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Evaluation & Testing

Following CDC guidance on the evaluation and testing for congenital and postnatal Zika virus infection can help to ensure that affected children receive the care and services they need.

Congenital Zika Virus Infection

Initial Evaluation

Because changes in the updated interim guidance for pregnant women may lead to fewer pregnant women without Zika symptoms being tested, it is critical that pediatricians ask about potential congenital Zika exposure for every newborn. All infants born to mothers with possible Zika virus exposure during pregnancy should receive a standard evaluation by a healthcare provider at birth and at each well-child visit. A standard evaluation should include the following:

  • A comprehensive physical exam (including growth parameters)
  • Age-appropriate vision screening and developmental monitoring and screening
  • Standard newborn hearing screening at birth, preferably using auditory brainstem response (ABR) methodology

As described in CDC’S Interim Guidance for the Diagnosis, Evaluation, and Management of Infants with Possible Congenital Zika Virus Infection — United States, October 2017, laboratory testing and additional clinical evaluation are recommended for infants with birth defects consistent with congenital Zika syndrome born to mothers with possible Zika virus exposure during pregnancy (regardless of the mother’s Zika virus testing results) and for infants without birth defects consistent with congenital Zika syndrome who were born to mothers with laboratory evidence of possible Zika virus infection during pregnancy.

For infants without clinical findings whose mothers did not have laboratory evidence of Zika virus infection during pregnancy, further testing for Zika virus and clinical evaluation are not recommended.

Recommendations for Clinical Evaluation

The initial evaluation of infants born to women with possible Zika virus exposure during pregnancy can be done before or after birth hospital discharge, taking into account hospital capabilities and the needs of the family.

Infants with birth defects consistent with congenital Zika syndrome born to mothers with possible exposure to Zika virus during pregnancy (regardless of mother’s Zika test results)

Healthcare providers should provide the following care for these infants:

  • Zika virus testing
  • Standard evaluation at birth
  • Head ultrasound within 1 month of birth
  • Comprehensive eye exam by an ophthalmologist within 1 month of birth
  • Automated auditory brain stem response (ABR) by 1 month of birth if infant passed the newborn hearing screen using only the otoacoustic emissions (OAE) method
  • Referral to a developmental specialist and early intervention services
  • Ensure family support is available
  • Consider consultation with specialists
    • Infectious disease specialist for evaluation of other possible infections and assistance with Zika diagnosis, testing, and counseling
    • Clinical geneticist to confirm phenotype and evaluate for other causes of clinical findings
    • Neurologist for comprehensive neurologic exam and consideration of other evaluations, like advanced imaging, within 1 month of age
    • Other potential specialists depend on clinical findings of the infant
  • Maintain vigilance for other clinical findings
    • Signs of increasing intracranial pressure (such as vomiting or mental status changes) should prompt imaging of the brain to assess for onset of hydrocephaly after birth
    • Monitor feeding for signs of dysphagia or problems with swallowing
    • Consider diaphragmatic paralysis among infants with respiratory distress

Infants without birth defects consistent with congenital Zika syndrome who were born to mothers with laboratory evidence of possible Zika virus infection during pregnancy

Healthcare providers should provide the following care for these infants:

  • Zika virus testing
  • Standard evaluation at birth
  • Head ultrasound within 1 month of birth
  • Comprehensive eye exam by an ophthalmologist within 1 month of birth
  • Automated auditory brain stem response (ABR) by 1 month of birth if infant passed the newborn hearing screen using only the otoacoustic emissions (OAE) method
  • Referral to specialists for any signs associated with congenital Zika virus infection

Infants without birth defects consistent with congenital Zika syndrome who were born to mothers without laboratory evidence of possible Zika virus infection during pregnancy

Testing infants in this group for Zika virus infection is not routinely recommended. Further evaluation beyond the standard evaluation and preventive care is not routinely indicated unless abnormalities are noted at any time.

