Isolation of rubella virus from products of conception

The products of conception from 50 therapeutic abortions for maternal rubella in either the first or second trimester were studied using virus isolation procedures. Rubella virus was isolated from 32 (64 per cent) of the specimens. Within the limits of the study it could be demonstrated that the rubella virus was capable of infecting the products of conception, and that once infection was established the virus persisted within the products of conception as late as the seventy-seventh day following the onset of rash.For 8 infected fetuses it was possible to study individual organs. The placenta was involved in all 8 cases. In addition, isolations were achieved from brain, eye, inner ear, heart, thymus, lung, kidney, adrenal glands, gonads, spleen and the gastrointestinal tract in 2 cases, and from the heart in 1 case.This study suggests that the establishment of infection by the rubella virus within the placenta may be a very frequent phenomenon with maternal rubella in the first 4 months of gestation. The possible critical role of the placenta in determining whether infection of other fetal tissues occurs is discussed.     

Picornavirus infection in early murine gestation: significance of maternal illness

To evaluate whether maternal illness following picornavirus infection during pregnancy adversely affects placental and fetal health, mice were inoculated with the GDVII strain of Theiler's murine encephalomyelitis virus or control cell lysate during days 4-7 of gestation. Gross appearance, histopathology and viral culture, and in situ hybridization positivity of placentae and fetuses from ill GDVII-infected, healthy GDVII-infected and control mice were compared. Twenty of 34 (59 per cent) GDVII-infected dams became clinically ill. More placenta-fetus pairs from ill mice were grossly abnormal (68 per cent) than from well GDVII-infected (51 per cent;P< 0.01) or control mice (9 per cent;P< 0.001). Virus was detected by in situ hybridization in 73 per cent of placentae and 29 per cent of fetuses from sick GDVII-infected dams, and in 85 per cent of placentae and 19 per cent of fetuses from healthy GDVII-infected mice (differences not significant). Histological abnormalities consisting of necrosis or an increase in hyaline tissue in the vascular labyrinth layer were similarly frequent in placentae from ill and well GDVII-infected mice (58 per cent versus 67 per cent, P=0.5). Viral RNA, inflammation and necrosis were evident in the heart, great vessels, brain and spinal cord of GDVII-infected fetuses. Infection with GDVII in early pregnancy produces a high rate of gross placental and fetal abnormalities. The rate of gross abnormalities exceeds the incidence of fetal infection and more closely parallels the rates of infection and histopathology in the placenta, suggesting that much of the damage to placenta-fetus pairs is a consequence of placental infection. In addition, the occurrence of viral-induced maternal illness is associated with additive risk to placental and fetal health not explained by an increased rate of placental or fetal infection.     

Protection of murine gestational tissues from picornavirus infection in the preimplantation period

Mice were inoculated with Theiler's murine encephalomyelitis virus (TMEV) on gestational days 1-3 (pre-implantation) or days 4-5 (peri- or post-implantation) or with control cell lysate (days 1-5). Dams were subsequently sacrificed between days 11-14 of gestation, and placentae and fetuses were harvested. Few placentae from dams inoculated with virus on days 1-3 were positive by virus culture (2 per cent) or in situ hybridization (6 per cent), and no fetuses were positive by either technique. In contrast, most placentae from dams inoculated with virus on days 4-5 were virus-positive by culture (96 per cent) or in situ hybridization (100 per cent), and a moderate number of fetuses were also positive (30 per cent by culture, 19 per cent by in situ hybridization). Necrosis was present more frequently in placentae from mice inoculated with virus on days 4-5 (55 per cent) than in placentae from dams inoculated with virus on days 1-3 (19 per cent) or with control cell lysate (18 per cent). Viral infection, mononuclear inflammation and cell necrosis were identified in the heart and great vessels of TMEV-infected fetuses. These results indicate that gestational tissues are largely protected from viral infection before implantation. After implantation, gestational tissues are more readily infected and damaged by maternal picornavirus infection.     

