Emerging Concepts in Congenital Cytomegalovirus

CMV is a member of the Herpesviridae family of viruses, and has a large, double-stranded DNA genome, with a total coding content of >230 kbp of sequence.¹⁰  CMV genomes consist of unique long and unique short segments, each bracketed by terminal repeats¹⁰ ; 4 different isomeric configurations of these segments are therefore possible in any given virus particle (Fig 1). Hence, some viral gene products are diploid, with duplicate copies in both the terminal repeat and internal repeat regions of the unique long or short regions of the 2 genome segments, respectively. From a clinical perspective, an understanding of these mechanisms of strain diversity informs the phenomenon of reinfection.¹¹  It has become increasingly clear that reinfection with novel variants can occur in women with prior prepregnancy immunity, and that such strain variants can be transmitted to the fetus.¹²  The rapid generation of novel CMV strains has also posed a challenge for vaccine and antiviral medication development given the rapidly-evolving nature of the “target.”¹³ 

The CMV genome includes many genes directed at evading, blocking, impairing, and modulating host viral defenses.^14–16  CMV envelope glycoproteins, structural proteins, and nonstructural proteins (Table 2) play important roles in the virus’s carefully choreographed host-immune evasion phenotype. These gene products function by interfering with the generation of CD4+ and CD8+ T-cell responses, impeding natural killer cell clearance of infection, altering host cytokine and chemokine responses, and blocking innate immune responses mediated by interferons.¹⁴  Together, CMV immune evasion genes function to subvert protective immunity and likely play a key role in promoting life-long infection, periodic reactivation, and reinfection with novel strain variants.

Advances in the molecular characterization of patterns of host gene expression have facilitated study of the host transcriptome as a biomarker for gauging the risk of cCMV sequelae.¹⁷  In one such study, blood transcriptional profiles were compared in infants with symptomatic and asymptomatic cCMV infection. A 16-gene host genetic “signature” was identified, which was associated with the development of SNHL in both infant categories.¹⁷  Understanding of the components of viral transcriptomics that contribute to outcomes may facilitate future clinical management by identifying infants that need more detailed audiological or neurodevelopmental follow-up or may be more likely to benefit from antiviral therapy. Similarly, viral strain variation has been shown to impact the pathogenesis of CMV-induced injury.^18,19  Although not yet available for bedside diagnostics, development of viral genotypic assays to identify “high-risk” variants may prove to be a useful adjunct to identifying reinfections of previously seropositive women during pregnancy, and to direct clinical management of infants with cCMV.

After an initial or primary infection, CMV establishes a lifelong infection in the host. Although the virus becomes quiescent, establishing latency in hematopoietic progenitor cells,²⁰  reactivation occurs throughout the lifetime of the host under conditions of stress, intercurrent illness, and immune suppression.²¹  Reactivation of latent CMV is thought to rarely cause disease in immunocompetent hosts but can lead to heightened morbidity and mortality in immunocompromised individuals.²⁰  Prior infection with CMV and circulating antibodies do provide some protection, however, lowering the risk of vertical transmission and severity of manifestations of congenital infections.²²  Because of the high prevalence of CMV in the US population, it is estimated that most infants with cCMV are born to women with preexisting exposure to the virus.²³ 

As an overview, most infants (85% to 90%) are born with clinically inapparent infections, and the remaining 10% to 15% are born with symptomatic disease (visible signs, laboratory abnormalities, and/or intracranial abnormalities).^24–26  The absence of visible signs at birth is not always associated with an asymptomatic infection (defined as having no signs, laboratory, or intracranial abnormalities at birth).²⁷  Some well-appearing infants with cCMV may still have intracranial involvement and thus be categorized as symptomatic.²⁸  Less than 5% of infants with cCMV present with the classic “blueberry muffin” triad (small for gestational age, direct hyperbilirubinemia, and a petechial rash). Infants with symptomatic disease are at heightened risk of neonatal death and of long-term neurodevelopmental sequelae.^25,27,29,30  SNHL may be present at birth or develop over childhood in those with asymptomatic or symptomatic infections.^26,31  Infants with isolated SNHL are generally considered as having an asymptomatic infection, although some experts argue that SNHL is a marker of central nervous system (CNS) involvement and should be treated similarly to symptomatic disease.²⁷ 

