Antibodies to dengue virus enhance infection by Zika virus

Zika virus

Model of Zika virus particle. E glycoprotein dimer is expanded at left.

It has been speculated that the development of neurological disease and fetal abnormalities after Zika virus infection may be due to the presence of  antibodies against other flaviruses that enhance disease. In support of this hypothesis, it has been shown that antibodies to dengue virus enhance infection of cells by Zika virus.

There are four serotypes of dengue virus, and infection with one of the serotypes generally leads to a self-resolving disease. When a different serotype is encountered, antibodies to the first serotype bind virus but do not block infection. Dengue virus then enters and replicates in cells that it does not normally infect, such as macrophages. Entry occurs when Fc receptors on the cell surface bind antibody that is attached to virus particles. The result is higher levels of virus replication and more severe disease. This phenomenon is called antibody-dependent enhancement, or ADE.

Because dengue and Zika viruses are antigenically related, an important question is whether antibodies to dengue virus can enhance infection with Zika virus. To answer this question, the authors used two broadly neutralizing anti-dengue virus monoclonal antibodies that had been previously isolated from patients who recovered from infection. These antibodies recognize the viral E glycoprotein; specifially, a loop of the protein involved in fusion of the viral and cell membranes. The amino acid sequence of this fusion loop region is the same in dengue virus and in Zika virus.

The two anti-dengue virus mAbs bind the Zika virus E glycoprotein and also recognize Zika virus infected cells. However, when mixed with Zika virus, they do not neutralize infectivity in a cell line made from rhesus monkey kidneys. But when the antibodies were tested in Fc receptor-bearing cells, Zika virus infection was enhanced by over 100 fold. In the absence of dengue virus antibodies, levels of Zika virus RNA are very low.

Serum from four patients who had recovered from dengue virus infection was also examined for enhancement of Zika virus infection. All four sera contain antibodies that neutralized all four serotypes of dengue virus, but only two neutralized Zika virus infection. All four human sera enhanced Zika virus infection of Fc receptor-bearing cells. Enhancement of Zika virus infection could be blocked when Fc receptors were blocked with anti-Fc receptor antibodies before virus infection. A control serum from a patient in Canada that did not contain antibodies to dengue or Zika viruses did not enhance Zika virus infection.

These findings have one caveat: enhancement of Zika virus infection by antibodies against dengue virus was measured by PCR amplification of infected cell RNA, not by measuring the yield of infectious virus. The assumption is that increased intracellular viral RNA means more virus released from the cell, but this remains to be confirmed.

It will be important to confirm these findings in animal models of Zika virus infection, and in humans. If true, they have wide implications. If antibodies against dengue virus enhance Zika virus infection in humans, more severe disease might be observed in areas such as Brazil where both viruses co-circulate. It will be necessary to determine if Guillain-Barré syndrome, other neurological complications, and birth defects correlate with antibodies to dengue virus. Perhaps Fc receptors on the placenta and neural tissues allow entry of Zika virus only when bound to dengue virus antibody. It is also possible that antibodies to Zika virus might enhance dengue virus disease.

These observations do not bode well for Dengvaxia, a tetravalent dengue virus vaccine that has been recently licensed in Brazil, Mexico, and the Philippines. Might anti-dengue virus antibodies induced by this vaccine make Zika virus disease more severe? This outcome would be a tragedy, as many years of work has gone into making this vaccine to prevent severe disease caused by dengue virus infections. Second generation dengue virus vaccines such as TV003 are already moving through clinical trials.

It is essential to determine as soon as possible if antibodies induced by Dengvaxia and TV003 enhance Zika virus disease. If so, it will be necessary to assess whether deployment of this vaccine should proceed.

Dengvaxia consists of the yellow fever virus vaccine strain 17D in which the E and prM viral membrane proteins are substituted with those of dengue virus. In contrast, the attenuated TV003 vaccine has only the dengue virus genome. Would a vaccine consisting of TV003 plus an attenuated Zika virus vaccine solve potential problems of antibody dependent enhancement of disease?

Update 4/28/16: Over on Twitter someone asked, “any idea why we DON’T see severe disease in parts of Africa/Asia where dengue and Zika co-circulate”? Good question, too long an answer for Twitter. The easiest is that in humans, there is no antibody depencent enhancement of Zika virus infection by dengue antibodies. But if there is ADE, then there are a number of possible explanations. First, there have not been enough cases in Africa, nowhere near the numbers in the Pacific and South America.