Serum- and cell-mediated immune protection of mouse placenta and fetus against a Brucella abortus challenge: expression of barrier effect of placenta

When mice are intravenously inoculated with a virulent Brucella abortus strain at day 12 to 14 of pregnancy and killed three to five days later, colonization of placentae, fetuses and spleens can be estimated by the frequency and level (bacterial count) of infection and by linkage between individual placental and paired fetal infections. This linkage indicates the placental barrier effect, defined as the number of non-infected fetuses linked to 100 colonized placentae. Immune mice serum raised against two Brucella fractions injected one day before challenge (1) restricted the placental colonization (the dose required to infect 50 per cent placentae was increased by 50 to 70 times compared to controls), (2) decreased the level of splenic and placental infection, and (3) increased the barrier effect so that most fetuses were protected even when linked to a heavily infected placenta. Immune (B + T) spleen cells from mice vaccinated with a Brucella cell-wall fraction transferred to recipients seven to eight days before mating, that is, 22 days before challenge (1) restricted the frequency of placental and fetal colonization, (2) decreased the level of splenic, placental and fetal infections, and (3) increased the barrier effect. However, separated B- and T-cells were less active, in particular on the level of fetal infection. In contrast with serum, the cells did not decrease infection of the fetuses linked to heavily infected placentae.     

Effects of cytomegalovirus infection in pregnant women to fetuses: study with DNA-DNA hybridization method]

DNA of cytomegalovirus (CMV) was examined in 131 placentae and 28 umbilical blood specimens by DNA-DNA hybridization. The result revealed that CMV DNA was detected in 4 of 50 placentae from abnormal fetuses (31 fetal deaths, 19 fetal deformities). 77 placentae from normal fetuses showed negative results. One of 2 cord blood samples from fetal deformities showed CMV DNA positive. 25 umbilical blood samples from normal term newborns showed negative. 4 placentae and 1 cord blood sample from premature infants showed negative results. The results indicate that CMV may play a great role in fetal death and fetal deformity through the infected maternal-fetal circulation.     

Placental pathology in congenital rubella

Two groups of placentae from 18 cases of maternal rubella were examined morphologically and virologically. Placentae in Group I (four cases) had a mean gestational age of 21 +/- 1.9 weeks, whilst those in Group 2 (14 cases) had a mean gestational age of 38 +/- 2.8 weeks. A tendency to hypoplasia was observed. The microscopic lesions were similar to those found in other viral infections but in each group some specific features were noted. Only placentae of Group I showed nodules of villi agglutinated by fibrin. This lesion suggested recent maternal infection. Attention is drawn to the presence of abnormal areas of lobular rarefaction due to dysmaturity of villous stem and terminal villi. This aspect was more diffuse and accentuated in Group 2 placentae. Villitis of reactive, necrotic, proliferative and reparative types was seen only in placentae of Group 2. Devastating villitis was not observed. Inclusions in placental cells suggested rubella infection. The lesions were non-specific and hence stress the need for virological examination of the placenta, immunofluorescence studies and electron microscopy to confirm the diagnosis.     

Prenatal detection of rubella-specific IgM in fetal sera

Serum specimens were obtained by fetoscopy at 19-25 weeks' gestation from four fetuses whose mothers had had confirmed rubella earlier in pregnancy. They were tested for rubella-specific IgM by antibody capture radioimmunoassay. No specific IgM was detected in one fetus and a healthy infant was delivered at term. Specific IgM was detected in the other three fetuses. In one case the level was low (1 unit) and this pregnancy went to term resulting in a neonate with clinical and laboratory evidence of congenital rubella infection. The remaining two fetuses had 2.8 and 2.4 units of specific IgM and the pregnancies were terminated. Blood obtained from these two fetuses after abortion showed levels of 5.4 and 2.9 units respectively. No specific IgM was detected in sera from eleven other fetuses aborted because of maternal rubella but five of these cases were terminated before 19 weeks and in five the interval between rash and abortion was three weeks or less. The results show that the human fetus can produce detectable specific IgM antibody by 19-20 weeks' gestation after exposure to rubella several weeks earlier. However, a larger study is required to define the reliability of fetoscopic blood sampling for the diagnosis of intrauterine infection.     

Detection of rubella virus in amniotic fluid by electron microscopy

We report in this paper the observation of rubella virus by electron microscopy in an amniotic fluid sample, collected from a pregnant woman with rubella infection. Virological investigations by inoculation of cell cultures with amniotic fluid and fetal blood remained negative, due probably to the presence of neutralizing antibodies in the samples. Electron microscopy is a rapid but weakly sensitive method to detect viruses in clinical specimens. However, this unusual observation would indicate that in some cases electron microscopy could be a useful technique to evidence a fetal rubella infection.     