Congenital CMV occurs in 3 to 6 infants in every 1000 live births in the United States each year.³²  Studies of cCMV prevalence in the United States have identified disparities among marginalized communities. A study of roughly 100 000 neonates screened at birth found cCMV prevalence to be highest among African American infants (9.5 per 1000 live births), followed by multiracial infants (7.8 per 1000 live births), with the lowest prevalence among non-Hispanic White infants (2.7 per 1000 live births) and Asian infants (1.0 per 1000 live births).³²  Even after adjusting for socioeconomic status and maternal age, African American infants were nearly twice as likely to have cCMV compared with non-Hispanic White infants.^32,33  Furthermore, Native Americans and African Americans have also been found to be roughly 2 times more likely, respectively, to die of cCMV than non-Hispanic White infants.³⁰  Social determinants of health and systemic racism may contribute to high rates of CMV seropositivity among pregnant women living in urban, low-income areas, particularly among those of racially marginalized groups.³⁴ 

Attention has recently been drawn to the role cCMV plays in intrauterine fetal demise and miscarriage. A large multisite US study examining intrauterine fetal demise due to infectious etiologies identified cCMV as the likely cause in 8% of cases.³⁵  This builds on prior work from Australia that identified CMV on fetal autopsy in 15% of stillbirths at >20 weeks gestational age.³⁶  A meta-analysis found maternal CMV infection during pregnancy to be a risk factor for spontaneous abortion (OR 1.61, 95% CI 1.14%–2.27%) and intrauterine fetal demise (5.74, 95% CI 2.04%–16.12%).³⁷ 

CMV is transmitted horizontally in bodily fluids, including urine, saliva, vaginal secretions, semen, breast milk, and plasma. Women with more than 1 child and those who work with children are at the highest risk of having an infant with cCMV, as close exposure to body fluids from young children increases the likelihood of horizontal transmission.³⁸  Toddler and preschool-aged children shed CMV in their saliva and urine for months after an infection, long after their symptoms have resolved.^39,40  Behaviors to lower an individual’s risk of horizontal transmission include hand hygiene after changing diapers, not sharing food or utensils with young children, and avoiding kissing young children on the lips.^24,41  Pediatricians are uniquely positioned to educate mothers about the risks of and prevention strategies against acquisition of CMV from their toddlers, which in turn may be of benefit for future pregnancies.^42–44  Prenatal education about CMV prevention behaviors is standard of care in several countries.^45–48  Furthermore, such behavioral interventions have been found to be acceptable to women and associated with lowered rates of seroconversion in pregnancy.^41,42,47  An on-going randomized controlled trial (RCT) of prenatal education, versus standard of care, may provide even more supportive evidence (Table 1).⁴⁹  Early data suggest that the prevalence of cCMV dropped dramatically in 2020 as compared with 2019, coinciding with the COVID-19 pandemic.⁵⁰  It may be that increased hand hygiene and the closure of childcare centers resulted in fewer opportunities for horizontal transmission and thus, fewer cases of cCMV.⁵⁰ 

Mechanisms of and protective factors against vertical transmission are an area of active inquiry. A recent meta-analysis of twin pregnancies affected by CMV found an elevated rate of vertical transmission (59% vs 31% in singleton pregnancies) and cCMV discordance in 50% of twin pregnancies (21 of 42).⁵¹  cCMV concordance occurred in 95% of monozygotic twin pairs and 38% of dizygotic twin pairs, resulting in an estimated hereditability of 94%.⁵¹  These findings point toward possible placental differences or genetic factors conferring increased susceptibility to vertical transmission of CMV.⁵¹  Genetic variations in maternal toll-like receptor single nucleotide repeats have been implicated both in increased⁵²  and decreased rates of CMV infection during pregnancy.⁵³ 

Prenatal treatment to lessen the risk of vertical CMV transmission have been the focus of some small studies and 2 recent RCTs.^54–57  Hughes et al sought to evaluate whether cytomegalovirus immune globulin lowered the risk of vertical transmission but the trial was stopped early because of futility and safety concerns.⁵⁵  A randomized double-blind placebo-controlled trial of valacyclovir in pregnant women with primary CMV infections in the first trimester found lower odds of vertical transmission in the treatment group.⁵⁷  The potential impact of in-utero antiviral treatment in ameliorating sequelae in CMV-infected fetuses has also been studied. An open-label trial of valacyclovir in women pregnant with mildly symptomatic cCMV-infected fetuses found that a greater proportion delivered infants with asymptomatic infections (82%) as compared with rates of untreated controls from the literature (43%).⁵⁸ 

