Zika Fear Heats Up, Cure Sought in Cold

Dr. Michael Rossmann (right) and Dr. Richard Kuhn stand with the cryo-electron microscope used to determine the structure of the Zika virus. Image: Purdue University, Mark Simons

Dr. Michael Rossmann (right) and Dr. Richard Kuhn stand with the cryo-electron microscope used to determine the structure of the Zika virus. Image: Purdue University, Mark Simons

North America is welcoming summer and with it the increased fear of a pronounced expansion of Zika from its southern neighbors. In April, the US Centers for Disease Control and Prevention (CDC) held a summit to formulate an action plan against the virus and shortly thereafter began issuing statements, including steps to prevent occupational exposure and a controversial proposal that asked women to postpone planned pregnancies.

No vaccine currently exists to prevent the spread of Zika, but scientists at facilities across the world are busy studying the virus, many of them with cryogenic systems.

A team led by Purdue University researchers using cryo-electron microscopy, for example, was the first to determine the structure of the Zika, revealing insights critical to the development of effective antiviral treatments and vaccines.

The team identified regions within the virus structure where it differs from other flaviviruses, the family of viruses—including dengue, West Nile, yellow fever, Japanese encephalitis and tick-borne encephalitic viruses—to which Zika belongs.

Any regions within the virus structure unique to Zika have the potential to explain differences in how the virus is transmitted and how it manifests as a disease, according to Dr. Richard Kuhn, director of the Purdue Institute for Inflammation, Immunology and Infectious Diseases (PI4D). Kuhn led the research team with Dr. Michael Rossmann, Purdue’s Hanley Distinguished Professor of Biological Sciences. The two have studied flaviviruses for more than 14 years and were the first to map the structure of any flavivirus when they determined the dengue virus structure in 2002. In 2003, they were first to determine the structure of West Nile and now they are the first to do so with Zika.

“The structure of the virus provides a map that shows potential regions of the virus that could be targeted by a therapeutic treatment, used to create an effective vaccine or to improve our ability to diagnose and distinguish Zika infection from that of other related viruses,” says Kuhn. “Determining the structure greatly advances our understanding of Zika, a virus about which little is known. It illuminates the most promising areas for further testing and research to combat infection.”

The team studied a strain of Zika virus isolated from a patient infected during an epidemic in French Polynesia and determined the structure to be 3.8Å. At this near-atomic resolution, key features of the virus structure can be seen and groups of atoms that form specific chemical entities, such as those that represent one of 20 naturally occurring amino acids, can be recognized, according to Rossmann.

“We were able to determine through cryo-electron microscopy the virus structure at a resolution that previously would only have been possible through X-ray crystallography,” Rossmann says. “Since the 1950s X-ray crystallography has been the standard method for determining the structure of viruses, but it requires a relatively large amount of virus, which isn’t always available; it can be very difficult to do, especially for viruses like Zika that have a lipid membrane and don’t organize accurately in a crystal; and it takes a long time. Now, we can do it through electron microscopy and view the virus in a more native state. This was unthinkable only a few years ago.”

The team found the structure to be very similar to that of other flaviviruses with an RNA genome surrounded by a lipid, or fatty, membrane inside an icosahedral protein shell. The strong similarity with other flaviviruses was not surprising, according to Purdue graduate student Devika Siroh, and is perhaps reassuring in terms of vaccine development already underway.

”Most viruses don’t invade the nervous system or the developing fetus due to blood-brain and placental barriers, but the association with improper brain development in fetuses suggest Zika does,” Sirohi says “It is not clear how Zika gains access to these cells and infects them, but these areas of structural difference may be involved. These unique areas may be crucial and warrant further investigation.”

The team also discovered differences between Zika and other flaviviruses, including a glycosylation site that protrudes from the surface of the virus. A carbohydrate molecule consisting of various sugars is attached to the viral protein surface at this site, and could be linked to viral infection and transmission.

In many other viruses it has been shown that as a virus projects a glycosylation site outward, an attachment receptor molecule on the surface of a human cell recognizes the sugars and binds to them, according to Kuhn.

“If this site functions as it does in dengue and is involved in attachment to human cells, it could be a good spot to target an antiviral compound,” Rossmann says. “If this is the case, perhaps an inhibitor could be designed to block this function and keep the virus from attaching to and infecting human cells.”

In the majority of infected individuals symptoms are mild and include fever, skin rashes and flulike illness, according to the World Health Organization (WHO), but Zika virus has been associated with both Guillain-Barré syndrome, an autoimmune disease that can lead to temporary paralysis, and microcephaly, a birth defect that causes brain damage and an abnormally small head in babies born to mothers infected during pregnancy.

Researchers at Florida State University, Johns Hopkins University and Emory University were the first to associate Zika’s link to microcephaly with more than anecdotal evidence. The team infected neural stem cells, stored them at cryogenic temperatures and then began monitoring them for changes.

Prof. Tang and graduate student Sarah Ogden in a lab at FSU. Image: Florida State University

Prof. Tang and graduate student Sarah Ogden in a lab at FSU. Image: Florida State University

“We’re trying to fill the knowledge gap between infection and the neurological defects,” says Dr. Hengli Tang, a professor of biological science at FSU. “This research is the very first step in that, but it’s answering a critical question. It enables us to focus the research. Now you can be studying the virus in the right cell type, screening your drugs on the right cell type and studying the biology of the right cell type.”

The team discovered that the virus targets a cell type called human embryonic cortical neural progenitors in as little as three days after being exposed to the virus. The researchers also discovered that these infected cells replicate the Zika virus, posing potential treatment problems, and that the virus is directly interfering with cell growth and function. Some of the cells died after being infected.

“It’s significant because we’re literally the first people in the world to know this, to know that this virus can infect these very important cells and interfere with their function,” Tang says. “Research is rewarding in general, but when you have something this timely and this clinically relevant, it’s extra satisfying because we’ll be helping people in the long run.”

Since 2015, 46 countries have reported new outbreaks of Zika, while 14 others have had ongoing outbreaks since 2007. Of the countries where Zika is circulating, 13 have reported an increased incidence of Guillain-Barré syndrome while eight countries or territories have reported an increase in microcephaly, according to WHO. In February WHO declared the Zika virus to be “a public health emergency of international concern.”

*Update August 29, 2016. The team of researchers from Florida State University, Johns Hopkins University and the National Institutes of Health has found existing drug compounds that can both stop Zika from replicating in the body and from damaging the crucial fetal brain cells that lead to birth defects in newborns.