Villous oedema of the placenta: a clinicopathological study

Villous oedema was observed in 259 placentae among 1925 consecutive singleton pregnancies of greater than 19 weeks gestation. It was present in 11 per cent of term placentae in which significant associations with fetal and neonatal death (P less than 0.03), and absence of maternal cigarette smoking (P less than 0.002) were found. In preterm placentae, the oedema was usually more severe, and its prevalence increased from 20 per cent for 33-37 weeks to 40 per cent for less than 33 weeks. Our analysis showed that for a given gestational age, villous oedema was not significantly related to chorioamnionistis, Apgar scores of less than 7 at 1 and 5 min, or neonatal death, an exception was for 33-37 weeks gestation, in the absence of chorioamnionitis, villous oedema was associated with low 1 min Apgar score. Immature intermediate villi are present in premature placentae as a normal developmental stage and in dysmature placentae as a result of villous maldevelopment. Since villous oedema closely resembles the 'stromal channels' in this villous type and shows significant association with prematurity and villous dysmaturity, we postulate that villous oedema is a lesion primarily of the immature intermediate villi. Both fetal and maternal factors are involved in its pathogenesis.     

Experimental primary cytomegalovirus infection in pregnancy: timing and fetal outcome

In contrast to intrauterine rubella infection, the relationship between timing of maternal cytomegalovirus (CMV) infection and fetal outcome has not been clearly defined. In order to investigate this relationship, a guinea pig model was utilized to assess the fetal consequences of maternal CMV infection during the first, second, or third trimester of pregnancy. Congenital infection occurred in 24 of 35 newborn guinea pigs (69%) delivered to mothers infected during the third trimester, with localization of virus to salivary gland in 17 of the 24 infected newborn guinea pigs. In contrast, only one of 28 (5%) progeny sacrificed following first-trimester maternal infection was congenitally infected (p less than 0.01). Second-trimester maternal infection was associated with an intermediate risk of intrauterine infection with transmission of virus to 17 of 54 progeny (33%) (p less than 0.01). Eight of the 10 fetuses delivered after second-trimester infection had virus in multiple organs including the brain. These data suggest that timing of maternal CMV infection is an important variable affecting fetal outcome, with increased risk of intrauterine infection when maternal infection occurs late in pregnancy. However, if fetal infection occurs earlier in pregnancy, it appears to present a greater threat to the fetus, with the potential for dissemination of virus in multiple fetal tissues, including the brain.     

Fetal and maternal white cells and B- and T-lymphocyte subpopulations in pregnant women with recent infection.

OBJECTIVE: To study maternal and fetal white cell counts, B- and T-lymphocyte subpopulations in pregnant women with evidence of recent infection. METHODS: Thirty-seven pregnant women with recent infection and 38 controls were studied. All were referred for fetal blood sampling to exclude congenital infection, or to perform fetal chromosome analysis. There were 16 infected fetuses: 9 cytomegalovirus (CMV), 4 rubella, and 3 toxoplasmosis. Maternal and fetal blood was taken and white cell counts, the percentage of CD3+, CD4+, CD8+, CD56+, HLADR+CD3+ T-lymphocyte subpopulations and CD19+ B lymphocytes were measured. RESULTS: The percentage of CD3+, CD8+, and HLADR+CD3+ lymphocytes were significantly higher in infected mothers compared to controls, while CD19+ and the CD4+/CD8+ ratio were lower. Infected mothers carrying infected fetuses had significantly lower white blood cell counts compared to those infected mothers without fetal infection. The percentage of HLADR+CD3+ T lymphocytes was significantly higher and the CD4+/CD8+ ratio lower in infected fetuses compared to controls and noninfected fetuses of infected mothers. Abnormal CD4+/CD8+ ratios and/or increased HLADR+CT3+ T lymphocytes were found in 8 of 10 fetuses with structural abnormalities and/or hematological/biochemical signs of systemic damage, and in 7 of 27 without (RR = 3.1, 95% CI = 1.5-6.3). CONCLUSION: Both infected fetuses and their mothers have significant identifiable changes in white cell counts and T-lymphocyte subpopulations compared to controls. These tests may help in diagnosing maternal and fetal infection.     

A prospective study of primary cytomegalovirus infection in pregnant women

During a four year study, sera were obtained from 5575 women attending for antenatal care and 3188 (57.2 per cent) were shown to possess complement fixing antibodies to cytomegalovirus (CMV). A total of 1608 seronegative women were followed to term and 14 (0.87 per cent) primary CMV infections occurred in either the second or third trimester. Transplacental spread of CMV occurred in 3 out of 12 (25 per cent) of the 14 babies born to infected mothers. All 14 babies were apparently normal at birth but short term clinical follow-up has already revealed that one child has impaired hearing whilst another is microcephalic. The women were also monitored serologically throughout pregnancy for evidence of rubella infections. Only 12 infections were detected and 7 of these occurred during the large rubella epidemic of 1978. Of the 10 pregnancies which were allowed to proceed to term, transplacental spread of rubella virus occurred in 1 out of 7 (14 per cent). During this study period, CMV infections occurred as frequently as did rubella infections. We therefore conclude that, apart from those years when extensive epidemics occur, many more pregnant women are infected with CMV than with rubella virus.     