Newborn screening for cCMV has become more common over the last several years, with the goal of early identification of and intervention for affected infants. Hearing-targeted cCMV screenings target testing only to infants who fail their newborn hearing screening, which is nearly eightfold more common in infants with cCMV (versus those without cCMV),⁵⁹  unlike a universal screening approach in which all neonates are tested. However, Fowler et al found that only 57% of infants with cCMV associated hearing-loss in the neonatal period were identified by hearing-targeted cCMV screening protocols, which is arguably suboptimal when compared with universal screening.⁵⁹  Seven states now have legislative mandates for hearing-targeted cCMV screening (Illinois, Iowa, Connecticut, New York, Utah, and Virginia), and 1 state (Minnesota) recently passed a universal screening bill.⁶⁰  There remains an active debate about the cost-effectiveness of cCMV screening programs, as well as the economic burden of cCMV.⁶¹ 

Studies support parental acceptance of and a desire for cCMV newborn screening.^62,63  Tastad et al found that 96% of women supported newborn screening once they were educated about the public health significance of the topic.⁶²  In a survey study of 3922 participants, most parents strongly or somewhat agreed that they would want to have their newborn tested for cCMV, even if it was not performed routinely (84%), if they had to pay out-of-pocket for the test (87%), or if CMV-related problems never developed in their child (84%).⁶³ 

Newer point-of-care testing platforms for cCMV have allowed for the growth of cCMV screening programs.⁶⁴  For context, screening or testing for cCMV may be performed using nucleic acid amplification testing (eg, polymerase chain reaction [PCR] using dried blood spot [DBS], saliva, or urine). The timing of screening and confirmatory testing for cCMV is important and must be performed on a fresh sample (eg, urine or saliva) before 21 days of age, after which it is challenging to distinguish a congenital infection from a postnatal exposure.²⁷ 

If saliva-based screening is employed, strong consideration should be provided to confirming positive saliva results with a urine PCR because of the substantial risk of a false positive result attributable to the presence of “carry-over” CMV DNA in saliva due to the presence of viral genome in breast milk from a seropositive nursing mother. The risk of false positive results can be minimized by collecting the specimen >1 hour after breastmilk consumption.⁶⁵  Retrospective cCMV testing can be performed on saved samples collected at birth, such as cord blood, umbilical stump, or DBS remnants. Recent work found the sensitivity of DBS CMV PCR in 2 laboratories to be 74% and 77%, with a 2-primer approach yielding a sensitivity of 86%, suggesting that newborn screening programs focused on DBS PCR may need to use multiprimer and multiplex approaches to maximize sensitivity.⁶⁶  Whereas somewhat higher than the 30% to 40% sensitivity reported a decade ago, present day DBS PCR sensitivity remains relatively low as compared with the near 99% sensitivity of saliva and urine assays.^64,66  This concern may be mitigated by the fact that DBS collection is a standard practice in newborns to screen for a variety of genetic and metabolic diseases; thus, the infrastructure exists to rapidly incorporate DBS screening into standard clinical care, greatly reducing the overall cost of such programs.

The presentation of cCMV varies widely across the spectrum of disease severity, although it is most commonly clinically inapparent.⁶⁷  A detailed review of the clinical signs and recommendations for management are beyond the scope of this paper; the following sources for reference on those subjects are recommended.^24,27  Once a diagnosis of cCMV is confirmed, additional diagnostic evaluation is critical in determining the presence of sequelae and the infection severity, which dictates further treatment and monitoring. Below, a summary of recent research surrounding the most common sequelae of cCMV is presented:

All infants with cCMV are considered to be at high risk for SNHL and warrant diagnostic testing and close audiologic follow-up, according to the most recent Joint Committee on Infant Hearing guidelines.⁶⁸  SNHL occurs in 15% to 25% of all children with cCMV over the course of childhood.^31,69  The development of SNHL is associated with first-trimester (versus third) infection, symptomatic disease, and intracranial involvement.^69,70  Unique to cCMV-related SNHL is the frequent progression and fluctuation of hearing thresholds, which occurs in roughly ∼45% of cases.^70,71  Work by Lanzieri et al reported SNHL at age 18 years in 25% of children with asymptomatic cCMV, 72% of whom had a delayed onset of SNHL.⁷² 

Symptomatic cCMV has been associated with several ophthalmologic manifestations, which can contribute to abnormal visual acuity and visual impairment over childhood and early adolescence. Children with symptomatic cCMV are more likely to have moderate or severe visual impairment compared with those with asymptomatic infection.⁷³  Overall, the presence of clinical symptoms of cCMV at birth, SNHL, and microcephaly have been found to be predictors of severe visual impairment.⁷³  Of note, children with asymptomatic infection and those with symptomatic disease with no evidence of retinal or posterior pathway involvement in early life have been found to have visual acuity comparable to healthy controls in adulthood.⁷⁴  Other work has also found little evidence of ocular sequelae in those with asymptomatic infections.^74,75  Yearly ophthalmologic evaluations are recommended for those with symptomatic cCMV disease.⁷³ 

