TWiV 353: STING and the antiviral police

On episode #353 of the science show This Week in Virology, the TWiVniacs discuss twenty-eight years of poliovirus shedding by an immunodeficient patient, and packaging of the innate cytoplasmic signaling molecule cyclic GMP-AMP in virus particles.

You can find TWiV #353 at www.microbe.tv/twiv.

Oral polio vaccine-associated paralysis in a child despite previous immunization with inactivated virus

Poliovirus by Jason Roberts

Poliovirus by Jason Roberts

Vaccine-associated poliomyelitis caused by the oral poliovirus vaccine is rare, but its occurrence in a healthy, immunocompetent 6-month old child was highly unusual because the child had been previously immunized with two doses of the injected, inactivated poliovirus vaccine (IPV).

The three poliovirus vaccine strains developed by Albert Sabin (OPV, oral poliovirus vaccine) contain mutations which prevent them from causing paralytic disease. When the vaccine is ingested, the viruses replicate in the intestine, and immunity to infection develops. While replicating in the intestinal tract, the vaccine viruses undergo mutation, and OPV recipients excrete neurovirulent polioviruses. These so-called vaccine-derived polioviruses (VDPV) can cause poliomyelitis in the recipient of the vaccine or in a contact. During the years that the Sabin poliovirus vaccines were used in the US, cases of poliomyelitis caused by VDPV occurred at a rate of about 1 per 1.4 million vaccine doses, or 7-8 per year. Once the disease was eradicated from the US in 1979, the only cases of polio were caused by the Sabin vaccine.

To prevent vaccine-associated poliomyelitis, in 1997 the US switched to an immunization schedule consisting of two doses of IPV followed by one dose of OPV. The US then switched to using IPV exclusively in 2000. The child in this case essentially had a polio immunization course similar to that utilized in the US from 1997-2000: two doses of IPV, one dose of OPV. Why did the child develop poliomyelitis?

One clue comes from the fact that after the switch to an IPV-OPV schedule in 1997, there were still three cases of VAPP in 1998 and three in 1999. Another hint comes from a study of immune responses in children given multiple doses of IPV. Most of the children receiving two doses of IPV produced antibodies against types 1 and 2 poliovirus (92 and 94%), but only 74% of children produced antibodies against type 3 poliovirus.

The final piece of information needed to solve this puzzle is that the child in this case had vaccine-associated poliovirus caused by the type 3 strain, which was isolated from his feces.

Therefore, the child in this case most likely did not produce sufficient antibodies to type 3 poliovirus after receiving the two doses of IPV. As a consequence, when he was given OPV, he developed type 3 vaccine-associated poliomyelitis.

This case of VAPP could have been prevented: the child was born in Canada, and as customary in that country since 1995, he received two doses of IPV. At 5 months of age the child and his family visited China, where his parents decided to continue his immunizations according to the local schedule. China still uses OPV, so that is what the child received.

India has been free of polio for three years

Poliovirus cutaway

Image credit: Jason Roberts

Three years ago today, on 13 January 2011, the last case of poliomyelitis was reported in India. This achievement represents a remarkable turnaround for a country where control of the disease had for years been extremely difficult. As recently as 2009 there were 741 confirmed cases of polio caused by wild-type virus in India. Being polio-free for three years is certainly a cause for celebration, but not for becoming complacent. Immunization efforts in India must not decline, because wild-type and vaccine-derived polioviruses continue to circulate and pose a threat to any unimmunized individual.

Wild polioviruses – those that have always been circulating in nature – continue to cause disease in Afghanistan and Pakistan, two countries close to India. Pakistan reported 58 polio cases in 2012, and 85 so far in 2013; for Afghanistan the numbers are 37 and 12. But distant countries can also transmit polio: recent outbreaks in the Horn of Africa and in Syria originated in Nigeria and Pakistan, respectively.