Virological studies on the feto-maternal tissues infected with rubella virus

In studies on the congenital rubella syndrome, trans-placental rubella virus (RV) infection was investigated in vitro with human chorionic, decidual and fetal tissues obtained by artificial abortion from RV-infected pregnant women showing high hemagglutination-inhibiting and complement-fixing RV antibodies (1:512 and 1:16). RV was isolated from both chorionic (CR) and the fetal cells (FR) derived from RV-infected pregnant woman and the neutralization test disclosed that their antigenicity and biological properties were similar to that of the standard RV strain, M-33. These CR and FR cells showed a constant release of RV ranging from 2 to 4 log10 FFU (focus forming unit)/0.1ml into the culture media. Moreover, positive staining by immunofluorescent technique (IF) over 70 days seems to indicate RV persistent infection in these cells. However, decidual cells derived from RV-infected pregnant woman gave negative results in the RV release and IF staining. The above evidence strongly indicates that the chorionic cells are easily infected and converted to the RV-carrier. One possible mode of trans-placental RV infection is via an initial infection of the chorionic cells followed by the establishment of persistent RV infection.     

胎儿先天性心脏病和风疹病毒感染的关系

目的 探讨和分析孕妇风疹病毒感染与胎儿先天性心脏病(简称先心病)的关系,为提高先天性风疹综合征产前诊断率寻求方法.方法 对超声心动图诊断并要求引产的38例先心病胎儿行脐带穿刺,采用酶联免疫吸附试验测定脐血风疹病毒特异性抗体IgM. 结果 38例先心病胎儿中18例脐血风疹病毒IgM(+),占47.4%,其余20例IgM(-).18例风疹病毒IgM(+)胎儿按心脏异常结构出现频率排序:室间隔缺损10例、肺动脉发育异常9例、房室瓣异常6例、大动脉转位5例、主动脉骑跨4例.IgM(+)和IgM(-)组中属于圆锥动脉分隔异常或圆锥动脉干间隔旋转不足或方向相反的病例数分别为11例(61.1%)和5例(25.0%)(P<0.05). 结论 风疹病毒与胎儿先心病的发生有一定关联,可能影响胎儿心室分隔和圆锥动脉干发育.B超提示胎儿室间隔缺损、肺动脉发育异常、心肌瓣膜异常、大动脉转位时,应考虑到风疹病毒感染的可能.     

Morphological aspects of the placenta in HIV pregnancies

Forty-nine placentae from HIV-seropositive mothers were collected in various hospitals in France and Belgium. Twenty [corrected] placentae with seven fetuses from interrupted pregnancies and 29 [corrected] placentae from spontaneous deliveries, including two stillborns and a set of twins, were studied morphologically. No significant abnormalities were observed in the aborted material. The placentae corresponding to deliveries presented no significant gross abnormalities but the ratio of fetal to placental weight was significantly decreased in the study group compared with the control group (6.13 versus 7.41; P less than 0.001), associated with a congestive and mature aspect of the parenchyma. Histologically a high incidence of chorioamnionitis (43 per cent) was found, contrasting with the absence of villitis. A relative villous hypercellularity was observed in the study group compared with the control group. Ultrastructural studies of 13 placentae corresponding to gestations of 10 to 40 weeks are presented. In six cases, retrovirus-like particles were found at various sites, such as villous fibroblasts, syncytiotrophoblast and endothelial cells, and in the free membranes.     

The risks of early cordocentesis (12-21 weeks): analysis of 500 procedures

Five hundred cordocenteses were performed between 12 and 21 weeks. The indications were thalassaemia (386), rapid karyotyping (97), feto-maternal allo-immunization (10), rubella (6), and toxoplasmosis (1). One hundred and ten pregnancies underwent termination on the basis of the result, while 20 of the 370 pregnancies intended to continue were lost to follow-up. Amongst these were 16 fetal losses (4.3 per cent) and 22 premature deliveries (5.9 per cent); no other complications were reported. Four adverse prognostic factors were identified: (a) cord bleeding; (b) fetal bradycardia; (c) prolonged procedure time; and (d) anterior insertion of the placenta. There was no 'obvious' difference in fetal loss rate with advancing gestation until 19-21 weeks, when the risk of fetal loss decreased to 2.5 per cent.     

