Indian Scientists Have a New Hypothesis of Why Zika is So Deadly to Brains

Amid a global flurry of research into Zika, a team of Indian scientists have quietly emerged with a plausible theory that explains how Zika infected mothers are giving birth to babies with malformed brains.

Transmission electron microscope image of negative-stained, Fortaleza-strain Zika virus (red), isolated from a microcephaly case in Brazil. Credit: NIAID

Transmission electron microscope image of negative-stained, Fortaleza-strain Zika virus (red), isolated from a microcephaly case in Brazil. Credit: NIAID

The rising incidents of rare neurological disorders in places of Zika transmission led bodies like WHO and the US’s CDC to declare earlier this year that it was highly likely that Zika infection and neurological disorders like microcephaly and Guillain-Barré Syndrome are linked. Though Zika virus has been found in the placenta and amniotic fluid of babies born with microcephaly and even in the brain of one foetus which died of severe complications, the exact mechanism by which the virus causes these disorders remains largely a mystery.

A group of scientists from the All India Institute of Medical Sciences, National Brain Research Centre and University of Delhi, found it intriguing that the Zika-related brain abnormalities looked a lot like the brain abnormalities caused when a process called retinoic acid signalling gets disrupted. They investigated this similarity by analysing the Zika virus genome and comparing it to the genes involved in retinoic acid signalling.

Their results, published in the journal Frontiers in Human Neurosciences on August 9, show how Zika virus could be interfering in this pathway and wreaking havoc in brain development in growing foetuses. “If our hypothesis is confirmed, it could be a game changer in therapeutic approaches towards Zika virus associated brain malformation in infected foetuses,” said Muneeb Faiq, one of the authors of the paper, in an email interview.

As of August 2016, 15 countries, out of the 66 where vector-borne Zika virus transmission has taken place since 2015, have reported Zika-related congenital brain malformations, says the World Health Organisation (WHO). India has managed to stay out of these lists so far but that does not mean it always will. “The pattern of its spread suggests the travel of this threat towards the eastern part of the world also,” said Faiq, adding that the widespread presence of dengue (which is transmitted by the same mosquito that transmits Zika, Aedis aegypti) makes India automatically susceptible to a Zika outbreak.

Naturally, authorities are keeping a close eye on things. ICMR is coordinating Zika surveillance in the country and Hyderabad-based Bharat Biotech is leading one of the vaccine development projects, confirmed ICMR director general Soumya Swaminathan. This imminent threat is what motivated Faiq and group to venture into the busy field of Zika research.

It starts with Vitamin A

Fruits and vegetables are good sources of Vitamin A, chemically known as retinol. The retinol consumed by a pregnant woman plays an important role in the development of the embryo. Inside the embryonic cell, the retinol is chemically converted to retinoic acid which is then transported into the nucleus. Inside the nucleus, retinoic acid binds at regions on the DNA called RARE sequences. This binding sets off a cascade of reactions resulting in the regulated release of molecules crucial in the formation of the primitive brain. Disruption of this pathway can cause abnormalities in the embryo, a condition called retinoic acid embryopathy.

Underdeveloped brains are of one the abnormalities, though Jonaki Sen, a neurobiologist who studies this pathway in IIT Kanpur, pointed out in an email interview that very few cases have been observed in humans. “Since retinoic acid is required for development of nearly every tissue in the embryo, I suspect that any mutation affecting its signalling substantially is likely to lead to severe malformations and early lethality in the embryo and hence are not detected,” she said, explaining why this could be so.

Among the observed symptoms of retinoic acid embryopathy include underdeveloped cerebellums, brain stems or brain lobes, enlarged lateral ventricles and a reduced number of folds in the brain. What was interesting to Faiq and team was that these symptoms are also seen in the Zika-linked microcephaly cases. Faiq agrees that microcephaly is a non-specific symptom that is a common feature seen in many disorders, but the fact that Zika virus has been found in the brain tissues of microcephalic foetuses where other causes have been ruled out suggested that there might be more to this link. Hence, the team decided to design a study to figure out if this was more than a coincidence.

When a virus infects a host, it inserts parts of its genetic material into the genome of the host cell. In cases where the viral inserts are similar to the important regions found in the host genes, the working of the cell can be disturbed. Molecules in the host cell mistake the viral genes for the host genes. The researchers wondered if something similar is happening when Zika virus infects an embryo’s neurons. So they scanned the genome sequences of different types of Zika viruses for the presence of RARE sequences using a computer programme. Their suspicion proved right. It turned out that the Zika genome contains RARE sequences, too.

“If Zika virus genome contains significant numbers of RARE sequence, it may insert additional amount of RARE sequences in genomes of the developing brain cells of the foetus. It is well known that excess amount of retinoic acid is can interfere with normal brain development. This manifests as microcephaly,” Faiq hypothesised.

It fit like a glove

More intriguingly, the scientists observed that the number of RARE sequence matches found in the Zika genomes was directly proportional to the virulence of that particular strain, i.e. the strains that were more likely to cause microcephaly had more RARE sequences. Faiq admits that they found this level of correlation surprising. “The direct genomic evidence fit like a glove, indicating that our speculation – that retinoic acid metabolism seems to be a key factor in Zika virus mediated microcephaly – was correct.”

To confirm further, the team also scanned the genomes of other viruses for the RARE sequences. Only those viruses that affected the nervous system, for example Chikungunya virus, were found to have the sequences. But even these viruses, the authors warn need to be able to cross from mother to foetus to be able to affect brain development. So viruses like Japanese encephalitis virus, though contain RARE sequences and affect brain cells, have not been known to cause microcephaly – possibly because maternal-foetal transmission of it has not yet been documented in humans. One exception was dengue virus, which showed no matches despite evidence of it affecting the nervous system in literature.

Sen, who is one of the few Indian scientists studying the impact of retinoic acid signalling on brain development in vertebrates, cautioned that further evidence of this mechanism is necessary. She said that directed experiments using brain organoids derived from human embryonic stem cells can be used as a model for this, though she admits that this may not be feasible at this time.

Faiq agrees that further studies must follow. He suggested that brain cell cultures and animal models will be helpful in proving our hypothesis. “This would involve infection with Zika virus and evaluation of retinoic acid metabolism and parallel monitoring of brain development.”

Uncovering the actual mechanism of the Zika-microcephaly link is our best bet at formulating a way to treat this condition. If, in the future, Faiq and team’s mechanism does prove to be valid then the retinoic acid pathway could be targeted. “A means could be designed to ensure the optimum metabolism of retinoic acid in developing brain,” said Faiq. He predicted that drugs can be designed to specifically target the pathway and prevent developmental defects like microcephaly.