There were certainly some cases of Guillain-Barré syndrome associated with some of the Pacific outbreaks – which I would consider more serious disease and could be potentiated by dengue antibodies.

I also think that Brazil is hyperendemic for dengue virus, with multiple serotypes circulating and people having multiple infections.

But the recent outbreaks are much larger than before, and dengue antibody induced complications of Zika virus infection might only be observed in larger outbreaks.

The authors of the paper discussed in this post suggest that the introduction of Zika virus into a completely naive population could also be a factor, as the age of exposure. Maybe a robust anti-Zika virus antibody response, in a non-naive population, can temper any effects of dengue mediated ADE.


TWiV 376: The flavi of the week is Zika

TWiVOn episode #376 of the science show This Week in Virology, the TWiV team discusses the latest data on Zika virus, including ocular defects in infants with microcephaly, and isolation of the entire viral genome from fetal brain tissue.

You can find TWiV #376 at

Zika virus

FlavivirusThe rapid spread of Zika virus through the Americas, together with the association of infection with microcephaly and Guillain-Barré syndrome, have propelled this previously ignored virus into the limelight. What is this virus and where did it come from?

Zika virus was first identified in 1947 in a sentinel monkey that was being used to monitor for the presence of yellow fever virus in the Zika Forest of Uganda. At this time cell lines were not available for studying viruses, so serum from the febrile monkey was inoculated intracerebrally into mice. All the mice became sick, and the virus isolated from their brains was called Zika virus. The same virus was subsequently isolated from Aedes africanus mosquitoes in the Zika forest.

Serological studies done in the 1950s showed that humans carried antibodies against Zika virus, and the virus was isolated from humans in Nigeria in 1968. Subsequent serological studies revealed evidence of infection in other African countries, including Uganda, Tanzania, Egypt, Central African Republic, Sierra Leone, and Gabon, as well as Asia (India, Malaysia, Philippines, Thailand, Vietnam, Indonesia).

Zika virus moved outside of Africa and Asia in 2007 and 2013 with outbreaks in Yap Island and French Polynesia, respectively. The first cases in the Americas were detected in Brazil in May 2015. The virus circulating in Brazil is an Asian genotype, possibly imported during the World Cup of 2014. As of this writing Zika virus has spread to 23 countries in the Americas.

The virus
Zika virus is a member of the flavivirus family, which also includes yellow fever virus, dengue virus, Japanese encephalitis virus, and West Nile virus. The genome is a ~10.8 kilobase, positive strand RNA enclosed in a capsid and surrounded by a membrane (illustrated; image copyright ASM Press, 2015). The envelope (E) glycoprotein, embedded in the membrane, allows attachment of the virus particle to the host cell receptor to initiate infection. As for other flaviviruses, antibodies against the E glycoprotein are likely important for protection against infection.

Zika virus is transmitted among humans by mosquito bites. The virus has been found in various mosquitoes of the Aedes genus, including Aedes africanus, Aedes apicoargenteus, Aedes leuteocephalus, Aedes aegypti, Aedes vitattus, and Aedes furcifer. Aedes albopictus was identified as the primary vector for Zika virus transmission in the Gabon outbreak of 2007. Whether there are non-human reservoirs for Zika virus has not been established.

Signs and Symptoms
Most individuals infected with Zika virus experience mild or no symptoms. About 25% of infected people develop symptoms 2-10 days after infection, including rash, fever, joint pain, red eyes, and headache. Recovery is usually complete and fatalities are rare.

Two conditions associated with Zika virus infection have made the outbreak potentially more serious. The first is development of Guillain-Barré syndrome, which is progressive muscle weakness due to damage of the peripheral nervous system. The association of Guillain-Barré was first noted in French Polynesia during a 2013 outbreak.

Congenital microcephaly has been associated with Zika virus infection in Brazil. While there are other causes of microcephaly, there has been a surge in the number of cases during the Zika virus outbreak in that country. Whether or not Zika virus infection is responsible for this birth defect is not known. One report has questioned the surge in microcephaly, suggesting that it is largely attributed to an ‘awareness’ effect.  Current epidemiological data are insufficient to prove a link of microcephaly with Zika virus infection. Needed are studies in which pregnant women are monitored to see if Zika virus infection leads to microcephaly.

