amniotic fluid

An epidemic of Zika virus infection began in Brazil in April 2015, and six months later there was a surge in the number of infants born with microcephaly. Confirming that Zika virus causes microcephaly will require much more information than is currently available. So far there have been few isolations of Zika virus RNA from microcephalic fetuses or amniotic fluid.

A single case report revealed the entire Zika virus genome in fetal brain tissue from a 25 year old who developed fever, muscle and eye pain, and rash during the 13th week of gestation in Natal, Brazil. The fetus was aborted at 28 weeks of gestation when fetal abnormalities, including microcephaly, were detected. Virus particles 42 to 54 nm in diameter were detected in the brain by electron microscopy.

It is probably not normal to have Zika virus in the fetal brain. However, its presence there might be a consequence of microcephaly, not a cause. As Dr. Steven Seligman writes at ProMedMail, “It is possible that brain tissue in cases of microcephaly become susceptible to Zika virus infection by a mechanism such as diminution of he blood-brain barrier.”

The entire Zika virus genome has also been detected in amniotic fluid, which surrounds the developing fetus. Two pregnant women from the state of Paraíba in Brazil reported clinical symptoms early in pregnancy consistent with Zika virus infection (fever, myalgia, rash). Microcephaly was diagnosed at 21 weeks gestation by ultrasound, and 7 weeks later samples of amniotic fluid were obtained by amniocentesis.

Amniotic fluid was centrifuged to purify virus particles, and RNA was extracted, copied into DNA by reverse transcriptase, amplified by polymerase chain reaction and subjected to deep sequencing.

The complete Zika virus genome sequence was obtained from one sample, and two smaller genome fragments from the second. Sequence analyses revealed 97-100% similarity with Zika viruses isolated from French Polynesia in 2013.

IgM antibodies to Zika virus were detected in both amniotic fluid samples, indicating that the fetus was likely infected and mounting an immune response against the virus (this antibody does not cross the placenta). In contrast, serum and urine from both mothers was negative for Zika virus IgM. This antibody appears first during infection, then subsides as levels of IgG antibody rise. It is possible that the mothers were infected with Zika virus early in pregnancy and cleared the infection, but the virus entered the fetus where it persisted.

Even if Zika virus does cause birth defects, a vaccine will likely not be available for another two years. In the meantime it would be highly advisable to practice mosquito avoidance and control.

Update 2/24/16: I asked Carolyn Coyne how a virus might reach the amniotic fluid. Her reply:

Amniotic fluid is mainly urine from the baby (after month 4ish) so if the virus is being shed in the urine, that is one way. Cells from the baby are also shed into the fluid (these are usually skin cells, but I imagine could also be from the mouth as the baby is usually drinking amniotic fluid at later stages of gestation). I will note that this is usually in a normal pregnancy and I imagine is the fetus were dying/dead (a fetus can die in utero and not be miscarried for a shockingly long time sometimes), the virus might easily enter the amniotic fluid as the fetus begins to decompose (which also happens in utero).

The two routes of entry are hematogenous or ascending. In hematogenous infections, virus present in the maternal blood would have to cross the placenta across the villous trees. In an ascending infection, the virus would be introduced into the vagina, then would have to bypass the cervix and still have to cross the placenta to access the fetus. Usually ascending infections are associated with bacteria (from UTIs mainly, but can be other). In either case, the placenta is there and would have to be crossed.

Update 2/25/16: A report in PLoS Neglected Tropical Diseases describes finding Zika virus RNA by RT-PCR in neuronal tissues but not in heart, lung, liver or placenta, in a stillborn infant with microcephaly and hydrops fetalis. The 20 year old woman from Salvador, Brazil denied having any symptoms consistent with Zika virus infection, but only one in five infections are symptomatic. As in the case described above, we do not know if Zika virus caused the fetal defects, or if virus was able to invade the fetus as a consequence of severe developmental damage.