- Introduction: The prevalence of viral infections in the amniotic fluid (AF) has not yet been ascertained. The aim of this study was to determine the prevalence of specific viral nucleic acids in the AF and its relationship to pregnancy outcome. Study design: From a cohort of 847 consecutive women undergoing midtrimester amniocentesis, 729 cases were included in this study after exclusion of documented fetal anomalies, chromosomal abnormalities, unavailability of AF specimens and clinical outcomes. AF specimens were tested by quantitative real-time PCR for the presence of genome sequences of the following viruses: adenoviruses, herpes simplex virus (HSV), varicella zoster virus (VZV), human herpesvirus 6 (HHV6), human cytomegalovirus (HCMV), Epstein-Barr virus (EBV), parvovirus B19 and enteroviruses. Viral nucleic acid testing was also performed in maternal blood and cord blood in the population of women in whom AF was positive for viruses and in a control group of 29 women with AF negative for viral nucleic acids. The relationship between the presence of viruses and pregnancy and neonatal outcome was examined. The correlation between the presence of nucleic acids of viruses in the AF and the concentration of the cytokine interleukin-6 (IL-6) and the T cell chemokine CXCL-10 (or IP-10) in AF and maternal blood were analyzed. Results: Viral genome sequences were found in 16 of 729 (2.2%) AF samples. HHV6 was the most commonly detected virus (7 cases, 1.0%), followed by HCMV (6 cases, 0.8%), parvovirus B19 (2 cases, 0.3%) and EBV (1 case, 0.1%), while HSV, VZV, enteroviruses and adenoviruses were not found in this cohort. Corresponding viral DNA was also detected in maternal blood of six out of seven women with HHV6-positive AF and in the umbilical cord plasma, which was available in one case. In contrast, viral DNA was not detected in maternal blood of women with AF positive for parvovirus B19, HCMV, EBV or of women with AF negative for viruses. HHV6 genome copy number in AF and maternal blood was consistent with genomic integration of viral DNA and genetic infection in all women. There was no significant difference in the AF concentration of IL-6 and IP-10 between patients with and without VIAC. However, for HCMV, there was a significant relationship between viral copy number and IP-10 concentration in maternal blood and AF. The group of women with AF positive for viral DNA delivered at term healthy neonates without complications in 14 out of 16 cases. In one case of HHV6 infection in the AF, the patient developed gestational hypertension at term, and in another case of HHV6 infection in the AF, the patient delivered at 33 weeks after preterm premature rupture of membranes (PPROM). Conclusion: Viral nucleic acids are detectable in 2.2% of AF samples obtained from asymptomatic women in the midtrimester. HHV6 was the most frequently detected virus in AF. Adenoviruses were not detected. Vertical transmission of HHV6 was demonstrated in one case.