BACTrIOLOGiCAL REVIEWS, Mar. 1973, P. 19-31 Vol. 37, No. 1Copyright i 1973 American Society for Microbiology Printed in U.S.A.

Viral TeratologyDAVID A. FUCCILLO AND JOHN L. SEVER

Infectious Diseases Branch, National Institute of Neurological Diseases and Stroke,Bethesda, Maryland 20014

INTRODUCTION .............................................................. 19MECHANISMS OF THE FETAL DAMAGE .......... .......................... 20VIRUSES TERATOGENIC IN MAN ............. .............................. 21Rubella................................................................ 21Cytomegalovirus ............................................................... 21Herpes Simplex Virus......................................................... 22

VIRUS IMPLICATED IN CONGENITAL INFECTIONS AND INFECTIONSIN NEWBORN INFANTS ................. ............................... 22

Mumps................................................................ 22Echoviruses ............................................................... 23Rubeola Virus ............................................................... 23Vaccinia Virus ............................................................... 23Varicella-Zoster Virus......................................................... 23Western Equine Encephalitis Virus ............. ............................... 23Variola Virus ............................................................... 24Inlfluenza Virus ............................................................... 24Hepatitis................................................................ 24Coxsackieviruses .............................................................. 24

VIRUSES TERATOGENIC IN ANIMALS .......... ............................ 25Blue Tongue of Sheep......................................................... 25Hog Cholera Virus ............................................................. 25Parvoviruses ............................................................... 26Reoviruses ............................................................ ... 26

DIAGNOSIS ............................................................ ... 26CONCLUDING STATEMENT .................................................. 28LITERATURE CITED .......................................................... 28

INTRODUCTION

Congenital malformations account for ap-proximately 14% of all deaths within the firstyear of life in the United States (19). In a recentstudy of 45,000 pregnancies from the Col-laborative Perinatal Research Study sponsoredby the National Institute of Neurological Dis-eases and Stroke, a rate of 414 malformationsper 10,000 births (4.1%) was found (8).Very little data are available to estimate the

relative importance of viruses as causes ofmalformations. Certainly, several viruses havebeen well established as causes of severe de-formities in man. The role of viruses in produc-ing the more subtle fetal defects is just beinginvestigated. For example, the frequency of lossof hearing as the only manifestation of therubella syndrome was recently documented bylong-term studies of patients with serologicallyand virologically documented infections (91).A very significant gap in our knowledge is the

role of perinatal viral infections in mentalretardation. Of the 3.5 million children borneach year in the U.S., approximately 3% are

mentally retarded (82). It appears that asmany as 10% of these cases may be attributedto infectious diseases (23). Specific studies ofthese preventable causes of mental retardationare needed to define this group precisely and toinstitute appropriate measures for treatment orprevention.Information available on viral teratogens is

still limited, but is continuously expanding asknowledge is acquired about the fetal responseto infection as well as viral biology and immu-nology. Control and prevention of viral infec-tions having teratogenic potential require: (i)recognition of the causal relationship betweenthe infection and the affected fetus or child; (ii)understanding the mechanisms responsible forthe infection and damage; (iii) knowledge ofthe biological characteristics of the virusesinvolved and their mechanism of transmission(88). Clinical and subclinical viral infections ofthe mother may lead to the invasion of thefetus. Thus, the identification of viruses whichhave teratogenic potential is often difficult,since many of the subclinical infections gounreported. Adequate data for both clinical

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and subclinical infections can be obtained onlyby use of prospective studies employing bothvirus isolation and serological studies. The useof new animal model systems now being devel-oped should also help in the understanding ofviral effects on the fetus (90).The frequency of clinically recognized infec-

tions during pregnancy has been determined inthe Collaborative Perinatal Research Study(92). Although the figures are at best anapproximation, they do provide an idea of therelative frequency of infection in the U.S. Thestudy population included 30,000 pregnanciesregistered between October 1958 and February1964. In this study population, approximately5% of the patients reported definite or pre-sumed viral infections. Excluding the commoncold, the most frequent infections were influ-enza, flulike illnesses, herpesvirus infections,viral gastroenteritis, and viral tonsillitis. Thespecific infections of mumps, rubella, chickenpox, viral bronchitis, measles, viral conjunLti-vitis, viral pneumonitis, and infectious mono-nucleosis occurred at rates between 1 and 15per 10,000 pregnancies. Serological tests doneon serial blood specimens substantiated re-ported infections in about two-thirds of 45 ru-bella cases, one-fifth of 11 rubeola cases, andalmost three-fourths of 46 mumps and vari-cella zoster cases. The data from clinicalrecognition alone frequently incorrectly esti-mated true frequency of infection, as indicatedby the serological studies. Combined clinicalreporting and serological information per-mitted the development of minimum estimatesof frequency: thus, for 10,000 pregnanciesunder nonepidemic conditions, the minimumfrequency of maternal infections was: mumps,10; rubella, 8; varicella zoster, 5; and rubeola,0.6. During epidemic years, the frequency ofclinical rubella may have reached 220 per10,000. However, since about one-third of theadult infections are subclinical, the total epi-demic frequency might have been as high as330 per 10,000.