Increased attention has been paid to the broad spectrum of neuroimaging abnormalities seen in cCMV, moving away from a dichotomous characterization of “normal” versus “abnormal.” Nonbinary severity rating scales covering the range of potential intracranial abnormalities may lead to more accurate neurodevelopmental prognoses.^76–81  A recent study found a MRI severity score to be a better predictor of adverse neurologic sequelae than the presence of symptoms at birth.⁸²  Expert cCMV clinical guidelines recommend all infants receive a cranial ultrasound (CUS), followed by a MRI, if abnormalities are seen, and antiviral treatment of those with CNS involvement.^27,48  Recent studies have highlighted the complementary nature of CUS and MRI for evaluating cCMV CNS involvement.^83,84  Work from the Netherlands found that 20% (93 of 480) of infants with cCMV with normal CUS findings had abnormalities found on MRI, 37% of whom were categorized as severely symptomatic based on MRI alone.⁸⁴  Vande Walle et al reported MRI abnormalities in 33% to 59% of infants with clinically inapparent cCMV in their cohort.⁸⁵  The clinical significance of subtle isolated MRI findings remains unclear.⁸⁶  Whereas severe lesions, such as polymicrogyria or lissencephaly, are likely irreversible regardless of antiviral treatment, some have proposed that infants with isolated white matter abnormalities may be more susceptible to treatment,⁸⁷  but this remains to be proven. Longitudinal studies of the neurodevelopmental outcomes of infants with isolated subtle MRI findings are needed to inform clinical guidance around neuroimaging and antiviral treatment.

CMV is a neurotropic virus, readily crossing the fetal blood-brain barrier and infecting and replicating in brain-resident cells, including astrocytes, neurons, and microglia.⁸⁸  The lesions caused by cCMV are best understood according to the timing of infection in pregnancy and corresponding stages of fetal brain development (Fig 2). Earlier trimester infections result in the most severe neuropathology because CMV disruption of fetal cortical development from aberrant neuronal migration, formation, and organization results in polymicrogyria, cerebellar hypoplasia, and more rarely, lissencephaly.^23,89,90  Infections later in pregnancy after the architecture of the fetal brain has developed may result in inflammation, causing white matter lesions and cysts.⁹¹ 

Potential molecular mechanisms of CMV-dysregulated cortical development have been explored. One study evaluated CMV-infected neural stem cells, demonstrating increased expression of the gene PAFAH1B1, encoding LIS1 (lissencephaly-1).⁹²  LIS1 is a component of the platelet activating factor acetyl-hydrolase complex, which is important in directing neuronal migration. Increased immunostaining for LIS1 in brains of congenitally infected fetuses, but not controls, has been found, strengthening the hypothesis that dysregulation of PAFAH1B1 provides at least one explanation for cCMV brain injury. Other molecular mechanisms may also mediate fetal CNS injury. CMV has been found to induce downregulation of the gene, nidogen 1 (NID1), a basement membrane protein important for neuronal migration, regulating Schwann cell proliferation, migration, and myelin production.⁹³  It is hypothesized that virus-induced downregulation of NID1 might offer CMV a means of increased distribution throughout the host.⁹³ 

The host inflammatory response can also contribute to CMV neuropathy. Infected microglia and astrocytes become active overproducers of cytokines, leading to a wide-spread proinflammatory response in the developing fetal brain.⁹⁴  Inflammatory infiltrates have been found in close approximation to CMV-positive neurons, suggesting a significant component of immune-mediated injury.^95,96  Similarly, CMV infection and inflammatory infiltrates have been found throughout the structures of the inner ear, including the stria vascularis, which plays a critical role in potassium and ion homeostasis in the cochlea.⁹⁷  It has been proposed that CMV-mediated dysregulation of such homeostatic mechanisms could, by altering the generation of endocochlear action potentials, produce SNHL.⁹⁸ 

Interventions and treatment of cCMV largely consist of standard management of sequelae (eg, antiepileptic drugs for seizures, physical therapy for gross motor delays), which are nonspecific to cCMV. Monitoring for audiologic, visual, and developmental sequelae is also a form of intervention. Below, recent findings in the field specific to the treatment of cCMV are highlighted.

In 2015, Kimberlin et al published the results of a RCT of a 6-month versus 6-week course of oral valganciclovir (the prodrug of IV ganciclovir) for infants with symptomatic cCMV, finding modest improvement in hearing and neurodevelopmental outcomes at 24-months in the group treated for 6 months.⁹⁹  Guidelines recommend valganciclovir treatment of infants with moderate-to-severe symptomatic cCMV. The possible benefits of therapy must be balanced with the risks of adverse events, particularly neutropenia.¹⁰⁰  The safety and efficacy of valganciclovir use in less severely affected infants has not been established; results of ongoing studies are much anticipated (Table 1).