Given the serious nature of Guillain-Barré and microcephaly, it is prudent for pregnant women to either avoid travel to areas that are endemic for Zika virus infection, or to take measures to reduce exposure to mosquitoes.

There are currently no antiviral drugs or vaccines that can be used to treat or prevent infection with Zika virus. We do have a safe and effective vaccine against another flavivirus, yellow fever virus. Substituting the gene encoding the yellow fever E glycoprotein with that from Zika virus might be a good approach to quickly making a Zika vaccine. However testing of such a vaccine candidate might require several years.

Mosquito control is the only option for restricting Zika virus infection. Measures such as wearing clothes that cover much of the body, sleeping under a bed net, and making sure that breeding sites for mosquitoes (standing water in pots and used tires) are eliminated are examples. Reducing mosquito populations with insecticides may also help to reduce the risk of infection.

Closing thoughts
It is not surprising that Zika virus has spread extensively throughout the Americas. This area not only harbors mosquito species that can transmit the virus, but there is little population immunity to infection. Infections are likely to continue in these areas, hence it is important to determine whether or not Zika virus infection has serious consequences.

Recently Zika virus was identified in multiple states, including Texas, New York, and New Jersey, in international travelers returning to the US . Such isolations are likely to continue as long as infections occur elsewhere. Whether or not the virus becomes established in the US is a matter of conjecture. West Nile virus, which is spread by culecine mosquitoes, entered the US in 1999 and rapidly spread across the country. In contrast, Dengue virus, which is spread by Aedes mosquitoes, has not become endemic in the US.

We recently discussed Zika virus on episode #368 of the science show This Week in Virology. You can be sure that we will revisit this topic very soon.

Added 1/28/16 9:30 PM: The letter below to TWiV provides more detail on the situation in Brazil.

Esper writes:

Hi TWIVomics

I hope this email finds you all well and free of pathogenic viruses.

My name is Esper Kallas, an ID specialist and Professor at the Division of Clinical Immunology and Allergy, University of São Paulo, Brazil.

I have been addicted to TWIV since a friend from U. Wisconsin participated in the GBV-C episode (David O’Connor, episode #260). Since then, never missed one episode. After long silent listening, I decided to write for the first time, motivated by the ongoing events in my country, potentially related to the Zika virus.

In the last episode, Emma wrote about events taking place in the small town of Itapetim, State of Pernambuco, Northeastern Brazil, which I will describe a bit later in this email. Before, let me bring some background information on the current situation.

Most believed Zika was a largely benign virus, causing a self-limited disease, clearly described in episode #368. Its circulation was documented after an outbreak became noticed in the State of Bahia (NE Brazil) by a group led by Guilherme Ribeiro, a talented young Infectious Diseases Scientist from Fiocruz (PMID: 26584464, Emerg Infect Dis. 2015 Dec;21(12):2274-6, free access)

However, things started to get awkward around October 2015, when a single hospital in Recife (NE Brazil) and some other practicing Obstetricians and Pediatricians from the region started reporting a mounting number of microcephaly cases in newborns, later confirmed by the national registry of newborns. The numbers are astonishing. The graph below depicts the number of cases per year prior to the surge in 2015. Only this year, 2,975 cases were reported by December 26, the vast majority in the second semester of the year. Cases are concentrated in the Northeast (map), with 2,608 cases, including 40 stillbirths or short living newborns.

Microcephaly, Brazil

In response to the situation, the Brazilian Ministry of Health has declared a national public health emergency (

The Brazilian Ministry of Health has been presenting updates every week (see link: It is important to observe some imperfections in these numbers: 1. There may be an over reporting after the news made to the big media, suggesting an association between microcephaly and Zika virus. 2. The criterion to consider a microcephaly case has been changed after the current epidemic from 33cm to 32cm; this is because 33cm of head circumference is sitting in the 10th percentile of newborns at 40 weeks of pregnancy and the adjustment would bring the limit to the 3rd percentile, increasing the specificity to detect a true microcephaly case (this may result in an over reporting in the beginning of the epidemic).