Viral infections which are known to producemalformations in the human embryo includerubella, cytomegalovirus (CMV), and herpessimplex virus (HSV). A number of other virusesare known to be associated with an increasedincidence of illness or fetal death. Others maybe implicated but, at present, data are insuffi-cient to establish a clear causal relationshipbetween the infection and the effects in thechild. The present report summarizes some ofthe information now available for these agentsand their effects on the developing fetus.

MECHANISMS OF THE FETALDAMAGE

Some viruses can infect the fetus at any timeduring gestation, and the frequency and type ofdefects produced often depend upon the time ofmaternal infection. If morphogenesis is incom-plete at time of virus infection, pregnancy mayterminate in abortion, malformations, degener-ative lesions of the fetus or stillbirth. If infec-tion takes place after the fetus has completelydeveloped, results may be abortion, stillbirth,prematurity, or pathogenic lesions. Malforma-tions (62) such as hydrocephalus due to steno-sis of the aqueduct of sylvius may be restrictedto infection at a late developmental stage.Diseases associated with congenital infec-

tion can be divided into two basic types: oneresembling the disease seen in the adult andthe other which is unique and characteristi-cally different from that of the adult. In uteroinfections resembling the adult disease includepolio, Western equine encephalitis, and small-pox. The presence or absence of receptor sitesmay account for this susceptibility or resist-ance of tissues to certain viruses (54). Theunique fetal type of diseases caused by viruseswhich usually have limited pathogenicity inadults are notably rubella, CMV, and dis-seminated infections due to herpesvirus andchickenpox.A number of factors are thought to be respon-

sible for the increased susceptibility of thefetus to certain viruses. First, all of theseviruses pass through the placenta and replicatein the fetus. The lack of antibody productionuntil about 20 weeks of gestation and theinsignificant amounts of circulating leukocytesuntil about the 12th week of gestation probablycontribute to the dissemination of virusthroughout the fetus. Rapidly dividing cells,such as those found in a developing fetus, areknown to be more favorable for the cultivationof viruses. Also, these fetal tissues produce lessinterferon than adult tissues.Many possible mechanisms of fetal damage

have been proposed for the teratogenic capabil-ities of viruses. Only limited data are availableto support any hypothesis. One of the mech-anisms involved with fetal damage is cellularnecrosis that is caused by the action of virusesin fetal tissues. Lytic viruses are associatedmost often with this mechanism, however,nonlytic viruses are not excluded. Herpesvirus,varicella, vaccinia, variola, and coxsackievi-ruses are examples of highly lytic viruses whichcan produce severe intrauterine or neonatal

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infection, with or without malformation anddeath (40). Viruses such as rubella and CMV,which produce lesser degrees of cellular necro-sis and cell lysis, more characteristically pro-duce widespread malformation with variabledegrees of tissue damage. These effects arethought to result from the interference ofgrowth or maturation of tissue with persistenceof infection (56). The limited replicative capac-ity of infected cells is probably the basis fordecreased growth and specific malformations(81).Another possible mechanism of fetal dam-

age, although highly speculative, is that thevirus may produce chromosomal injury andthereby induce congenital malformation orabortion (101). Rubella virus grown in vitro hasbeen shown to inhibit mitosis and increase thenumber of chromosomal breaks (78). Herpes-virus and rubeola have also been shown toproduce abnormal mitosis (72). To date, how-ever, no such changes have been found in vivo.

Generalized vasculitis of the placenta andthe fetus is another theoretical mechanismwhich may be associated with congenital mal-formations (62). In cases of congenital rubella,vasculitis of cerebral arteries and stenosis oflarge blood vessels have been reported (103).Vascular lesions of the cerebral circulation maylead to the production of small foci of calcifica-tion. This same mechanism may be involved incongenital cytomegalic disease, as well as inherpesvirus type II malformations (98). In-travascular coagulation may be manifested byclinical signs such as thrombocytopenia andpurpura (38). This mechanism could accountfor the similarities of the clinical manifesta-tions of rubella, CMV, and herpesvirus.Thrombosis and vasculitis of the small bloodvessels could produce organ hypoxia, and thusreduce the rate of cellular division. This sameeffect could take place with placental infectionand produce indirect damage without fetalinvolvement (62).

