TWiV 415: Ebola pipettors and the philosopher’s clone

Jeremy Luban, Aaron Lin, and Ted Diehl join the TWiV team to discuss their work on identifying a single amino acid change in the Ebola virus glycoprotein from the West African outbreak that increases infectivity in human cells.

You can find TWiV #415 at microbe.tv/twiv, or listen below.

Click arrow to play
Download TWiV 415 (67 MB .mp3, 110 min)
Subscribe (free): iTunesRSSemail

Become a patron of TWiV!

Increased infectivity of Ebola virus glycoprotein from West Africa

filovirionWhen viruses cross species, serial transmission may lead to the selection for mutations that confer improved replication or transmission in the new host. Identifying such mutations in human viruses is extremely difficult: we cannot conduct the appropriate experiments in humans, and often do not have viral isolates spanning the time from spillover through prolonged circulation. The 2013-2016 outbreak of Ebola virus in West Africa is unique because viral genome sequences were obtained early and throughout the epidemic. The results of two new studies (link to paper one, link to paper two) suggest that some of the observed mutations increase infectivity for human cells. The impact of these mutations on infection of humans, and their role in the West African outbreak, remain unknown.

Many mutations have been identified among the many hundreds of genome sequences obtained during the recent Ebola virus epidemic. One stands out: a mutation that leads to a single amino acid change in the viral glycoprotein, from alanine to valine at position 82 (A82V). This change arose early in the outbreak (it was first observed in Guinea in March 2014) and was subsequently found in most of the isolates. It has never been observed in previous Ebolavirus outbreaks.

The effect of the A82V change on viral infectivity was determined by building pseudotyped viral particles – in this case, HIV particles with the Ebola virus glycoprotein. Human cells in culture were infected with pseudotyped viruses with the Ebola virus glycoprotein with either alanine or valine at position 82. Infectivity was measured by quantifying the production of a protein from the HIV genome. The results show that A82V increases infectivity by twofold. The effect is also observed in cells from non-human primates, but not from rodents, dogs, or cats. However, the A82V change decreased infectivity in bat cells.

The A82V change is located at the binding site of the Ebola virus glycoprotein with the cell fusion receptor, NPC1. It appears to increase the fusion activity of the viral glycoprotein.

Other amino acid changes in the Ebola virus glycoprotein were also observed to increase infectivity in human cells, and decrease infectivity in bat cells.

The pattern of increased infectivity in primate cells, and decreased infectivity in bats, is consistent with the hypothesis that the outbreak virus came from bats, and after circulation of the virus in humans, it lost some ability to infect bat cells while becoming better at infecting human cells. However there is still no solid proof that bats are a reservoir of Ebolaviruses.

What does increased infectivity have to do with infection of humans? The idea is that the mutation increases the efficiency of virus entry into cells, and hence increased viral gene expression is observed. Fewer viruses needed to infect a cell, the better chance of initiating an infection. But is the two-fold increase observed in cells enough to impact infection in humans?

The assays used in these papers measure protein production from an HIV genome. The experiments need to be repeated using bona fide Ebola virus, to make sure that the mutations have the same effect. The changes might have impacts on other stages of viral replication. Furthermore, the impact of the changes in the viral glycoprotein should be assessed in animal models, to determine if improved infectivity has any impact on pathogenesis and transmission. Ultimately, we can’t prove that these mutations have any effect in humans – the needed experiments cannot be done.

I’m curious about why the A82V change was not seen in previous Ebola virus outbreaks. Those were in different parts of Africa – could the changes be driven by population genetics, ecology, or other factors? It will be important to determine if the same change is selected in future outbreaks.

The authors are sufficiently cautious in their conclusions. From paper #2:

Despite the experimental data provided here, it is impossible to clearly establish whether the adaptive mutations observed were in part responsible for the extended duration of the 2013–2016 epidemic. Indeed, it seems likely that the prolonged nature of the outbreak in West Africa was primarily due to epide- miological factors, such as an increased circulation in urban areas that in turn led to larger chains of transmission.

From paper #1:

Our findings raise the possibility that this mutation contributed directly to greater transmission and thus to the severity of the outbreak. It is difficult to draw any conclusions from this hypothesis, though…

As I feared, press coverage of these findings has been inaccurate. For example, a BBC headline proclaims “Ebola adapted to easily infect people”. Even the journal Cell, which published both papers, made an incorrect conlcusion: see the screen capture below from the journal website.key mutations ebola virusBoth Cell and the BBC might have taken too literally the unfortunate title of one of the papers,  “Human adaptation of Ebola virus during the West African Outbreak.” The results suggest adaptation to human cells, not to humans. The title of the second paper is sufficiently careful: “Ebola virus glycoprotein with increased infectivity dominated the 2013-2016 epidemic”. But that’s not a BBC headline.