The association between Zika virus infection and microcephaly was suspected since the beginning, when Brazilian health authorities ruled out other potential causes, together with the fact that the microcephaly epidemic followed Zika virus spread. Further evidences were the two positive RT-PCR for Zika RNA in two amniotic fluids obtained from two pregnancies of microcephalic fetuses and a stillborn microcephaly case with positive tissues for Zika RNA. In fact, French Polynesia went back to their records and also noticed an increase of microcephaly case reporting, following their epidemic by the same virus strain in 2013 and 2014.

Now, Zika virus transmission has been detected in several countries in the Americas (

Although strong epidemiological data suggest the association between Zika virus and the microcephaly epidemic, a causal link between the virus and the disease is still lacking and is limited to few case reports. Many questions still remain. Does the virus damage embryonic neural tissue? What is the percentage of fetuses getting infected when the mother acquires Zika virus during pregnancy? Does the stage of pregnancy interfere with virus ability to be transmitted to the fetus and the development of neurologic effects? Are there other neurological defects related to Zika virus infection? Is there another cofactor involved, such as malnutrition or other concurrent infection? All these questions are exceedingly important to provide counseling to pregnant women and those who are planning to become pregnant, especially in Northeastern Brazil. In fact, Brazilian authorities have been recommending avoiding pregnancy until this situation is further clarified.

The microcephaly epidemic impact is unimaginable. It is a tragedy. These children are compromised for life and the impact on their families is beyond any prediction.

Back to the story sent by Emma. A small town in the North of Pernambuco State, named Itapetim, has almost 14 thousand inhabitants and has reported 11 cases of microcephaly in the past 3 months. This very same town has been suffering from a prolonged drought, since September 2013 when the last reservoir went dry. Perhaps the storage of clean water or the limited resources has led to the best environment for arbovirus spread and the development of microcephaly.

But the Zika virus’s impact may be reaching further. An increase in Guillain-Barré syndrome cases has also been noticed in the Northeast of Brazil, possibly related to the epidemic.

Several groups have been trying to establish animal models to study the interaction of Zika virus with neural tissue. The forthcoming developments are critical to better understand the virus immunopathology and confirm (or refute) the association between the virus infection and neurologic damage in fetuses and in the infected host developing Guillain-Barré syndrome. Many things still shrouded in mystery.

Keep on the good work. I will keep on listening!


TWiV 212: Apocalypse TWiV 122112 212

On episode #212 of the science show This Week in Virology, the TWiVerers answer listener email about genetically modified chickens, a hendra vaccine for horses, online education, curing color blindness, Roosevelt and polio, Th cells, and much more.

You can find TWiV #212 at

Safety of influenza 2009 H1N1 vaccine

whoA number of readers have asked when we would have information about the safety of the influenza 2009 H1N1 vaccine. The World Health Organization has just released briefing note #16 “Safety of pandemic vaccines” in which they summarize vaccination information from 16 countries in which 80 million doses of vaccine have been administered.

Side effects commonly reported include swelling, redness, or pain at the injection site, which usually resolve soon after vaccination. Fever, headache, fatigue, muscle aches, and a variety of allergic reactions, occurring shortly after vaccine administration, have also been reported less frequently.

There have been fewer than ten cases of Guillain-Barre syndrome reported in H1N1 vaccine recipients. These numbers are consistent with normal background rates of the illness.  All the individuals have recovered.

Some deaths have been reported in people who have been vaccinated. These are all investigated by WHO, and so far in no case is there a direct link to H1N1 vaccine as the cause of death.

There have been no differences in the safety profile of inactivated vaccines with or without adjuvant, and infectious attenuated vaccines..

WHO concludes:

Although intense monitoring of vaccine safety continues, all data compiled to date indicate that pandemic vaccines match the excellent safety profile of seasonal influenza vaccines, which have been used for more than 60 years.


twiv-200Hosts: Vincent Racaniello and Jason Rodriguez

On episode #50 of the podcast “This Week in Virology”, Vincent and Jason review influenza 2009 H1N1 vaccine trials and protection against the virus conferred by the 1976 swine flu vaccine, then move on to a virus called XMRV and its possible role in prostate cancer.