VIRUSES TERATOGENIC IN MAN

RubellaGregg, in 1941, initially reported the associa-

tion between maternal rubella and congenitalmalformations (30). Originally, it was thoughtthat all fetal defects were produced duringinfection in the first trimester. It is now clearthat fetal infection following rubella in thesecond trimester can produce less obvious, butnevertheless severe, damage of the fetus (58,61). The major damage to the fetus include

heart lesions, cataracts, deafness, microen-cephaly, and mental retardation (64, 83). Ap-proximately 50% of mothers infected in thefirst month of pregnancy have abnormal chil-dren, 22% in the second month, and 6 to 8% inthe third through the fifth months. Defectsassociated with rubella in the second trimesterinvolve problems of communication based onhearing loss, mental-motor retardation, andeye defects. Children with congenital rubellafrequently remain infected for 6 months orlonger after birth. Virus may be present in thenaso-pharynx, lymphocytes, or spinal fluid. By1 year of age, only about 10% are still infected.However, in one reported case, the virus wasisolated from a cataract of a 3-year-old child(60). Most children shed virus while they havehigh circulating antibody titers, indicating apossible defect in the cellular immune mecha-nism.

Live vaccines produced in duck and rabbittissue culture are now being administered tomillions of children to reduce the susceptiblepool and thereby reduce the exposure of preg-nant women to the wild virus. Only limiteddata are available on the possible transmissionof the vaccine strain of the virus to the fetus.Therefore, it is imperative that pregnantwomen do not receive the vaccines.

CytomegalovirusThe major manifestations of congenital cy-

tomegalic inclusion disease (CID) include mi-crocephaly or hydrocephaly, chorioretinitis,blindness, hepatosplenomegaly, encephalitiswith cerebral calcifications, and mental retar-dation. Occasionally, these children havepneumonia, anemia, thrombocytopenia withpetechiae at birth, and rarely congenital heartdisease (33).One study has reported that congenital CID

infection accounts for 1 or 2% of all infantdeaths (70). A high percentage of the infantswho survive often have neurological sequelaethat include blindness and mental retardation(34). In one study, approximately 10% of mi-crocephalic mentally retarded children hadserological evidence of CID infection (100).Further investigations are needed to definemore precisely the relative importance of CIDin the production of mental retardation.During pregnancy, approximately 3.5% of

pregnant women develop significant levels ofantibody to CMV (89). Other studies havedemonstrated that about 3% of women excretevirus in the urine during pregnancy, and that

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about 1% of the newborn infants excrete cy-tomegalovirus in the urine at birth (10, 39).

In a recent study, a total of 545 urinespecimens were obtained from newborns withinthe first 24-h of life and cultured for thepresence of cytomegalovirus. Three childrenhad viruria which persisted throughout a pe-riod of 1 year of observation (10). None of thechildren had the classical findings of CID,although minimal abnormalities such as tran-sient petechiae and hepatosplenomegaly werenoted. After 1 year of age, two were entirelynormal and one had findings of mild spasticity.It is quite evident, therefore, from these andother studies, that congenital infection withCMV is often not fatal and perhaps frequentlysubclinical. Large-scale longitudinal studies ofinfected infants are needed to define preciselythe effect of perinatal CMV infection.As with rubella infection, CMV persists in

the infants for months or years in the presenceof neutralizing antibody. Lymphocyte culturesfrom children with CID have demonstrated thepresence of virus for at least several months(55).At present no specific therapy is available.

Drugs such as cytosine arabinoside have beenused for treatment of children, with little effecton excretion of virus (53).

Herpes Simplex VirusRecent studies have clearly indicated that

two strains of HSV exist. Type 1 infects the oralarea and type II infects the genital area (66).Type I usually causes the common cold sore orfever blister, whereas type II produces cervicitisand vaginitis and is thought to be venereallytransmitted. These two strains differ consider-ably in their antigenic and biological proper-ties. Approximately 95% of herpes infections ofthe genital area are due to type II. In a series of28 cases of neonatal herpetic infection, 25 werefound to be due to type II herpesvirus (67).

Congenital infection with HSV may producea syndrome quite similar to that found withcongenital CMV infection. Newborns may havemicrocephaly, cerebral calcification, chorioret-initis, and mental retardation (98). If trans-placental or transamnionic infection occurnear delivery, a spectrum of clinical disease inthe newborn may range from a few cutaneousvesicles to a fatal generalized disease withjaundice, fever, hepatosplenomegaly, encepha-litis, circulatory coagulation, and death (63,96).Neonatal HSV infection can occur as a

fulminating or as a slow, progressive disease

with delayed death or recovery with or withoutsequelae.

