A modified RT-PCR technique to screen for viral RNA in the semen of hepatitis C virus-positive men.

BACKGROUND Our objective was to use an adapted RT-PCR technique to assess the presence of hepatitis C virus (HCV) in semen and also in different density gradient semen fractions collected from men with chronic viral hepatitis participating in an assisted reproduction programme. METHODS This study included 50 semen samples from 35 HCV(+) men, with active viral replication assessed by RT-PCR, collected the day of oocyte retrieval and used for assisted reproduction. These samples were subjected to standard assisted reproduction sperm preparation conditions, using density-gradient centrifugation with 45 and 90% layers. Aliquots of semen, 45 and 90% fractions, and embryo culture media were frozen at -80 degrees C for subsequent virological analyses. All aliquots were tested with a commercially available HCV RNA assay, adapted for use with semen after a number of technical changes. This assay yielded a sensitivity of 50-100 HCV RNA copies/ml and strongly diminished the effect of seminal amplification inhibitors. RESULTS HCV RNA was detected in 7/50 (14%) semen samples tested, 5/35 (14.3%) men. HCV RNA was found in only 1/50 45% fractions but never in the 90% fraction or embryo culture media. Sera from 3/5 men contained 3.19-7.40 x 10(5) IU/ml, while the two others had 4.5 and 11.7 x 10(6) IU/ml. However, HCV RNA was quantified at <600 IU/ml in the HCV(+) semen of these five patients. The ongoing pregnancy rate was of 20% (10/50) with one delivery at the time of the present report. No anti-HCV antibody was found in any of the women or the newborn. CONCLUSIONS Although HCV is present at low concentrations in the semen of a few HCV(+) patients, no purified sperm fraction (i.e. 90% fraction) used in assisted reproduction was HCV(+) and no seroconversion was observed in the women and the newborn, thereby suggesting a very low risk of virus transmission. Nevertheless, because the presence of HCV in semen implies a possible risk of nosocomial contamination, safety regulations must be strictly applied in assisted reproduction laboratories.