Perhaps a greater threat are vaccine-derived polioviruses. The Sabin poliovirus vaccines, which have so far been the mainstay of the polio eradication effort, comprise infectious viruses that are taken orally. Upon replication in the intestinal tract, the vaccine strains confer immunity to infection, but they also revert and become capable of causing paralysis. Such vaccine-derived polioviruses circulate and can cause outbreaks of polio. Because India has been using Sabin poliovirus vaccines intensely for many years, there is no doubt that vaccine-derived polioviruses are circulating in that country. If polio vaccine coverage drops, there will be outbreaks of polio caused by vaccine-derived strains. Even if wild polioviruses disappeared from the globe, as long as Sabin vaccines are used, vaccine-derived polioviruses will circulate. The solution to this conundrum is to switch to Salk’s inactivated poliovirus vaccine and wait for the Sabin-derived strains to disappear. This switch is now part of the WHO’s eradication plan (it wasn’t always), but it will not be easy: Salk vaccine must be injected, and therefore requires trained health care personnel; administering Sabin vaccine requires no special skills. But we cannot simply stop immunizing with Sabin vaccine – that is a recipe for outbreaks of polio.

According to the World Health Organization, being free of wild polio for three years means that the virus is probably no longer endemic in India. However, WHO does not certify individual countries as polio-free; rather it declares a WHO region polio-free when all countries in the Region have not reported a case of wild polio for 3 years in the face of highly active surveillance. The Americas, the Western Pacific, and European regions have been declared polio-free by WHO. India is part of the South-East Asia region, which also includes Bangladesh, Bhutan, Democratic People’s Republic of Korea, Indonesia, Maldives, Myanmar, Nepal, Sri Lanka, Thailand, and Timor-Leste, none of which have reported a case of polio for 3 years. WHO will decide in March whether to declare this region polio-free. That would leave the regions of Africa and the Eastern Mediterranean as the last known reservoirs of wild poliovirus.

India polio-free for one year

Year in polio 2011A year has passed since the last reported case of poliomyelitis in India, which occurred on 13 January 2011 in a two year old girl in Howrah, West Bengal. If no additional cases are reported in the next few weeks (some samples are currently being tested for the virus), then it will mark the first time that India has been polio free for one year.

This achievement represents a remarkable turnaround for India, where control of the disease had for years been extremely difficult. As recently as 2009 there were 741 confirmed cases of polio caused by wild-type virus (as opposed to vaccine-derived virus) in India. The tide turned in 2010 with only 42 confirmed polio cases, and in calendar year 2011 there was just one. That is why the 2011 map marking locations of confirmed wild polio cases in India (see figure) shows only one red dot (paralysis caused by type 1 poliovirus) in the country. The blue dots indicate cases caused by type 3 poliovirus.

The challenge now is to keep India free of polio. The map shows why this will be difficult – there are many red dots (cases of type 1 polio) in neighboring Pakistan and Afghanistan. Poliovirus does not respect national borders – China had been free of polio since 1999, but now there are red dots in that country. That outbreak was imported from Pakistan. Even the polio cases in more distant countries such as Africa constitute a threat. As long as there is polio somewhere, all countries must maintain extensive immunization programs. Whether or not that will happen depends upon money, determination, and allowing immunization campaigns to proceed without interruption.

Once polio was eradicated from the United States, the only poliomyelitis was caused by the Sabin vaccine. Consequently this country switched to the use of inactivated vaccine in 2000. As other countries eliminate the disease, vaccine-associated poliomyelitis will become more prominent. If eradication of polio is achieved, the world will have to switch to using inactivated poliovaccine.

Related:

Wild poliovirus in China
Dreaming of inactivated poliovaccine
Poliomyelitis after a twelve year incubation period
Poliovirus vaccine litigation

 

Transgenic mice susceptible to poliovirus

pvr transgenic mouseYesterday I terminated the last remaining mice in my small colony, including the line of poliovirus receptor transgenic mice that we established here in 1990. Remarkably, I had never written about this animal model for poliomyelitis which has played an important role in the work done in my laboratory.

While I was still working on poliovirus as a postdoctoral fellow with David Baltimore, I became interested in how the virus causes disease. There were no convenient animal models to study poliovirus pathogenesis, so I began to think about the cellular receptor for the virus and how it could be used to make a mouse model for infection. When I moved to Columbia University Medical Center in 1982, I decided to identify the cellular gene for the poliovirus receptor. This work was carried out by the second graduate student in my lab, Cathy Mendelsohn. She identified a gene from human cells that encoded a protein which we believed to be the cellular receptor for poliovirus. When this human gene was expressed in mouse cells, it madepvr model them susceptible* to poliovirus infection (the mouse cells were already permissive for poliovirus replication). The gene encodes a transmembrane glycoprotein (illustrated) that we called the poliovirus receptor (PVR), later renamed CD155. Over the years we worked extensively on PVR, with the goals of understanding its interaction with poliovirus during entry into the cell. In one project we collaborated with Jim Hogle, David Belnap, and Alasdair Steven to solve the structure of poliovirus bound to a soluble form of PVR. The image of that complex decorates the banner at virology blog and twiv.tv.