Present evidence suggests that transplacen-tally acquired type I does not protect infantsfrom infection with type II virus (67). In theCollaborative Study, three cases of congenitalHSV infection were found. Serological studiesshow that, early in pregnancy, all had antibodyagainst type I and that seroconversion to typeII occurred at the time of delivery. Thus thefetus was not protected by transplacental typeI antibody.Treatment of HSV infection in the newborn

is still not possible. Drugs such as iododeox-yuridine (IUDR) has been tried without muchsuccess (107). Studies with other durgs are nowin progress. The greatest risk to the fetusappears to occur when the mother developsgenital herpes near the time of delivery. Cae-sarean section has not been thoroughly stud-ied but may be a method of avoiding newborninfection when lesions are found in the mother.

VIRUSES IMPLICATED INCONGENITAL INFECTIONS AND

INFECTIONS IN NEWBORN INFANTS

MumpsIn a prospective study, maternal mumps

infection during the first trimester was as-sociated with increased fetal mortality. Sixtypercent of the abortions occurred within 2weeks of onset of maternal disease. However,fetal infection was not demonstrated by viralisolation (97).Endocardial fibroelastosis in infants may be

associated with congenital mumps in somecases (94). This association was based on datashowing that infants with this disease have ahigh incidence of positive skin test to mumpsantigen. Other workers, however, have not beenable to confirm this association nor demon-strate hemagglutinating antibodies to mumpsvirus in the sera of patients with this disease(29).

In a recent animal study, pregnant Rhesusmonkeys were inoculated with mumps virusduring the first third of gestation. Replicationof virus was detected in the fetal or placentaltissues for only 1 week. Full-term infant mon-keys demonstrated delayed hypersensitivity tomumps virus but no antibody (99). This splitimmunological response lends some support toresults also found in humans (1). However,there was no report of endocardial fibroelasto-sis in the infant monkeys.

In the Collaborative Perinatal Research

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Study, there were 10 cases of endocardialfibroelastosis, and in one there was evidence ofmaternal mumps.

Animal studies have been reported in whichhydrocephalus was produced in suckling ham-sters inoculated intracerebrally with a strain ofmumps virus (42). Hydrocephalus was due toaqueductal stenosis which occurred as a latesequela several weeks after the acute infection.This was the first demonstration that mumpsvirus could cause a definite central nervoussystem anomaly. Since that report, there havebeen two cases of hydrocephalus noted as a latesequela occurring 27 and 20 months after acuteinfection in 5- and 8.5-year-old children (12,102). Whether mumps is indeed involved inthis neurological syndrome will depend upon

further isolation and serological studies.Other animal models, preferably primates,

must also be developed to determine whethermumps virus is involved in utero in the produc-tion of hydrocephalic newborn infants.An attenuated live mumps vaccine which

appears to produce effective levels of mumpsantibody titers has been developed.

EchovirusesVery little information is available on the

echo viruses and their capability of producingcongenital infections. These viruses have beenisolated from asymptomatic infants and fromnewborn cases of aseptic meningitis, diarrhea,febrile illnesses, and rashes (84). The echo-viruses appear to have the capacity of infectingthe placenta and also transmitting the infec-tion to the fetus (9). A recent report describesthree infants congenitally infected withEcho-19 (77). Pathological findings closely re-

semble those found with HSV which is a

generalized infection with massive terminalhemorrhage involving most organs of the body.Echo virus 14 was reported to be isolated

from the liver of a newborn infant with fatalhepatic necrosis (41). High immunoglobulin Mtiters were present at 6 days of age, indicatingthat possible infection was acquired in utero.

Rubeola Virus

Only limited information is available con-

cerning rubeola and its role in perinatal infec-tions. Since the advent of a live rubeola vac-cine, information on this virus and its relation-ship to congenital infection in the U.S. ispractically nonexistent. However, congenitalinfection has been reported to be involved witha high incidence of stillbirths and abortions(24, 76). Fetal infections occurring before term

may produce newborns with the clinical diseaseor development of disease soon after birth.Giant-cell pneumonia has also been reported innewborns with this disease (22). It has alsobeen reported that mothers who had measlesduring the first trimester have increased ratesof premature infants when compared to normalrates.At the present time, there are two live

measles vaccines licensed for use in the U.S.There is some evidence of reinfection of personswith low levels of antibody.

Vaccinia VirusCongenital infection with vaccinia virus

rarely occurs as a result of reimmunization ofpregnant women (24). Primary maternal vacci-nation in the first or second trimesters maylead to fetal infection and possibly stillbirth,abortion, or prematurity and death (40). Fetalinfection develops as a disseminated-type dis-ease with focal lesions occurring on the skin,lungs, liver, and other organs of the body.Lesions may be confused with those producedby herpes or varicella virus. Complicationsassociated with vaccination are eczema vac-cinatum, vaccinia necrosium, and postvac-cinial encephalitis (65). Infants or adults withexfoliative dermatitis, especially eczema,should not be vaccinated nor exposed to re-cently vaccinated individuals. The low inci-dence of smallpox in the U.S. plus the com-plications associated with vaccination has in-fluenced the Public Health Service to recom-mend discontinuing of routine smallpox vacci-nations in the U.S. as of September 1971.

