
 References
 
Our patient was a Caucasian man, participating in a semen donation programme of the semen bank of the Center for Reproductive Medicine of the Academic Medical Center from 1994 until 1998. He was 44 years old when he entered the programme in 1994. His history revealed a hepatitis A and a gonorrhoea infection in 1981 and a hepatitis B infection in 1982. He denied having homosexual contacts at that time. After an interval of 2 years, he started donating semen again in September 2000, because of requests for second children from this donor by successfully treated couples. At this time, he was found to be HIV negative, but urine ligase chain reaction (LCR) revealed a chlamydia infection that was treated with azithromycin. Subsequently, chlamydial DNA was undetectable in November 2000.

In total, 12 semen samples were collected between September 2000 and June 2001. According to protocol, the donor was screened approximately every 6 months for HIV, hepatitis B/C and chlamydia. Semen was cryopreserved and quarantined, which implies that it is not used for any fertility treatment until a negative test, at least 6 months after the donation, is available. In June 2001, the HIV-1 test was positive. For this reason, semen donation was stopped. For research purposes, he was asked to donate semen again in January 2003.

The donor had acquired the HIV-1 infection through unsafe homosexual contact. The HIV-1 infection was classified as category A1, according to the Centers for Disease Control and Prevention (CDC) classification system of 1993 (Centers for Disease Control and Prevention, 1992Go). Therefore, he did not receive any antiretroviral therapy and was still therapy naive at the end of July 2004. Blood HIV-1 viral load increased from 2192 copies/ml in July 2001 to 97 057 copies/ml in November 2003. CD4 counts were stable over this period, between 540 and 940 cells/mm3. His blood measurements showed antibodies against hepatitis A, hepatitis B, cytomegalovirus (CMV), Epstein–Barr virus (EBV) and toxoplasmosis, with no signs of recent infections. Physical and genital examination revealed no abnormalities.

Table I shows the results of the retrospective HIV-1 testing of the 12 cryopreserved semen samples, which were collected between the last negative HIV-1 test in September 2000 and the first positive HIV-1 test in June 2001. Qualitative HIV-1 RNA screening was done by nested PCR on the cryopreserved semen. The lower detection limit of this test was determined by spiking 2 x 106 spermatozoa from the September 2000 sample with 1– 100 000 HIV particles using HIV reference virus stock (Layne et al., 1992Go). After isolation and amplification, 10 HIV-1 RNA molecules could be detected in the background of 400 000 spermatozoa. The whole isolation and amplification process was repeated once, with the same results. All samples were tested in duplicate.


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Table I. Qualitative HIV-1 RNA detection by nested PCR and quantitative HIV-1 RNA detection on the cryopreserved semen between the last negative and first positive HIV test

 
The qualitative HIV-1 test was positive for the first time in the semen sample of November 9, 2000. The test remained positive until January 2001, but from February 2001 until April 2001, HIV-1 RNA could not be detected. In May 2001, one of the duplicate tests became positive and both tests were positive again in June 2001.

A quantitative real-time HIV-1 RNA assay, developed on an ABI Prism® 7000, was then used to determine the amount of HIV-1 RNA. The lower detection limit of this assay was five copies of HIV-1 RNA molecules per 100 000 spermatozoa. At most time points, undetectable viral loads were observed. Very low viral loads were observed from November until December 2000. During the same period, the donor reported flu-like symptoms. In view of the data summarized in Table I, we conclude that the primary HIV-1 infection occurred at the beginning of November 2000.

In Table II, semen variables before and after HIV-1 infection are presented. According to our protocol for semen donors, all semen samples were produced after 2–5 days of sexual abstinence. After HIV-1 infection, semen volume, sperm motility and the percentage of spermatozoa with normal morphology were reduced compared with before HIV-1 infection. Although spermatozoa concentration increased after HIV-1 infection, semen total count (TC) and total motile count (TMC) did not change.


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Table II. Semen variables before and after HIV-1 infection

 

    Discussion
 Top
 Abstract
 Introduction
 Case report
 Discussion
 Acknowledgements
 References
 
In the donor described in this case report, semen volume, sperm motility and the percentage of spermatozoa with normal morphology were reduced after acquisition of HIV-1. Since no statistical test exists for repeated measurements in one person, these findings cannot be tested statistically. Whether the described changes truly reflect an effect of HIV-1 on testicular function remains unclear, since only one person was involved. Semen parameters are known to be variable on repeated sampling in one man, and the effects of HIV infection on testicular function might differ between individuals.

