Scientists from the Instituto de Química of the Universidade de São Paulo (USP) and Fiocruz at Rio de Janeiro and Bahia have identified major lipid alterations in the plasma of newborns exposed to the Zika virus. These results can contribute to the early diagnosis and monitoring of congenital Zika, both in babies with microcephaly and asymptomatic ones. When babies were born with microcephaly during the 2015-2016 Zika outbreak in Brazil, the government declared a public health emergency, although the association between the disease and microcephaly was not yet confirmed. It is now known that the Zika virus reaches the placenta and triggers inflammation that can cause placental insufficiency, resulting in the deficiency on the delivery of lipids and leading to brain and retinal deficits during fetal development.

“With these results, we have a molecular signature that could be used as a biomarker for children who were exposed to the virus during the prenatal period,” said Marcos Yukio Yoshinaga, a postdoctoral fellow at the laboratory of Professor Sayuri Miyamoto, from the IQ-USP and the RIDC Redoxoma, and coordinator of the research published in the journal PLOS Neglected Tropical Diseases.

Congenital Zika, characterized by the virus transmission of the virus from mother to baby during pregnancy, is a syndrome with a broad spectrum of clinical presentations, ranging from asymptomatic cases to microcephaly and other neurodevelopmental abnormalities manifested in early childhood. Zika virus-exposed newborns who do not have microcephaly can develop significant abnormalities 1 to 3.5 years after birth as revealed by brain imaging and neurodevelopmental evaluations.

During early brain development, lipids play a central role in metabolism, membrane structure, and signaling. Transplacental lipid transport is delicately regulated during pregnancy. Changes in placental lipid metabolism can impact embryonic development, especially in the brain and eyes that are highly dependent on the transfer of polyunsaturated fatty acids, such as docosahexaenoic (DHA) and arachidonic (ARA) acids, across the placenta.

Co-author of the article, Isadora Cristina de Siqueira, a infectologist and researcher at Fiocruz-Bahia, points out that most studies on congenital Zika infection found in the literature are related to children’s epidemiological and clinical descriptions. “We believe that there are a huge number of children who were affected with a lighter condition and who need long-term follow-up. We don’t have anything palpable as laboratory markers, we don’t have a follow-up or severity biomarker. This study brings new information about the pathogenesis of the disease, it shows that less affected children also have changes in lipids, so it brings laboratory markers that can be used in practice”. She also highlighted the importance of the collaboration between researchers and institutions from three Brazilian states.

For Professor Sayuri Miyamoto, the quality of the experimental analysis and the precision in the identification of the lipid species were fundamental for the research results. “The lipidomics platform implemented in Redoxoma allowed this new study showing the potential of this technique to discriminate important markers for diseases.”

Oxidized lipids

In 2016, Isadora conducted a study at the Jose Maria Magalhães Netto public maternity hospital in Salvador, which was one of the first cities hit by the epidemic of babies with microcephaly, and established active surveillance to identify pregnant women who had reports suggestive of Zika during pregnancy. At that time, researchers collected blood samples from newborns umbilical cord. For the present study, samples from three groups were used: 10 uninfected babies, 9 asymptomatic infected, and 11 infected that presented microcephaly.

The samples were subjected to lipid extraction and lipidomics analysis by ultra-high-performance liquid chromatography coupled with mass spectrometry. “We used an approach called untargeted lipidomics, that is, we do not have pre-established targets”, explained Adriano Britto Chaves-Filho, a postdoctoral fellow at IQ-USP and first author of the article with Nieli Rodrigues da Costa Faria, then a postdoctoral fellow at the Laboratório de Flavivírus at Fiocruz in Rio de Janeiro, under the supervision of researcher Ana Maria Bispo de Filippis. As there are few published works on newborn plasma, the method was ideal for discovering lipids in this medium. In the samples, 274 individual lipid species were identified, distributed in 20 classes and subclasses.

The results revealed that, in newborns with microcephaly, the plasma concentrations of hydroxyoctadecadienoic acid (HODE), mainly the 13-HODE isomer, were higher compared to the other two groups (newborns without microcephaly exposed to the virus and the control group). HODE is an oxidized fatty acid, derived from linoleic acid, and is considered an oxidative stress marker, mainly in plasma. Total HODE concentrations were also associated with levels of other oxidized lipids and various circulating free fatty acids in newborns, indicating a possible plasma lipidome signature of microcephaly.

The researchers also observed that the group of newborns infected but without microcephaly exhibited higher plasma concentrations of the lipid lysophosphatidylcholine than the other groups, suggesting a possible interruption in the transport of polyunsaturated fatty acids across the fetal blood-brain barrier. Disruption of the absorption of these polyunsaturated fatty acids can lead to various brain and eye damage in babies. This happens, for example, when a very rare mutation inactivates the MfSd2A protein, responsible for the transport of polyunsaturated fatty acids, especially DHA, to the brain, and causes microcephaly. “When this protein doesn’t work, you have less transport of polyunsaturated substances to the brain and an accumulation of lysophosphatidylcholine, exactly the phospholipid that we saw accumulated as a molecular signature of asymptomatic babies. And recently it was reported that Zika inhibits the action of this protein,” said Marcos.

The mechanisms by which Zika virus infection leads to brain defects are not known. However, according to the researchers, observational and experimental studies have documented that the virus targets placental cells, resulting not only in increased systemic inflammation but also in significant changes in placental lipid metabolism.

Now, according to the researchers, further studies with larger groups will be needed to investigate the role of individual lipids in Zika virus neuropathogenesis and to transform the plasma lipid profile into a marker for early diagnosis of newborns with suspected exposure to the virus.

The article Plasma lipidome profiling of newborns with antenatal exposure to Zika virus, by Nieli Rodrigues da Costa Faria, Adriano Britto Chaves-Filho, Luiz Carlos Junior Alcantara, Isadora Cristina de Siqueira, Juan Ignacio Calcagno, Sayuri Miyamoto, Ana Maria Bispo de Filippis, and Marcos Yukio Yoshinaga can be read here.