Varicella-Zoster VirusVaricella (chickenpox) and zoster (shingles)

lesions are caused by the same virus. Congeni-tal infection associated with maternal chicken-pox or herpes zoster infection has been reported(2, 16). Infection early in gestation has led toprematurity, stillbirth, or abortion. Congenitalinfection acquired late in pregnancy is oftenmanifested at birth or in the newborn period bythe presence of the characteristic chickenpoxskin lesions or severe pneumonia (3). Zosterlesions have also been observed with theircharacteristic segmental distribution. This dis-ease is usually mild, although it may exist insevere disseminated forms which end in deathof the infants. No vaccine is available for thisdisease.

Western Equine Encephalitis VirusTransplacental passage of Western equine

encephalitis virus has been demonstrated with

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serological evidence by a number of investiga-tors (21, 93). Congenital encephalitis has beenfound in the newborn infant at time of orshortly after delivery. No congenital abnor-malities have been described.

Variola Virus (Smallpox)The effect of variola virus on the fetus has

not been considered a problem in the U.S.,since it is almost completely absent from thiscountry. It also has a very low incidence inmost of the rest of the world. This virus can betransmitted to the fetus as a result of maternalinfection (13, 25). The results, similar to thosepreviously described for vaccinia virus, aredeath, abortion, or congenital disease with skinlesions and pneumonia in the neonatal infant.No congenital malformations have been as-sociated with this virus.

Influenza VirusFor many years, influenza virus has been

regarded as a potential teratogenic agent forthe human fetus. However, there are probablyjust as many publications showing correlationbetween the disease and congenital malforma-tions (20, 32, 36, 85, 109, 110) as there arenegative reports denying this effect (18, 108).The reasons for these contradictory reports canbe explained in part by the methods employedfor collecting data. Many studies have de-pended upon the self-diagnosis of the diseaseand self-reporting to correlate infection withpregnancy. Often there is little or no serologicalbackup for the histories. This method can bevery inaccurate and misleading. Other studieshave depended upon the evaluation of clinicalsymptoms and signs. With this type of ap-proach, the inapparent or subclinical infectionsare overlooked. Most investigations have beenmade with very small numbers of patients andcontrols.Some of the positive findings reported for

influenza infections during pregnancy includeincreased incidence of fetal death, anenceph-aly, abortions, stillbirths, prematurity, andcongenital malformations. Most of the dataavailable on these associations are at a very lowlevel of statistical significance.

Experimental animal studies tend to supportthe conclusion that maternal infection doescause fetal abnormalities (62). Abnormalitiesof the nervous system and fetal wastage havebeen demonstrated in mice infected with influ-enza virus early in pregnancy. Malformationsin chicken embryos have been induced withinfluenza A virus. Abnormalities similar to

those occasionally found in human fetuses wereproduced in chicken embryos, e.g., abnormaltube flexure and failure of neural tube closure.Viral infection was limited to extranueral tis-sues suggesting an indirect effect on or-ganogenesis (43). Hydrocephalus has been re-ported by others as a result of influenza virusbeing inoculated intracerebrally into sucklinghamsters and mice (45).More extensive data with laboratory support

will be required to establish the role of influ-enza on the course of pregnancies.

Hepatitis

Two types of hepatitis designated as A and Bhave been described. Type A produces infec-tious hepatitis of short incubation, and type Bproduces serum hepatitis of long incubation.The hepatitis virus responsible for these twodiseases has not been isolated. A hepatitis-associated antigen shows a strong relation-ship to serum hepatitis (11). This antigen or itsantibody may be found in serum from patientswith this disease. The availability of serologi-cal tests is important in the diagnosis and inepidemiological studies of this disease (80).

Hepatitis-associated antigen can cross theplacenta and appear in the cord blood (46, 72).This type B hepatitis may also be transmittedby close contact from mother to infant afterbirth in the neonatal period (79).The availability of a serological test now

helps to differentiate this hepatitis from otherclinical hepatitis (produced by other virusessuch as rubella, coxsackie, CMV, herpes, andvaricella) in the newborn infant.

Congenital hepatitis may result in stillbirthor abortion of the fetus. Newborn infants canappear with jaundice, hepatosplenomegaly, he-molytic anemia with a liver function test in-dicating an obstructive type of jaundice. About80% of the infants survive for at least 1 year;50% of these exhibit a chronic persistent liverdisease. There have been examples of mothersrepeatedly giving birth in successive pregnan-cies to infants with hepatitis, thus indicating achronic maternal infection (5).Experimental animal model systems have

been found but do not demonstrate the his-topathological changes found in the humandisease. Apparent viral propagation with anti-body formation has been shown (57).