Imaging

A head ultrasound is recommended within 1 month of birth for

  • Infants with birth defects consistent with congenital Zika syndrome born to mothers with possible Zika virus exposure during pregnancy (regardless of mother’s Zika test results), and
  • Infants without birth defects consistent with congenital Zika syndrome who were born to mothers with laboratory evidence of possible Zika virus infection during pregnancy.

For infants without birth defects consistent with congenital Zika syndrome born to mothers who were exposed to Zika virus but without laboratory evidence of Zika virus infection, a head ultrasound is not routinely recommended.

Testing

Who to Test

Testing is recommended for

  • Infants with birth defects consistent with congenital Zika syndrome born to mothers with possible Zika virus exposure during pregnancy (regardless of mother’s Zika test results), and
  • Infants without birth defects consistent with congenital Zika syndrome who were born to mothers with laboratory evidence of possible Zika virus infection during pregnancy.

Laboratory testing of infants should be performed as early as possible, preferably within the first few days after birth, although testing specimens within the first few weeks to months after birth might still be useful.  However, distinguishing between congenital, perinatal, and postnatal infection is difficult in infants who live in areas where there is ongoing transmission of Zika virus and who are not tested soon after birth

Testing infants without birth defects consistent with congenital Zika syndrome who were born to mothers with possible Zika virus exposure during pregnancy but without laboratory evidence of possible Zika virus infection during pregnancy is not routinely recommended.

Diagnosis & Testing

The optimal assays, specimens, and timing of testing for congenital Zika virus infection are unknown. Recommended laboratory testing for possible congenital Zika virus infection includes evaluation for Zika virus RNA in infant serum and urine and Zika virus IgM antibodies in serum. In addition, if cerebrospinal fluid (CSF) is obtained for other purposes, NAT and IgM antibody testing should be performed on CSF because CSF was the only sample that tested positive in some infants with congenital Zika virus infection. Testing of cord blood is not recommended because it can yield false positive and false negative test results.

A Zika virus NAT positive result in an infant sample confirms the diagnosis of congenital Zika virus infection. Zika virus IgM detected in an infant with a negative NAT result should be interpreted as probable congenital Zika virus infection. If neither Zika virus RNA nor Zika IgM antibodies are detected on the appropriate specimens (e.g., serum or urine) obtained within the first few days after birth, congenital Zika virus infection is unlikely.

The plaque reduction neutralization test (PRNT), which measures virus-specific neutralizing antibodies, can be used to help identify false-positive results. If the infant’s initial sample is IgM non-negative and NAT negative, but PRNT was not performed on the mother’s sample, PRNT for Zika and dengue viruses should be performed on the infant’s initial sample. If Zika virus PRNT is negative, this suggests that the infant’s Zika virus IgM test is a false positive.

PRNT cannot distinguish between maternal or infant antibodies. Maternal antibodies in the infant are expected to wane by 18 months. PRNT might help confirm or rule out infection in a sample collected from an infant aged 18 months whose initial sample collected at birth was IgM non-negative and neutralizing antibodies were detected by PRNT in either the infant’s or mother’s sample. If PRNT results at 18 months are negative, the infant is considered not to have congenital Zika virus infection. If PRNT results are positive, congenital Zika virus infection is presumed, but postnatal infection cannot be excluded, especially among infants living in an area with active Zika virus transmission.