For infants with severe-to-profound SNHL, cochlear implantation can now be performed before 9 months-of-age.¹⁰¹  Early cochlear implantation and language exposure has been associated with improved spoken language outcomes in children with SNHL.¹⁰²  Children deafened by cCMV have been found to have adequate access to sound for speech production, but their spoken language scores tend to be lower than children deafened by other etiologies (eg, Connexin 26 mutations), which is likely related to other cCMV comorbidities.¹⁰³ 

Most infants born with asymptomatic cCMV have been found to have neurodevelopmental outcomes in the typical range,²⁶  though recent work has highlighted some areas of risk. In 1 of the only longitudinal studies of children with asymptomatic cCMV followed into adolescence, Lopez et al found that those with SNHL (but not those with typical hearing) had lower measures of verbal skills as compared with hearing controls, which was likely attributable to hearing loss.¹⁰⁴  There were no differences in nonverbal intelligence or academic achievement in math and reading between adolescents with cCMV-associated SNHL and unaffected sibling controls.¹⁰⁴  Pinninti et al identified a high prevalence of vestibular (45%), gaze (46%), and balance (30%) disorders in children with asymptomatic cCMV with and without hearing loss.¹⁰⁵ 

Symptomatic cCMV is associated with greater risk of long-term disabilities. Lanzieri et al followed 76 children with symptomatic cCMV through median age 13-years, finding 43% with intellectual disability, 74% with SNHL, 27% with visual impairment, and 42% with both intellectual disability and SNHL.¹⁰⁶  Congenital CMV is also thought to be prevalent in children with cerebral palsy, although there does not seem to be a particular phenotype of cerebral palsy associated with cCMV.¹⁰⁷  Some preliminary studies point to a possible correlation between symptomatic cCMV and autism spectrum disorder; additional research is needed to establish causation.¹⁰⁸ 

Several candidate vaccines against CMV are currently in development (Table 1).⁷  Whereas the ideal population for a CMV vaccine would be young children who would carry immunity into adulthood, similar to the measles, mumps, and rubella vaccine, developers are first focusing on women of childbearing age. A vaccine in this population may serve to reduce horizontal infection to the mother, and thereby reduce vertical transmission to the fetus. Prospective studies of long-term immunity conferred by a CMV vaccine may provide support for childhood immunization. Strain variation has posed a challenge for vaccine development; although the molecular mechanisms leading to novel variants are different for CMV (a DNA virus) and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) (a RNA virus), strain variation complicates CMV vaccine design, just as it does for COVID-19 immunization strategies.¹⁰⁹ 

Subunit vaccines against CMV have focused on the 3 distinct regions of the CMV virus particle: the capsid, the tegument layer, and the envelope (Fig 3 and Table 2). The icosahedral capsid consists of 162 capsomere subunits that enclose the viral genome.¹¹⁰  In the mature virus, the capsid is surrounded by the tegument, a layer containing a 65-kilodalton phosphoprotein that is a major T-cell target referred to as pp65.¹¹¹  Finally, surrounding the tegument is the outer lipid bilayer envelope, containing virally encoded glycoproteins.¹¹²  The glycoprotein immunogens currently being explored in clinical trials as subunit vaccine candidates are outlined in detail in Table 1.

In addition to vaccine development, other areas of high priority in cCMV research include DBS PCR assay improvement, therapies to reduce vertical transmission, as well as safe and effective antiviral medications for all neonates with cCMV. In terms of clinical management, movement away from the binary categorization of cCMV as asymptomatic versus symptomatic toward a spectrum of disease may lead to more nuanced studies and treatment recommendations. Research is needed to improve our understanding of developmental outcomes and trajectories, based not only on presentation at birth, but also services and supports in childhood that may ameliorate outcomes. Identification of biomarkers of disease severity will aid in developing more nuanced studies and ultimately, interventions. Lastly, better elucidating the economic burden of cCMV disease, as well as costs of screening, treatment, and prevention programs is of high priority.⁶¹  These studies should include measures of direct healthcare costs, but also indirect costs, such as caregiver burden and quality of life.⁶¹  A better understanding of the societal impact of cCMV is important for public health prioritization to reduce pregnancy loss, childhood death, and disability from this common yet often inapparent virus.^113,114 

cCMV

congenital cytomegalovirus

CMV

cytomegalovirus

DBS

dried blood spot

PCR

polymerase chain reaction

SNHL

sensorineural hearing loss