Coxsackieviruses

There are more than 30 serological types ofcoxsackieviruses which are divided into two

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groups, A and B. Group B has been associatedwith congenital and neonatal infection. Theseviruses have been reported to have been iso-lated from stillbirths and aborted fetuses. Inone study of 50 stillborn infants and neonataldeaths, coxsackie B antigen was present in themyocardium of five (17). Myocarditis is theclinical finding most frequently associated withcoxsackievirus infection (14). However, dis-seminated infection involving multiple organsystems has been reported. In a recent prospec-tive study of 630 mothers of anomalous infantsand controls, maternal seroconversion to cox-sackieviruses appeared to be increased duringpregnancy with a variety of congenital defects(15). Coxsackie B2 and B4 were associated withurogenital anomalies, and type A9 with defectsof the digestive system. Cardiovascular anom-alies were associated with coxsackie B, andB4 infection during pregnancy, and the likeli-hood of congenital heart disease was increasedby maternal infection with two or more cox-sackie B viruses rather than with one. Anoma-lous infants weighed less than their controlsand were more often male. Further studies arein progress.Maternal or postnatally acquired infection

may be clinical or subclinical. Clinical manifes-tations can be mild or severe and includeupper respiratory tract infection, pneumonitis,pleurodynia, myocarditis, pericarditis, andaseptic meningitis. Diagnosis therefore israther difficult, and further confirmatory stud-ies should be done with both virological andserological support.Animal model systems have not yet been

developed to help confirm the relationship ofcoxsackieviruses to in utero infection.

VIRUSES TERATOGENIC IN ANIMALS

Blue Tongue of SheepBlue tongue is an animal disease produced

by a small RNA-containing virus which pri-marily affects sheep. The disease is transmit-ted naturally by an inset vector, and thereforethe virus was originally grouped with the ar-boviruses. Blue tongue virus is now provision-ally classified as a reovirus. The name of thisdisease is derived from the clinical pictureobserved in adult animals. There is hyperemiawith cyanosis and edema of the tongue, mouth,eyes, and nose. Mortality usually due to pneu-monia is 1 to 5%.Malformations and other fetal damage were

noted when pregnant ewes were immunizedwith an attenuated chicken embryo-grown vac-cine. Vaccination of pregnant ewes during 5 to

7 weeks' gestation produced abnormalities in20 to 50% of the fetuses (95). Stillbirths andneonatal deaths increased, and extensive de-fects such as deafness, blindness, shortenedlimbs, and deformities of the spine and neckwere observed. Diffuse anomalies of the centralnervous system were a common finding withfetal infection (31).

Additional studies demonstrated that thenature of malformations were dependent uponthe gestational age when infection occurred.Direct intramuscular inoculation of the fetus inthe first half of the 150-day gestation periodproduced consistent anomalies. Inoculation at50 days of gestation led to hydranencephaly, at75 days caused porencephaly, and at 100 daysproduced no gross malformations (74, 75). Thisincreased susceptibility of immature cells wasfurther demonstrated by studies with bluetongue virus vaccine adapted to newborn mice(68). However, it was shown to be a veryselective infection of the germinal zones andnot soley dependent on immaturity or mitoticactivity.Attempts to modify blue tongue virus-

induced malformations with cyclophospha-mide and antithymocyte serum did not causemice to be more susceptible to infection (69).An interesting observation is that the fetus

can be transplacentally infected even in thepresence of maternal immunity. This wouldindicate some intracellular transport mecha-nism of the virus to the fetus, possibly bycirculating lymphocytes or another cellularcomponent.

Hog Cholera VirusHog cholera virus is an RNA-containing

virus resembling the myxovirus and is classi-fied provisionally as a Toga virus. Initial stud-ies demonstrated that fetal abnormalitiescould be produced by an attenuated rabbit-modified hog cholera virus injected into preg-nant dams 10 to 16 days after breeding. Thesestudies showed that 38% of the newborn pigshad edema, ascites, and deformities of the ears,nose, legs, and kidneys (111). Additional stud-ies confirmed these observations, but also dem-onstrated anomalies of the nervous systemwhen dams were injected at 20 to 90 days ofgestation. Cerebral hypoplasia and hypomye-linogenesis with congenital tremors were pro-duced in almost 50% of the newborns (26).Newborn pigs infected in utero also harbored apersistent virus in their viscera, making itpossible to transmit infection to other membersof the herd.Recent studies indicate that malformations

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are caused by a persistent, immunologicallytolerant, and noncytolytic infection of the fe-tus. The mechanisms of malformation appearsto be a selective inhibition of cell division andfunctions (44).