Nevertheless, the decrease in semen volume could be indicative of dysfunction of the accessory glands; the prostate and seminal vesicles. Dysfunction of the accessory glands could also be an explanation for the more viscous sperm that is often found in HIV-1-infected subjects (Crittenden et al., 1992Go; Dondero et al., 1996Go). A reduction in sperm motility in HIV-1-infected men has been described before (Crittenden et al., 1992Go; Dulioust et al., 2002Go), although an explanation for this finding is lacking. A decrease in the percentage of spermatozoa with normal morphology has been reported in one other study (Muller et al., 1998Go), while others reported normal values for sperm morphology (Krieger et al., 1991Go; Politch et al., 1994Go; Dulioust et al., 2002Go). In this donor, the semen TC and TMC remained stable. The observed increase in semen concentration appeared to be caused by a decrease in semen volume in this donor.

Consistent with most other studies on semen quality of asymptomatic HIV-1-infected men, nearly all semen parameters matched the World Health Organization criteria for normal human semen at all time points in this donor (Krieger et al., 1991Go; Crittenden et al., 1992Go; World Health Organization, 1992Go; Dondero et al., 1996Go).

On PCR testing, the amount of virus was expressed per 105 spermatozoa. The spermatozoa were counted and a constant input of 105 spermatozoa was used for PCR testing. Each straw also contained a low variable number of non-sperm cells (NSCs). The NSCs were calculated using the original concentration of NSCs and a known dilution factor (Table I). The small number of NSCs in the sample most probably had no influence on the sensitivity of the PCR test since the spermatozoa were always by far the majority of cells in the test.

HIV-1 is most probably present as cell-free virus in seminal plasma and as cell-associated virus in seminal NSCs, such as lymphocytes, monocytes and macrophages (Dulioust et al., 1998Go). It seems unlikely that spermatozoa themselves can become infected with HIV-1, since there are no HIV-1 CD4 receptors or co-receptors present on spermatozoa and vasectomy does not influence the amount of cell-free virus in semen (Krieger et al., 1998Go; Kim et al., 1999Go).

In retrospect, HIV-1 was detectable in semen at the same time the patient suffered from flu-like symptoms, indicative of a primary HIV-1 infection. This is consistent with two other case reports where HIV-1 RNA was present in semen during primary HIV-1 infection (Tindall et al., 1992Go; Dyer et al., 1997Go).

On qualitative screening, HIV-1 RNA was intermittently detectable in the semen samples. This pattern of intermittent shedding is often found in HIV-1-positive men (Gupta et al., 2000Go; Bujan et al., 2002Go, 2004Go). It is hypothesized that a fluctuation of viral transcription from infected cells in the prostate is the cause of this intermittent shedding of HIV-1 in semen (Gupta et al., 2000Go). Also local factors such as inflammation of the male genital tract can increase the amount of HIV-1 RNA in semen, probably by an increase of lymphocytes and macrophages (Cohen et al., 1997Go).

Quantitative testing of HIV-1 RNA revealed a very low copy number of HIV-1 RNA during primary infection and an undetectable viral load in semen at all other time points. Because the qualitative test is more sensitive than the quantitative test, it was not always possible to quantify HIV-1 RNA precisely, despite a positive qualitative test. In these samples the quantity is presumably <5 copies per 100 000 spermatozoa. The highest amount of HIV-1 RNA in semen was detected during primary HIV-1 infection, when blood viral load was probably very high. This observation supports our recently published hypothesis that HIV-1 RNA in semen could be spill-over from blood (Lowe et al., 2004Go).

To our knowledge, we are the first to describe semen parameters before and after acquiring HIV-1 in the same individual. In all other studies published, men were already HIV-1 infected by the time the first semen analysis was performed. However, our results show that HIV-1 infection may lead to a reduction in semen volume, sperm motility and the percentage of spermatozoa with normal morphology. The clinical significance of these findings is not clear. Larger longitudinal studies on the effects of HIV-1 on semen quality will have to be carried out to confirm our findings.


    Acknowledgements
 Top
 Abstract
 Introduction
 Case report
 Discussion
 Acknowledgements
 References
 
The authors would like to thank Fokla Zorgdrager for her PCR work. This research has been funded by grant number 7003 from AIDS Fonds Netherlands


    References
 Top
 Abstract
 Introduction
 Case report
 Discussion
 Acknowledgements
 References
 
Bujan L, Daudin M, Alvarez M, Massip P, Puel J and Pasquier C (2002) Intermittent human immunodeficiency type 1 virus (HIV-1) shedding in semen and efficiency of sperm processing despite high seminal HIV-1 RNA levels. Fertil Steril 78, 1321–1323.[CrossRef][ISI][Medline]

Bujan L, Daudin M, Matsuda T, Righi L, Thauvin L, Berges L, Izopet J, Berrebi A, Massip P and Pasquier C (2004) Factors of intermittent HIV-1 excretion in semen and efficiency of sperm processing in obtaining spermatozoa without HIV-1 genomes. AIDS 18, 757–766.[CrossRef][ISI][Medline]

