Correlates of placental infection with cytomegalovirus, parvovirus B19 or human herpes virus 7.

Vertical transmission of viruses is an important cause of morbidity in the fetus and neonate. Placental viral infection indicates risk of vertical transmission, but not always transmission to, or disease of the fetus. Specimens from mothers and babies from three groups-two prospective and one retrospective cohort-were tested for pathogens of teratogenic potential using multiplex PCR. Placental infection was present in 13% of the 105 samples collected. Assessment of the prospective cohorts showed cytomegalovirus (CMV) detected in 4% of placentae from unselected women, parvovirus B19 in 1% and Ureaplasma parvum in 1% of placentae. In a retrospective cohort of women at high risk of transmitting congenital infection due to seroconversion during pregnancy, miscarriage or stillbirth, CMV was detected in 64% and human herpes virus type 7 in 9% of placentae. Of 14 PCR-positive placentae, two were associated with the birth of a living symptomatic infant, two with stillbirth, one with miscarriage, and two with elective terminations of pregnancy. Directed laboratory assessment of women at high risk of transmitting congenital infection, on the basis of clinical or laboratory markers, is important for accurate diagnosis of adverse outcomes of pregnancy. However, routine screening for viruses in the placentae from women with a low-risk serological profile for transmitting congenital infection is unlikely to result in significant numbers of additional diagnoses and is confounded by inadequacy of current diagnostic methods. The major pathogen detected in all cases of placental infection associated with fetal death was human CMV.

Development of multiplex PCRs for detection of common viral pathogens and agents of congenital infections.

Potential causes of congenital infection include Toxoplasma gondii and viruses such as cytomegalovirus (CMV), enterovirus, hepatitis C virus, herpes simplex virus types 1 and 2 (HSV-1 and -2), human herpesvirus types 6, 7, and 8, lymphocytic choriomeningitis virus, parvovirus, rubella virus, and varicella-zoster virus. Testing for each of these agents using nucleic acid tests is time consuming and the availability of clinical samples such as amniotic fluid or neonatal blood is often limited. The aim of this study was to develop multiplex PCRs (mPCRs) for detection of DNA and RNA agents in the investigation of congenital infection and an mPCR for the viruses most commonly requested in a diagnostic virology laboratory (CMV, Epstein-Barr virus, enterovirus, HSV-1, HSV-2, and varicella-zoster virus). The assays were assessed using known pathogen-positive tissues (cultures, placentae, plasma, and amniotic fluid) and limits of detection were determined for all the agents studied using serial dilutions of plasmid targets. Nested PCR was performed as the most sensitive assay currently available, and detection of the amplicons using hybridization to labeled probes and enzyme-linked immunosorbent assay detection was incorporated into three of the four assays. This allowed detection of 10 to 10(2) copies of each agent in the samples processed. In several patients, an unexpected infection was diagnosed, including a case of encephalitis where HSV was the initial clinical suspicion but CMV was detected. In the majority of these cases the alternative agent could be confirmed using reference culture, serology, or fluorescence methods and was of relevance to clinical care of the patient. The methods described here provide useful techniques for diagnosing congenital infections and a paradigm for assessment of new multiplex PCRs for use in the diagnostic laboratory.

Diagnosis of and screening for cytomegalovirus infection in pregnant women.

No single diagnostic test for cytomegalovirus (CMV) infection is currently available for pregnant women at all stages of gestation. Improved accuracy in estimating the timing of primary infections can be used to identify women at higher risk of giving birth to congenitally infected infants. A diagnostic algorithm utilizing immunoglobulin G (IgG), IgM, and IgG avidity was used to prospectively screen serum from 600 pregnant women enrolled from two groups: < or =20 weeks gestation (n = 396) or >20 weeks gestation (n = 204). PCR testing of urine and/or blood was performed on all seropositive women (n = 341). The majority (56.8%) of women were CMV IgG seropositive, with 5.5% being also CMV IgM positive. In the IgM-positive women, 1.2% had a low-avidity IgG, indicating a primary CMV infection and a high risk of intrauterine transmission. Two infants with asymptomatic CMV infection were born of mothers who had seroconverted in the second trimester of pregnancy. Baseline, age-stratified CMV serostatus was established from 1,018 blood donors. Baseline seropositivity from a blood donor population increased with age from 34.9% seroprevalence at less than 20 years of age to 72% seroprevalence at 50 years of age. Women at high risk of intrauterine transmission of CMV were identified at all stages of gestation. Women infected with CMV during late gestation may be more likely to transmit the virus, so failure to detect seroconversions in late gestation may result in failure to detect infected neonates.