TWiV 394: Cards in a hand

Vincent and Alan speak with Erica Ollmann Saphire about her career and her work on understanding the functions of proteins of Ebolaviruses, Marburg virus, and other hemorrhagic fever viruses, at ASM Microbe 2016 in Boston, MA.

You can find TWiV #394 at microbe.tv/twiv, or listen or watch the video below.

Click arrow to play
Download TWiV 394 (65 MB .mp3, 89 min)
Subscribe (free): iTunesRSSemailGoogle Play Music

Become a patron of TWiV!

TWiEVO 5: Looking at straw colored fruit bats through a straw

TWiEVOOn episode #5 of the science show This Week in Evolution, Sara Sawyer and Kartik Chandran join Nels and Vincent to talk about how the filovirus receptor NPC1 regulates Ebolavirus susceptibility in bats.

You can find TWiEVO #5 at microbe.tv/twievo, or you can listen below.

Click arrow to play
Download TWiEVO 5 (73 MB .mp3, 98 min)
Subscribe (free): iTunesRSSemail

TWiV 309: Ebola email

On episode #309 of the science show This Week in Virology, the TWiVocytes answer questions about Ebola virus, including mode of transmission, quarantine, incubation period, immunity, and much more.

You can find TWiV #309 at www.microbe.tv/twiv.

Ebola virus arrives in New York City

This morning I received this email from President Lee Bollinger:

Dear fellow members of the Columbia community:

As you may have seen in the media, Dr. Craig Spencer is being treated for Ebola at Bellevue Hospital in Manhattan. Dr. Spencer, an emergency department physician at NewYork-Presbyterian/Columbia University Medical Center, recently returned from a humanitarian mission with Doctors Without Borders to one of the outbreak areas in Western Africa. We admire and appreciate all of those willing to do this vital and selfless public health work around the globe.

It’s critical to bear in mind what our public health and infectious disease experts have emphasized – that the risk to people in New York City and at Columbia remains extremely low. If you or anyone has any concerns, please visit the University’s Ebola Preparedness site or the New York City Department of Health Ebola update page. You may also contact Student Health Services or Workforce Health and Safety for Faculty/Staff with Hospital Responsibilities.

We must keep Dr. Spencer in our thoughts and wish him a full and speedy recovery, as we do the vulnerable populations he serves. We will also continue to keep the Columbia community informed as we learn more from City, State, and Federal health officials.

Sincerely,
Lee C. Bollinger

The transition between incubation period (when there are no symptoms) and the first clinical signs is a dangerous period. During this time the patient may continue to move around in public despite having fever and other indications of infection. It will be important to trace as many of this physician’s contacts as possible, a difficult task in a city of over 8 million people. Apparently the physician traveled around the city, using the subways, the night before having a fever. Whether any virus is shed during this time, in amounts sufficient to infect others, is unknown, but could be determined by studying the contacts of such infected individuals.

TWiV 297: Ebola! Don’t panic

On episode #297 of the science show This Week in Virology, the TWiVites present an all-ebolavirus episode, tackling virology, epidemiology, and approaches to prevention and cure that are in the pipeline.

You can find TWiV #297 at www.microbe.tv/twiv.

Ebolavirus vaccines and antivirals

guinea-liberia-sierra-leone-2014As the epidemic of Zaire ebolavirus in Western Africa continues (1,779 cases and 961 deaths in four countries), many are questioning why there are no means of preventing or stopping infection. In the past two decades there has been substantial research into developing and testing active and passive vaccines and antiviral drugs, although none have yet been licensed for use in humans.