Centers for Disease Control and Prevention (1992) 1993 revised classification system for HIV infection and expanded surveillance case definition for AIDS among adolescents and adults. MMWR Recomm Rep 41, 1–19.[Medline]

Cohen MS, Hoffman IF, Royce RA, Kazembe P, Dyer JR, Daly CC, Zimba D, Vernazza PL, Maida M, Fiscus SA et al. (1997) Reduction of concentration of HIV-1 in semen after treatment of urethritis: implications for prevention of sexual transmission of HIV-1. AIDSCAP Malawi Research Group. Lancet 349, 1868–1873.[CrossRef][ISI][Medline]

Crittenden JA, Handelsman DJ and Stewart GJ (1992) Semen analysis in human immunodeficiency virus infection. Fertil Steril 57, 1294–1299.[ISI][Medline]

Dondero F, Rossi T, D'Offizi G, Mazzilli F, Rosso R, Sarandrea N, Pinter E and Aiuti F (1996) Semen analysis in HIV seropositive men and in subjects at high risk for HIV infection. Hum Reprod 11, 765–768.[Abstract]

Dulioust E, Tachet A, De Almeida M, Finkielsztejn L, Rivalland S, Salmon D, Sicard D, Rouzioux C and Jouannet P (1998) Detection of HIV-1 in seminal plasma and seminal cells of HIV-1 seropositive men. J Reprod Immunol 41, 27–40.[CrossRef][ISI][Medline]

Dulioust E, Du AL, Costagliola D, Guibert J, Kunstmann JM, Heard I, Juillard JC, Salmon D, Leruez-Ville M, Mandelbrot L et al. (2002) Semen alterations in HIV-1 infected men. Hum Reprod 17, 2112–2118.[Abstract/Free Full Text]

Dyer JR, Gilliam BL, Eron JJ, Jr, Cohen MS, Fiscus SA and Vernazza PL (1997) Shedding of HIV-1 in semen during primary infection. AIDS 11, 543–545.[ISI][Medline]

Gupta P, Leroux C, Patterson BK, Kingsley L, Rinaldo C, Ding M, Chen Y, Kulka K, Buchanan W, McKeon B et al. (2000) Human immunodeficiency virus type 1 shedding pattern in semen correlates with the compartmentalization of viral quasi species between blood and semen. J Infect Dis 182, 79–87.[CrossRef][ISI][Medline]

Kim LU, Johnson MR, Barton S, Nelson MR, Sontag G, Smith JR, Gotch FM and Gilmour JW (1999) Evaluation of sperm washing as a potential method of reducing HIV transmission in HIV-discordant couples wishing to have children. AIDS 13, 645–651.[CrossRef][ISI][Medline]

Krieger JN, Coombs RW, Collier AC, Koehler JK, Ross SO, Chaloupka K, Murphy VL and Corey L (1991) Fertility parameters in men infected with human immunodeficiency virus. J Infect Dis 164, 464–469.[ISI][Medline]

Krieger JN, Nirapathpongporn A, Chaiyaporn M, Peterson G, Nikolaeva I, Akridge R, Ross SO and Coombs RW (1998) Vasectomy and human immunodeficiency virus type 1 in semen. J Urol 159, 820–825.[CrossRef][ISI][Medline]

Layne SP, Merges MJ, Dembo M, Spouge JL, Conley SR, Moore JP, Raina JL, Renz H, Gelderblom HR and Nara PL (1992) Factors underlying spontaneous inactivation and susceptibility to neutralization of human immunodeficiency virus. Virology 189, 695–714.[ISI][Medline]

Lowe SH, Sankatsing SU, Repping S, Veen FV, Reiss P, Lange JM and Prins JM (2004) Is the male genital tract really a sanctuary site for HIV?. Arguments that it is not. AIDS 18, 1353–1362.[CrossRef][ISI][Medline]

Muller CH, Coombs RW and Krieger JN (1998) Effects of clinical stage and immunological status on semen analysis results in human immunodeficiency virus type 1-seropositive men. Andrologia 30 (Suppl 1), 15–22.

Politch JA, Mayer KH, Abbott AF and Anderson DJ (1994) The effects of disease progression, zidovudine therapy on semen quality in human immunodeficiency virus type 1 seropositive men. Fertil Steril 61, 922–928.[ISI][Medline]

Tindall B, Evans L, Cunningham P, McQueen P, Hurren L, Vasak E, Mooney J and Cooper DA (1992) Identification of HIV-1 in semen following primary HIV-1 infection. AIDS 6, 949–952.[ISI][Medline]

World Health Organization (1992) WHO Laboratory Manual for the Examination of Human Semen and Sperm-Cervical Mucus Interaction, 3rd edn. Cambridge University Press, Cambridge, UK.

Submitted on January 23, 2004; resubmitted on June 1, 2004; accepted on August 11, 2004.