Shortly after identifying PVR as the cellular receptor for poliovirus, a new student, Ruibao Ren, joined my lab. For his project I suggested he create transgenic mice with the human gene for PVR. We already knew that synthesis of PVR in mouse cells allowed the complete poliovirus replication cycle. Together with Frank Costantini and JJ Lee, Ruibao produced PVR transgenic mice and showed that they were susceptible to poliovirus infection. The illustration at top left shows a PVR transgenic mouse with a paralyzed left hind limb after poliovirus inoculation.

Poliovirus transgenic mice were used for many years in my laboratory to study how the virus causes disease, and to identify the mutations that attenuate the neurovirulence of the Sabin vaccine strains. A good summary of this work can be found in my review, ‘One hundred years of poliovirus pathogenesis‘. But there is a dark side of this story that I wish to briefly recount. When we first developed PVR transgenic mice, my employer decided to patent the animals. Until the patent issued, we could not share the transgenic mice with other researchers. As a consequence, others developed their own lines of PVR transgenic mice. One of these lines has been qualified by the World Health Organization to determine the neurovirulence of the Sabin vaccine strains. However, Columbia University realized little income from the PVR transgenic mice – such animals cannot be patented in Europe. By patenting the mice, we simply delayed research progress. Because of this experience I am personally very wary about patenting biological discoveries.

There are several reasons why I decided to stop doing research with mice. The cost of housing and breeding mice is very high, nearly $1.00 US per cage per day, and I simply don’t have the funds to support such work. More importantly, no one in my laboratory has any interest in working with mice: the last student to do mouse work left years ago. Although there are many interesting experiments to be done using viruses and mice, that line of work ended for the Racaniello lab on 11 July 2011.

*A susceptible cell bears the receptor for the virus; a permissive cell allows viral replication. A susceptible and permissive cell allows the complete viral replication cycle.

 Ren, R., Costantini, F., Gorgacz, E., Lee, J., & Racaniello, V. (1990). Transgenic mice expressing a human poliovirus receptor: A new model for poliomyelitis Cell, 63 (2), 353-362 DOI: 10.1016/0092-8674(90)90168-E

Poliomyelitis after a twelve year incubation period

Sabin type 2 poliovirusAnalysis of poliovirus recovered from the stool of a patient with fatal poliomyelitis revealed that she had been infected with the virus 12 years earlier, probably when one of her children received the oral poliovirus vaccine. This case has the longest known incubation period for vaccine-derived poliomyelitis, and highlights our still rudimentary understanding of how poliovirus causes disease.

The patient in this case, a 44 year old woman from Minnesota, had been diagnosed with common variable immunodeficiency (CVI) in 1991.  Patients with this disease lack B lymphocytes and therefore cannot produce antibodies that help control microbial infections. For example, individuals with CVI often develop chronic enterovirus infections. Furthermore, after receiving oral poliovirus vaccine, CVI patients may shed infectious virus for long periods of time, often in the absence of clinical disease. To help control infections, CVI patients are regularly given pooled immune globulin harvested from healthy individuals.

The patient’s first symptoms were cough, runny nose, malaise, and low grade fever, which resolved in 4 days. Two days later she experienced leg cramps which lead to leg weakness, a fall, and hospitalization. The weakness spread to the upper extremities and involved severe muscular pain. Respiratory failure developed and the patient died after 92 days of hospitalization. The initial mild disease, followed by neurological symptoms, is a classic course of poliomyelitis.

Poliovirus was isolated from a stool sample taken at hospital day 74. Sequence analysis revealed that it was derived from the Sabin type 2 vaccine strain. Because poliovirus evolves at a rate of 1% per year, it was possible to determine that the patient had become infected with the Sabin vaccine approximately 11-13 years previously. It is likely that the patient acquired the infection when her 13 year old child received poliovirus vaccine in 1995. By that time the patient had been diagnosed with CVI, and no member of her household should have received oral poliovirus vaccine. It is recommended that CVI patients and their household contacts receive inactivated poliovirus vaccine.