ParvovirusesThree viruses, H-1, rat virus, and feline

panleukopenia virus, belong to the group at onetime called picodna but now termed par-voviruses. They are all singe-stranded DNAviruses characterized by their propensity toattack replicating cells, destructive actionupon the developing cerebellum, and ability tocross transplacental barriers. H-1 was report-edly isolated from normal human fetuses andplacentas (105). It has many of the characteris-tics of the rat virus (RV), and the two are con-sidered closely related (106). When injectedinto pregnant rats, RV produces congenitalinfection which results in fetal death, reab-sorption, fetal survival with hepatitis, andcerebellar hypoplasia (47, 50). H-1 virus pro-duces similar effects in pregnant hamsters(104). A mongoloid-like deformity is producedin these animals which superficially resemblespatients with Down's syndrome, but with theabsence of chromosomal abnormalities (6).The relationship of H-1 virus or RV to hu-

man disease is still unknown. As previouslynoted, isolation of this virus has been reportedfrom human placentas and embryos. Serologi-cal studies in man, however, have failed toshow any relationship between maternal anti-body to H-1 or RV and pregnancies whichterminate in stillbirth, abortion, mongolism, orcongenital defects (71).

Feline panleukopenia virus is serologicallydistinct from H-1 and RV. For many years thecerebellar hypoplasia of kittens now known tob6-caused by feline panleukopenia virus wasregarded as a common genetic disorder ofcats (48). The virus selectively attacks theexternal germinal layer of the cerebellum dur-ing the active phase of histogenesis, result-ing in a congenital cerebellar hypoplasia in catand ferret fetuses (49).The degree of pathogenetic damage induced

by virus in the cerebellums of kittens eitherinoculated directly in utero or by intravenousinoculation of a pregnant cat was proportionalto the duration of their infections (52). Viruscan be isolated from the kidneys of infectedkittens for many weeks or months after birth,even in the presence of passively acquiredantibodies. The possibility that this virus orothers like it being involved in production of

cerebellar hypoplasia in man has not yet beenthoroughly explored.

ReovirusesReoviruses are ubiquitous in nature, and

their role in the etiology of human disease hasnot been well defined (27). Spontaneous dis-ease caused by reovirus has commonly beenobserved in laboratory colonies, primarily insuckling mice. Congenital and neonatal infec-tion in animals have been reported (37). Preg-nant mice were inoculated with type 2 virus on1, 3, 6, 10, and 15 days of gestation. A pro-longed infection of the fetus was produced.About 25% of the newborn mice developedinterstitial pneumonia and subcortical renaltubular necrosis. Virus can be isolated from allorgans in high titer at 15 to 36 days of life.Another 50% show growth retardation andconjunctivitis. Developmental anomalies werenot observed.

Reovirus type 1, when inoculated into suck-ling hamsters, ferrets, rats, and mice via cere-bral or intraperitoneal routes, regularly causeobstructive hydrocephalus (51). The virus se-lectively attacks the ependymal and choroidplexus epithelium within 4 to 6 days afterinoculation (59). Nearly 100% of the hamstersdeveloped an ependymitis followed by grosshydrocephalus. Newborn ferrets respond simi-larly by tolerate infection longer before devel-oping hydrocephalus. Rats and mice also de-velop hydrocephalus but have widespread sys-temic disease causing runting with gastrointes-tinal tract illness.

DIAGNOSISRetrospective and prospective investigations

have been employed to obtain data on thefrequency of various virus infections duringpregnancy and to estimate the teratogenicpotential of many agents. The restrospectiveanalysis depends upon the recognition of anabnormality in the newborn and then attemptsto associate this with a disease which occurredduring the mother's pregnancy. The main lim-iting factors are the inability to document theinfection and failure to recognize inapparentinfections. The prospective approach providesfor the observation of the mother throughouther pregnancy. This approach is usually themost preferred, even though it is more difficultand time consuming. Clinical infections can bedocumented, and with serology inapparent in-fections can be recognized.

Isolation of a virus from a newborn or fetus isthe most direct method of establishing the

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diagnosis of congenital infection. The develop-ment of micro tissue culture and serologicaltechniques has reduced the cost of viral diagno-sis, thus making it a feasible procedure formost diagnostic laboratories (28, 87). Virusisolation results must be evaluated with theclinical history and antibody assays in both themother and infant. This is especially true ininterpretation of the isolation of entero virusesfrom throat swabs or stool specimens of themother or the newborn. Isolation of a virusfrom spinal fluid, urine, or lesion is usuallymore readily accepted as supportive evidenceof infection.Some viruses such as rubella, HSV, and

CMV produce chronic infection and are ex-

creted for many months after birth. Rubellavirus has been recovered from the throats ofcongenital newborns from birth to 2 years ofage. Virus has been isolated from cataracttissue taken from children as old as 31 months.Rubella virus has also been recovered fromspinal fluid and urine. These same sites arealso useful for isolation of other myxovirus suchas rubeola and mumps.CMV may be isolated from both throat

cultures and urine at approximately the samerate. More recently, cervical cultures have beenreported to show much higher frequencies ofinfection. Since CMV is very slow-growing,cultures should be kept for at least 6 weeksbefore considered negative. The virus is alsovery heat labile and thus may be lost byfreezing and thawing. Specimens should betransported rapidly to the laboratory and keptcool, not frozen, before they are inoculated intohuman fibroblastic tissue culture.HSV has been cultured from skin lesions,

throat swabs, and spinal fluids. This virusreproduces so rapidly that cytopathic effect intissue culture can be observed within 24 to 48 hafter inoculation.