Infant test results**

NAT IgM Interpretation
Positive Any result Confirmed congenital Zika virus infection
Negative Non-negative Probable congenital Zika virus infection†§
Negative Negative Congenital Zika virus infection unlikely†¶

Footnotes
**Infant serum, urine, or cerebrospinal fluid
Laboratory results should be interpreted in the context of timing of infection during pregnancy, maternal serologic test results, clinical findings consistent with congenital Zika syndrome, and any confirmatory testing with plaque neutralization testing (PRNT)
§ If Zika virus PRNT is negative, this suggests that the infant’s Zika virus IgM test is a false positive
Congenital Zika virus infection is unlikely if specimens are collected within the first few days after birth; however, healthcare providers should remain alert for any new findings of possible congenital Zika virus infection

Possible Limitations of Infant Laboratory Testing

Optimal tests, types of specimens to test, and timing to test for congenital Zika virus infection are unknown. In addition, two recent studies describe a small number of infants with clinical findings consistent with congenital Zika syndrome in whom results of laboratory testing for Zika virus infection were negative (1,2). Negative test results might occur in an infant with clinical findings of possible congenital Zika virus syndrome for several reasons:

  • The clinical findings are due to another cause
  • Testing was incomplete (e.g., RNA testing without antibody testing), performed on suboptimal specimens (e.g., cord blood rather than blood obtained from the infant), or performed too late (e.g., after RNA and IgM antibodies had cleared or waned) (3)
  • The fetus did not mount an IgM antibody response; it is unknown if some fetuses infected early in gestation might not mount an IgM antibody response (4)
Remain Alert to Possible Clinical Findings

For infants without clinical findings of congenital Zika syndrome and without laboratory evidence of Zika virus infection born to women with possible Zika virus exposure during pregnancy, healthcare providers should remain alert for any new findings of possible congenital Zika virus infection. If findings suggestive of congenital Zika syndrome are identified at any time, refer to appropriate specialists and evaluate for congenital Zika virus infection.

Postnatal Zika Virus Infection

Criteria for testing will vary by state. However, postnatal Zika virus disease should be suspected in an infant or child aged <18 years who 1) traveled to or resided in an area with risk of Zika virus infection within the past 2 weeks or who might have been exposed to Zika virus through sexual contact with a partner who traveled to or resided in an area with risk of Zika, and 2) has one or more of the following manifestations: fever, rash, conjunctivitis, and arthralgia. Because perinatal transmission of Zika virus from mother to infant during delivery is possible, Zika virus disease should also be considered in an infant 1) who has one or more of the following manifestations: fever, rash, conjunctivitis, and arthralgia during the first 2 weeks of life, and 2) whose mother was potentially exposed to Zika virus within approximately 2 weeks of delivery.

Arthralgia can be difficult to detect in infants and young children and can manifest as irritability, walking with a limp (for ambulatory children), difficulty moving or refusing to move an extremity, pain on palpation, or pain with active or passive movement of the affected joint.

Diagnosis & Testing

Zika virus NAT should be performed on serum and urine collected <14 days after onset of symptoms in patients with suspected Zika virus disease. A positive Zika virus NAT confirms Zika virus infection. However, because Zika virus RNA in serum and urine decreases over time, a negative NAT does not rule out Zika virus infection; in this case, serologic testing should be performed. If Zika virus NAT results are negative for both serum and urine specimens, serum should be tested by antibody detection methods.

Serologic assays can also be used to detect Zika virus-specific IgM and neutralizing antibodies, which typically develop toward the end of the first week of illness. A positive IgM result does not always indicate Zika virus infection and can be difficult to interpret because cross-reactivity with related flaviviruses (e.g., dengue, Japanese encephalitis, West Nile, yellow fever viruses) can occur. A positive Zika virus IgM result may reflect previous vaccination against a flavivirus, previous infection with a related flavivirus, or current infection with a flavivirus, including Zika virus.

PRNT can be performed to measure virus-specific neutralizing antibodies to confirm primary flavivirus infections and differentiate from infections with other related viruses. PRNT can be performed to measure virus-specific neutralizing antibodies to Zika virus, but neutralizing antibodies may still yield cross-reactive results in a person who was previously infected with another flavivirus, such as dengue, or has been vaccinated against yellow fever or Japanese encephalitis.

