The case against an association between HIV-1 and sperm: molecular evidence
Introduction
It is estimated that over thirty million individuals worldwide have been infected with Human Immunodeficiency Virus type 1 (HIV-1), and over 70% of these have acquired their infection from genital or anal intercourse (UNAIDS Fact Sheet, 1996). Despite these statistics and the fact that HIV transmission remains unabated in many regions of the world, data is either lacking, or controversial, concerning which components of genital tract secretions are associated with infectious virus and how HIV-1 is transmitted across mucosal surfaces (Anderson, 1992, Scofield, 1992). Studies with the most dichotomous results are those that have investigated whether HIV binds to, or infects, spermatozoa. Inherent problems with many of these studies are the use of methods such as electron microscopy which are relatively subjective in their analyses, and the assessment of virus in semen sub-populations which have been improperly purified. A critique on the use of microscopic techniques is presented in this issue by Pudney and co-workers (Pudney et al., 1998). The objectives of this paper are (a) to review the techniques used to purify cell populations from semen, and (b) to present a review of the literature and data from our laboratory on the measurement of HIV proviral DNA and RNA in semen cell subpopulations by polymerase chain reaction (PCR) methodology.
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Isolation and characterisation of semen cell sub-populations
The four major populations of cells in semen are spermatozoa, immature germ cells, leukocytes and epithelial cells. A study in our laboratory of 17 normal men revealed highly variable numbers of each sub-population when assessed by immunocytochemistry (Table 1). Other studies from our laboratory indicate that semen cellular parameters change in HIV seropositive men, but that this is highly influenced by disease stage and treatment. Sperm parameters are generally within the normal range in
Measurement of HIV DNA and RNA in semen cell sub-populations
To address the question of which semen cell sub-populations are associated with HIV-1, we have undertaken a substantial number of experiments involving either HIV-1 negative ejaculates spiked with virus, or semen samples from HIV-1 positive individuals.
In a series of experiments, we pooled semen samples from healthy seronegative men, and incubated the washed cellular fraction with various concentrations of macrophage tropic and T cell tropic HIV for 1 h at 37°C. Subsequent to removal of an
Factors affecting viral load in semen
The concentration of HIV in semen varies considerably between individuals. Higher viral loads are associated with CD4 counts of <200 (Anderson et al., 1992, Xu et al., 1997), with concomitant sexually transmitted infections (Cohen et al., 1997) and with asymptomatic genital tract inflammation as assessed by PMN numbers (Anderson et al., 1992, Xu et al., 1997). In one study of HIV-1 seropositive men without symptomatic genital tract infections, all men with semen leukocyte counts over one
Conclusions and future directions
Clearly, further studies are needed to address issues of sexual transmission of HIV. In particular, well-designed studies using samples from a greater number of patients infected with different clades of HIV-1 should continue to address whether HIV is incorporated into, or bound to, cells other than leukocytes in semen. Studies should be made on fresh samples using techniques which ensure purity of cell populations and sensitive, reproducible techniques to quantify virus. A greater
Acknowledgements
This research was financially supported by contract CSA-88-020 from the Contraceptive Research and Development Program (CONRAD) of the US Agency for International Development, which receives funds for AIDS research from an interagency agreement with NICHD, and grants AI-35564 and HD-33276 from the National Institutes of Health (NIH) The views of the authors do not necessarily reflect the views of CONRAD. The authors wish to thank Joseph Politch, PhD, for the graphics.
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