Persistence of virus shedding with thesecongenital infections may be a factor in trans-mission of disease. Therefore, precautionsshould be implemented regarding exposure ofpregnant women, other infants, and nursingpersonnel caring for these infected infants.

Enteroviruses are relatively stable, and spec-imens can be stored at -20 C for short periodsof time. They can be isolated from stool, throatswabs, and occasionally spinal fluid.Vaccina and varicella viruses can be recov-

ered from skin lesions. Vaccinia as well as theother poxvirus, variola, can be recovered frompurulent or crusted lesions. Varicella on theother hand must be cultured before lesionsbecome crusted. They must be obtained from

the nonpurulent fluid. Isolates in human orsimian cells must be passed by transferringviable infected cells.A number of serological tests are available

for determination of antibody against the vi-ruses previously described (86). In order ofwidest application to the least commonly usedtests are: complement fixation, hemagglutina-tion inhibition, neutralization, passive hemag-glutination, and various precipitation tests.Selection of the method depends upon the virusstudied; some tests have distinctive advan-tages over others. The presence of antibodiesprovides indisputable proof of previous expo-sure to an antigen. However, high antibodytiters in a single specimen cannot be reliedupon entirely as presumptive evidence of arecent infection. The best approach is to dem-onstrate seroconversion with acute and con-valescent sera. Seroconversion can be regardedas a change from no detectable antibody topresence of antibody or a change in antibodytiter from one level to a fourfold higher level.With this approach, serological confirmation ofa recent infection can be documented and alsoprove that it occurred during pregnancy.Sero-immunoglobulins consist mainly of

three classes; IgG, IgM, and IgA. IgG normallycrosses the placenta during gestation and maybe found in fetal blood (4). Thus IgG antibodypresent in maternal blood will also be found inapproximately the same titer in the infant.Passively transferred antibody has a half-life of3 to 4 weeks. Therefore, a serum obtained froman infant at 4 or 5 months of age should show asignificant natural drop in IgG antibody titer orbe completely undetectable. However, if thespecific antibody level to an infectious agentremains the same or becomes elevated, it maybe a clue to congenitally or neonatally acquiredinfection.

Since IgM antibody is not passed transpla-centally from mother to fetus under normalcircumstances, an elevated IgM in cord serumis indicative of immunoglobulin synthesis bythe fetus in response to antigenic stimulation.Results of studies have indicated that mostinfants with severe congenital infections haveraised IgM levels. The value of this test hasbeen compared to a white-blood count in that itis usually increased in viral and bacterialinfection but is not specific. Approximately20% of children with increased IgM in thenewborn period can be identified as havingbeen infected with HSV, CMV, rubella virus,toxoplasmosis, or syphilis. The remaining 80%are unexplained. Long follow-up of a largenumber of newborns will be needed to deter-

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mine the strength of correlation of in uteroinfection with the presence of high levels ofIgM. Tests for specific IgM antibody utilizingthe indirect fluorescent-antibody techniqueshave been developed for rubella (7), CMV (35),and HSV (67). Although the availability ofthese tests is still restricted to the largerlaboratories, the specific IgM antibody testsshould prove extremely helpful in establishingthe early diagnosis of congenital infections.Appropriate treatment or other procedurescould then be instituted.

Concluding StatementThe role of viruses in congenital malforma-

tion will become more clearly evident as newtechniques for diagnosis become available.Further development of animal models of con-genital viral infections will help contribute toour knowledge of the number of variableswhich influence the frequency of congenitalinfection and their pathogenic processes.The possibility that viruses may remain in a

latent stage after being congenitally acquiredraises the question of the extent of their con-tribution to neurologic dysfunction in adultlife. Relationship of these viruses in initiationof malignancy and immunological dysfunctionsis also a possibility.

Since some viruses such as CMV and herpes-virus have the capability of latency, productionof safe vaccines may be difficult. The develop-ment of chemotherapeutic agents, therefore,becomes a very important area for study.Effective control of these diseases requiresincreased knowledge of the parasitic relation-ship of viruses to cells at a molecular level.

ACKNOWLEDGMENTSWe gratefully acknowledge the assistance in the

preparation of this manuscript provided by DianneEdwards and Renee Traub.

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