Immunologic causes for poor outcome of pregnancy complicated by primary cytomegalovirus (CMV) infection are only partially understood. Maternal and pup tumor necrosis factor (TNF) and natural killer cell (NK)-like activity associated with primary gestational CMV infection initiated in either the first or third trimester equivalent in the inbred guinea pig model were investigated. Poor pregnancy outcome defined as fetal resorptions, premature delivery, stillbirths, and intrauterine growth retardation occurred with infection at either gestational time. Induction of TNF and NK activity by CMV infection during pregnancy correlated with resorptions in early pregnancy infection and with premature labor in late pregnancy infection. Stillbirths occurred with CMV infection at either time. Regardless of the gestational time of CMV acquisition, poor outcome correlated with higher maternal NK and TNF responses during the first weeks after maternal virus acquisition. Furthermore, CMV infected dams with loss of ≥ 50% of conceptus had higher TNF responses than infected dams with < 50% conceptus loss. Likewise, pups born in litters from CMV-infected dams with resorptions and/or premature labor also had enhanced NK activity and TNF response to CMV compared with pups born to dams not having resorptions or premature labor. TNF responses in the delivered pups of infected dams were higher than from pups of uninfected dams regardless of litter outcome, whereas pup NK responses were enhanced only in pups from litters of dams with premature labor or resorptions. Enhanced NK and TNF activity appear to be associated with premature delivery and other poor outcomes of pregnancy. J. Med. Virol. 60:230–236, 2000. © 2000 Wiley-Liss, Inc.