Using antibodies to treat infection with ebolaviruses with antibodies is probably the best known therapy, because it was used to treat a two Americans who were infected while working in Liberia. They received a mixture of three monoclonal antibodies (called ZMapp) which had been previously shown to block infection of cells with ebolaviruses, and prevent lethal infection of non-human primates when given within 24-48 hours after infection. These are mouse monoclonal antibodies that have been ‘humanized’ so that when given to people they do not induce an antibody response against the antibodies. Humanization involves changing the amino acids of the antibody molecule from mouse to human, except in the part of the antibody that binds antigen. The antibodies are then synthesized in tobacco plants and purified. Administering anti-viral antibodies to patients, also called passive immunization, was done long before vaccines were available. Serum from patients who had recovered from a particular disease would be given to others who had recently been infected, in order to prevent disease. Such therapy was used to save the life of virologist Jordi Casals, who had become infected with Lassa virus while isolating the virus from the blood of a patient, Penny Pinneo. The serum administered to Casals was obtained from Pinneo, who had recovered from the infection. The American doctor infected with Zaire ebolavirus while working in Liberia was also given serum from a boy who had recovered from infection.

As ZMapp has not yet been subjected to human clinical trials to determine its safety and efficacy, its use in an infected human is considered unusual. A phase I clinical trial needs to be done to ensure that the preparation of monoclonal antibodies is safe in humans. Determining whether monoclonal antibody therapy for ebolavirus infection is effective is more difficult. Such testing could only be done during an outbreak, during which it would not be ethical to withhold treatment from the control group. Nevertheless it is clear that such mixtures of monoclonal anti-viral antibodies could potentially save many lives during outbreaks.

While passive immunization has value in saving lives, its protection is temporary: the antibodies given to patients do not endure. A better approach is immunization, which not only induces anti-viral antibodies, but creates immune memory, so that subsequent infections are accompanied by another round of antibody production. The catch is that it takes about two weeks after immunization for antibodies to reach sufficient protective levels. Nevertheless, a vaccine would likely have had substantial impact on the current outbreak, which began in March 2014 and has continued for 5 months.

A number of experimental vaccines against ebolaviruses are in development. In one approach, the glycoprotein of vesicular stomatitis virus is replaced with the corresponding protein of different ebolaviruses. These vaccines protect non-human primates from lethal infection. A similar approach using an attenuated rabies virus to deliver the ebolavirus glycoprotein also protected non-human primates from infection, as did immunization with an adenovirus encoding the ebolavirus glycoprotein.  This vaccine candidate has been shown to be safe and immunogenic in phase I clinical trials. Another vaccine approach entails production of the ebolavirus glycoprotein in E. coli. Immunization of mice with the purified protein leads to the production of neutralizing antibodies. Because protein-based vaccines do not replicate, the immune response may need to be boosted by using an adjuvant that stimulates the innate immune system and leads to better antibody production. A double-stranded RNA adjuvant has been shown to augment the immune response against a non-infections, virus-like particle vaccine containing the Ebola virus glycoprotein but not the viral genome.

Antivirals certainly have a place in control of viral disease, and a number of promising candidates to control infection with ebolaviruses have been developed. One is a nucleoside analog which is incorporated into RNA by the viral RNA polymerase and leads to chain termination. It blocks replication of ebolaviruses in culture cells, and protects mice and nonhuman primates from lethal infection. This compound, called BCX4430, is a broad spectrum antiviral that inhibits the replication of not only members of the Filoviridae, but also Arenaviridae, Bunyaviridae, Orthomyxoviridae, Picornaviridae, Paramyxoviridae, Flaviviridae, Coronaviridae. Another inhibitor of viral RNA synthesis is favipiravir, which has the advantage of being in late stage clinical development for the treatment of influenza. This compound inhibits replication of ebolaviruses in cultured cells and reduces disease severity and mortality in a mouse model of disease.

It is likely that the extent of the current outbreak of Ebola virus disease, the largest to date, will provide impetus to move some of these treatments into human trials. But consider that all the research on active and passive vaccines and antivirals for ebolaviruses required work in BSL-4 laboratories. Those who call for the shuttering of BSl-4 laboratories need to take note and move away from their unrealistic and unreasonable position.

Zaire ebolavirus in West Africa

Dr. Tom Solomon is Director of the Institute for Infection and Global Health at the University of Liverpool. Here he speaks with Vincent Racaniello about the 2014 outbreak of Zaire ebolavirus in West Africa. Dr. Solomon discusses why the epidemic has spread, how it might be curtailed, the return of two infected health workers back to the United States for treatment, and the possibility that he might be traveling to the affected region to assist with medical care.

 
 

TWiV 283: No Reston for the weary

On episode #283 of the science show This Week in Virology, Jens Kuhn speaks with the TWiV team about filoviruses, including the recent Ebola virus outbreak in Guinea.

You can find TWiV #283 at www.microbe.tv/twiv.