Confirmation of CVS mosaicism in term placentae and high frequency of intrauterine growth retardation association with confined placental mosaicism.

About 2 per cent of specimens from chorionic villus sampling (CVS) analysed either on direct preparation of cytotrophoblast cells or after culture of mesenchymal stroma reveal confined placental mosaicism (CPM), most commonly involving chromosomal trisomy. A significantly higher rate of prenatal loss (22 per cent) as well as the presence of intrauterine growth retardation (IUGR) has been reported among pregnancies with CPM. To evaluate more precisely the effect of these aneuploid cell lines confined to the placenta on intrauterine fetal growth and fetal survival, we have studied 34 term placentae from pregnancies with CPM diagnosed on CVS and confirmed identical mosaicism in 17 of these placentae. There was a direct correlation between a high number of aneuploid cells present at CVS and a high likelihood of their detection in term placenta. Also, the proportion of aneuploid cells in the mosaic term placentae correlated with that observed in CVS specimens. Among 17 gestations with confirmed CPM at delivery, there were six cases of IUGR identified, five in liveborns and one associated with intrauterine death.     

Prenatal diagnosis of congenital cytomegalovirus infection in 189 pregnancies with known outcome

Prenatal diagnosis (PD) of fetal cytomegalovirus (CMV) infection was performed in 242 pregnancies, with known outcome in 189 cases. In 141/189 pregnancies, PD was carried out on account of suspicious maternal CMV serology up to gestational week (WG) 23, and in 48 cases on account of abnormal ultrasonic findings detected between WG 18 and 39. Chorionic villus samples (n = 6), amniotic fluid (AF, n = 176) and/or fetal blood specimens (n = 80) were investigated for detection of virus by cell culture, shell vial assay, PCR and/or CMV-specific IgM antibodies. Of 189 fetuses correctly evaluated by CMV detection either in fetal tissue following therapeutic abortion/stillbirth (n = 24) or in urine of neonates within the first 2 weeks of life (n = 33), 57 were congenitally infected. In women with proven or suspected primary infection, the intrauterine transmission rates were 20.6% (7/34) and 24.4% (10/41), respectively. Of the congenitally infected live-born infants, 57.6% (19/33) had symptoms of varying degree. The overall sensitivity of PD in the serologic and ultrasound risk groups was 89.5% (51/57). A sensitivity of 100% was achieved by combining detection of CMV-DNA and CMV-specific IgM in fetal blood or by combined testing of AF and fetal blood for CMV-DNA or IgM antibodies. There was no instance of intrauterine death following the invasive procedure. The predictive value of PD for fetal infection was 95.7% (132/138) for negative results and 100% (51/51) for positive results. Correct results for congenital CMV infection by testing AF samples can be expected with samples obtained after WG 21 and after a time interval of at least 6 weeks between first diagnosis of maternal infection and PD. In case of negative findings in AF or fetal blood and the absence of ultrasound abnormalities at WG 22-23, fetal infection and neonatal disease could be excluded with high confidence. Positive findings for CMV infection in AF and/or fetal blood in combination with CMV suspicious ultrasound abnormalities predicted a high risk of cytomegalic inclusion disease (CID). Furthermore, detection of specific IgM antibodies in fetal blood was significantly correlated with severe outcome for the fetus or the newborn (p = 0.0224). However, normal ultrasound of infected fetuses at WG 22-23 can neither completely exclude an abnormal ultrasound at a later WG and the birth of a severely damaged child nor the birth of neonates which are afflicted by single manifestations at birth or later and of the kind which are not detectable by currently available ultrasonographic techniques.     

Transvaginal sonography-detection of findings suggestive of fetal chromosomal anomalies in the first and early second trimesters.

Over a 4-year period, 14 dyskaryotic fetuses were diagnosed by amniocentesis, performed after early detection of malformations using transvaginal sonography (TVS). These 14 dyskaryotic fetuses were detected out of 4878 sonographic screenings performed by TVS between 9 and 16 weeks' gestation. Twenty-eight per cent of the referrals were at high risk and 72 per cent were at low risk for fetal malformations. Two hundred and twenty-nine fetuses (4.7 per cent) of the screened population had 265 anomalies, 39 per cent of them being transient. In 7 of the 14 dyskaryotic fetuses (50 per cent), the sonographically detected anomalies were transient, being undetected by follow-up sonographic screenings at later gestational ages (greater than or equal to 18 weeks). Postponing the first sonographic scan aimed at malformation detection to a later gestational age may lead to transient anomalies and their associated dyskaryosis being missed.