Click the arrow above to play, or right-click to download TWiV #50 (54 MB .mp3, 74 minutes)

Subscribe to TWiV in iTunes, by the RSS feed, or by email

Links for this episode:
One dose of influenza 2009 H1N1 vaccine without adjuvant is enough
Partially completed study on influenza 2009 H1N1 vaccine with MF59 adjuvant
1976 swine flu vaccine induces cross-reactive antibodies against influenza 2009 H1N1 strain
Explanation of hemagglutination-inhibition and microneutralization assays
FDA approves influenza 2009 H1N1 vaccine
XMRV is present in malignant prostatic epithelium and is associated with prostate cancer
Identification of a novel gammaretrovirus in prostate tumors
CDC page on Guillain-Barré syndrome

Weekly Science Picks
Jason Glass Microbiology
Vincent FluWeb Influenza Historical Resources Database

Send your virology questions and comments (email or mp3 file) to or leave voicemail at Skype: twivpodcast. You can also send articles that you would like us to discuss to delicious and tagging them with to:twivpodcast.

TWiV 43: Virus classification

twiv-200Hosts: Vincent Racaniello and Dick Despommier

In episode 43 of the podcast “This Week in Virology”, Vincent and Dick explain virus classification, and revisit the 1976 swine flu immunization campaign and Guillain-Barré syndrome.

Click the arrow above to play, or right-click to download TWiV #43 (32 MB .mp3, 46 minutes)

Subscribe to TWiV in iTunes, by the RSS feed, or by email

Links for this episode:
International Committee on the Taxonomy of Viruses (ICTV)
CBS 60 Minutes on 1976 swine flu vaccine (thanks Gus and Swiss compass!)
Vaccines and Guillain-Barré
NINDS Guillain-Barré Information
Reflections on the 1976 swine flu vaccination program
Swine influenza outbreak at Fort Dix, 1976

Weekly Science Picks
Dick Five Kingdoms: An Illustrated Guide to the Phyla of Life on Earth by Lynn Margulis and Karlene V. Schwartz
Vincent A Genetic Switch by Mark Ptashne

Send your virology questions and comments (email or mp3 file) to or leave voicemail at Skype: twivpodcast

Swine flu at Fort Dix

swineThe death of a dozen pigs from swine influenza last week in the Philippines reminded me of an incident at Fort Dix, NJ in 1976. The infection of humans with a strain of swine influenza lead to a nationwide immunization campaign to curb a pandemic that never occurred.

An explosive outbreak of febrile respiratory disease raced through the 19,000 personnel at Fort Dix in January 1976. Virological laboratory studies revealed the presence of a new swine influenza strain which was named A/New Jersey/76 (Hsw1N1). The virus infected 230 soldiers and caused severe respiratory disease in 13, including one death.

At the time it was believed that a swine virus had caused the 1918-19 influenza pandemic. Therefore scientists were concerned that the virus had returned to Fort Dix and would soon cause another catastrophic outbreak. Dr. Edwin Kilbourne, a noted influenza researcher, and others convinced the US Public Health Service to contract for the production of 150 million doses of vaccine. In March of 1976 President Gerald Ford announced a program to inoculate every man, woman and child in the United States against swine flu. Immunizations began in October, but only 45 million doses had been distributed when the program was halted in December. By then it was clear that A/New Jersey/76 was going nowhere. An unfortunate consequence was that many individuals developed Guillain-Barré syndrome, a neurological disease involving muscle weakness, paralysis, and sometimes death.

Why didn’t A/New Jersey/76 spread to the general population? One factor was the limited contact between basic trainees and others who more frequently travel outside the facility. Older personnel may have been immune, because military influenza vaccine formulations from 1955 through 1969 contained a swine influenza component. Competition with concurrent circulating influenza virus strain, A/Victoria, might have  limited the impact of A/New Jersey virus which is believed to transmit poorly among humans.

In retrospect, the swine flu program had many flaws. The vaccine should have been stockpiled until it was clear that an epidemic was taking place. Today we realize that the 1918 influenza virus is derived from an avian strain, not a swine strain – had this information been available in 1976, the immunization campaign would not have taken place. Presumably these and many other errors will not be repeated when the time comes to immunize against the next pandemic strain.

To this day the origin of A/New Jersey/76 virus is an enigma. One theory is that a swine virus was brought to Fort Dix early in 1976 as recruits returned after the holidays. However, none of the personnel who were interviewed admitted to having contact with pigs. The virus seems to have circulated at Fort Dix for about a month, then disappeared.