Journal of Medical Virology 64:499±504 (2001) Complete Replication of Human Cytomegalovirus in Explants of First Trimester Human Placenta Liliana Gabrielli, 1 Luisa Losi, 2 Stefania Varani, 1 Tiziana Lazzarotto, 1 Vincenzo Eusebi, 2 and Maria Paola Landini 1 * 1 Department of Clinical and Experimental Medicine, Section of Microbiology, St. Orsola General Hospital, University of Bologna, Bologna, Italy 2 Department of Oncology, Bellaria Hospital, Bologna, Italy Tissue integrity and viability of ®rst trimester placenta explants were obtained in culture for 3 weeks. Explants were infected with human cytomegalovirus (HCMV), several cycles of HCMV replication were obtained and the pro- gression of the infection was observed within a tissue that maintains its normal cellular organiza- tion. In agreement with recent clinical data, 3 weeks were necessary for the virus to colonize the placenta fully. Complete HCMV replication was observed in trophoblasts, followed by subse- quent transmission of the infection to the stromal ®broblasts and fetal endothelial capillary cells. Viral DNA replication was monitored and the production of infectious viral progeny documen- ted. J. Med. Virol. 64:499 ± 504, 2001. ß 2001 Wiley-Liss, Inc. KEY WORDS: cytomegalovirus; placenta explants; trophoblast INTRODUCTION Human cytomegalovirus (HCMV) is the leading cause of congenital infection in developed countries, occurring in 0.3±2% of all live births [Peckham, 1991]. More than 10±15% of congenitally infected newborns are symptomatic at birth, and 10±15% of the asympto- matic infected infants will develop long-term neurologic sequelae such as mental retardation, deafness and visual impairment, thus making CMV the leading infectious cause of CNS damage in children [Boppana et al., 1992]. It is generally believed that intrauterine transmis- sion occurs transplacentally and that the virus spreads by blood from mother to fetus [Benirschke and Kaufmann, 1990]. Transplacental viral transmission has been hypothesized for three main reasons: 1) congenital HCMV infections are associated commonly with chronic villitis [Altshuler and Russell, 1975; Garcia et al., 1989], 2) HCMV infection of villous stroma, endothelial cells and macrophages has been shown in placenta tissue from fetuses infected congeni- tally [Sinzger et al., 1993], and 3) productive HCMV infection of the trophoblasts has been demonstrated in in vitro-cultured primary villous trophoblast cells [Halwachs-Baumann et al., 1998; Hemmings et al., 1998]. Little is known, however, about the mechanism of HCMV passage from maternal blood to fetal tissues or the progression of infection at placental level. In vitro studies have unequivocally demonstrated that human trophoblasts are permissive to the complete replicative cycle of HCMV in primary cultures [Fisher et al., 2000; Halwachs-Baumann et al., 1998; Hem- mings et al., 1998]. Nevertheless, a complete HCMV replication could not be demonstrated in trophoblasts of in vivo infected placentas from congenitally infected fetuses [Sinzger et al., 1993]. Neither immunohisto- chemical studies of human placenta from fetuses infected congenitally nor primary in vitro cultures of trophoblasts allow the evaluation of how the virus spreads into the placental tissue. In this study explants of ®rst trimester human placentas were infected in vitro and used to investigate the permissiveness of trophoblasts and other fetal cells in an organotropic model in which a normal tissue organization is maintained. MATERIAL AND METHODS Placenta Explants Placenta tissues of 12 weeks' gestation were obtained from elective abortions by vacuum aspiration. The tissues were washed ®ve times with cold sterile physiological solution containing vancomycin (100 mg/ ml), penicillin (1,000 U/ml), fungizone (25 mg/ml) and Grant sponsor: Ministry of Public Health (ISS, AIDS Projects); Grant sponsor: Ministry of Education; Grant sponsor: University of Bologna; Grant sponsor: St. Orsola General Hospital. *Correspondence to: Maria Paola Landini, MD, Laboratorio di Virologia, Policlinico S. Orsola - Malpighi, Via Massarenti n.9, 40138 Bologna, Italy. E-mail: viroland@med.unibo.it Accepted 26 September 2000 ß 2001 WILEY-LISS, INC.