Ordering Tests

Zika virus testing is performed at many state and territorial health departments, at CDC, and at commercial laboratories that perform Zika testing using a validated assay with demonstrated analytical and clinical performance. Healthcare providers should contact their local, state, or territorial health department to facilitate testing. See Testing for Zika Virus for information on Zika virus testing.

Testing Challenges

Zika virus testing in infants and children has several challenges. NAT tests may not detect Zika virus RNA in an infant who had Zika virus infection in utero or in a child if the period of viremia has passed. Serologic tests for Zika virus can be falsely positive because of cross-reacting antibodies against related flaviviruses (e.g., dengue and yellow fever viruses). Plaque-reduction neutralization testing (PRNT) can be performed to measure virus-specific neutralizing antibodies to Zika virus, but neutralizing antibodies may still be cross-reactive.  In addition, PRNT cannot distinguish between maternal and infant antibodies in specimens collected from infants at or near birth. Based on what is known about other congenital infections, maternal antibodies are expected to become undetectable by 18 months of age and might become undetectable earlier.  It is important to work closely with local, state, or territorial health departments to ensure that the appropriate test is ordered and interpreted correctly.

Clinician Resources

Evaluation and Management for Infants with Possible Congenital Zika Infection


Implementing CDC Guidance for Clinical Management and Evaluation of Infants Born to Mothers with Possible Zika Virus Exposure During Pregnancy; and Testing of Placental, Fetal, or Infant Autopsy Tissues


Zika Test Result Interpretation Table


Measuring Infant Head Circumference: An instructional video for healthcare providers


Measuring Head Circumference

Footnotes

*Laboratory evidence of maternal Zika virus infection includes 1) Zika virus RNA detected by nucleic acid testing (NAT) (e.g., rRT-PCR) in any clinical specimen; or 2) positive Zika virus IgM antibodies with confirmatory neutralizing antibody titers obtained via plaque reduction neutralization testing (PRNT). Confirmatory neutralizing antibody titers are needed in addition to IgM antibodies for laboratory evidence of maternal Zika virus infection.

§ An epidemiologic link includes travel to or residence in an area with risk of Zika or sex without a condom with a partner who traveled to or lived in such an area.

For pregnant women with possible Zika virus exposure who seek care >12 weeks after symptom onset or possible exposure, IgM antibody testing might be considered. However, a negative IgM antibody test or rRT-PCR result >12 weeks after symptom onset or possible exposure does not rule out recent Zika virus infection because IgM antibody and viral RNA levels decline over time.

References

  1. de Araujo TV, Rodrigues LC, de Alencar Ximenes RA, de Barros Miranda-Filho D, Montarroyos UR, de Melo AP, et al. Association between Zika virus infection and microcephaly in Brazil, January to May, 2016: Preliminary report of a case-control study. Lancet Infect Dis 2016;16(12):1356–1363.
  2. Melo AS, Aguiar RS, Amorim MM, Arruda MB, Melo FO, Ribeiro ST, Batista AG, Ferreira T, Dos Santos MP, Sampaio VV, Moura SR, Rabello LP, Gonzaga CE, Malinger G, Ximenes R, de Oliveira-Szejnfeld PS, Tovar-Moll F, Chimelli L, Silveira PP, Delvechio R, Higa L, Campanati L, Nogueira RM, Filippis AM, Szejnfeld J, Voloch CM, Ferreira OC Jr, Brindeiro RM, Tanuri A. Congenital Zika virus infection: Beyond neonatal microcephaly. JAMA Neurol 2016;73(12):1407–1416.
  3. Honein MA, Dawson AL, Petersen EE, Jones AM, Lee EH, Yazdy MM, et al. Birth defects among fetuses and infants of US women with evidence of possible Zika virus infection during pregnancy. JAMA 2017;317(1):59–68.
  4. Alford CA, Foft JW, Blankenship WJ, Cassady G, Benton JW. Subclinical central nervous system disease of neonates: A prospective study of infants born with increased levels of IgM. J Pediatr 1969;75:1167–1178.
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