If we assume that the patient excreted poliovirus for 11-13 years (it is formally possible that the patient recently acquired vaccine-derived poliovirus from other sources), then an interesting question is why neurological disease took so long to develop. The answer is unknown, but may be related to the fact that in healthy persons, paralytic disease only occurs in 1% of infected individuals. Why most poliovirus-infected individuals do not develop neurological disease remains a mystery. The results of experiments in a mouse model for poliomyelitis have provided some clues. In mice, poliovirus invasion of the nervous system is limited by a strong innate immune response – the production of interferons – in non-neural tissues. In addition, viral invasion of the nervous system is dependent on transport along axons, which is an extremely inefficient process. Whether the same factors limit poliovirus invasion of the human central nervous system is not known.

Another unanswered question is why the patient was not protected by the pooled immune globulin which she received every three weeks. These preparations are tested to ensure that they contain a minimum titer of antibodies against polioviruses. It is possible that the amount of antibody needed to protect individuals with a chronic and acute poliovirus infection are different. Nevertheless, the administration of pooled immune globulin apparently failed to prevent the initial infection, the chronic infection, and paralytic disease.

This case emphasizes the need to continue research on poliovirus. Our knowledge of how the virus causes disease is still rudimentary – as is evident by our failure to understand the 12 year incubation period of the case described here. Although the polio eradication campaign has had excellent results, it can be compromised by vaccine-associated disease. Individuals with B lymphocyte deficiencies will be a reservoir for the virus and can lead to infections if immunization levels drop. It would be highly beneficial to identify all individuals with chronic poliovirus infections, and treat them with antivirals. Unfortunately, such compounds do not exist – underscoring the need to continue research to identify drugs that can be used to treat poliovirus infection.

DeVries AS, Harper J, Murray A, Lexau C, Bahta L, Christensen J, Cebelinski E, Fuller S, Kline S, Wallace GS, Shaw JH, Burns CC, & Lynfield R (2011). Vaccine-derived poliomyelitis 12 years after infection in Minnesota. The New England journal of medicine, 364 (24), 2316-23 PMID: 21675890

Lancaster KZ, & Pfeiffer JK (2010). Limited trafficking of a neurotropic virus through inefficient retrograde axonal transport and the type I interferon response. PLoS pathogens, 6 (3) PMID: 20221252

Is bivalent poliovirus vaccine a good idea?

polio-immunizationA new bivalent poliovirus vaccine, consisting of infectious, attenuated type 1 and type 3 strains, has been deployed in Afghanistan. The use of this vaccine was recommended by the Advisory Committee on Poliomyelitis Eradication, the global technical advisory body of the Global Polio Eradication Initiative. Considering the polio experience in Nigeria, the elimination of type 2 poliovirus from the vaccine might have serious consequences.

There are three serotypes of poliovirus, all of which can cause poliomyelitis. Infection with one serotype of the virus does not confer protection against the other two; therefore poliovirus vaccines have always included all three serotypes (they are trivalent). The attenuated vaccine that is used in the eradication effort is an infectious vaccine. The vaccine is ingested, the viruses replicate in the intestine, and immunity develops. Viruses of all three serotypes undergo genetic changes during replication in the alimentary tract. As a consequence, the vaccine recipient excretes polioviruses that can cause paralysis. These so-called vaccine-derived polioviruses (VDPV) can cause outbreaks of poliomyelitis in non-immune people, as described in Polio among the Amish.

Poliovirus type 2 was declared eradicated from the globe by the World Health Organization in 1999. When type 2 poliovirus was eliminated, many countries began using monovalent type 1 and type 3 vaccines: one vaccine for type 1 and another for type 3. As a consequence of this immunization strategy, population immunity to type 2 poliovirus declined. Not unexpectedly, there was an outbreak of type 2 poliovirus in Nigeria in 2006. The surprise was that the outbreak was caused by a poliovirus type 2 vaccine strain.

Before 2003, the year that Nigeria began a boycott of polio immunization, the trivalent polio vaccine was used. Immunization resumed with monovalent types 1 and 3 vaccine in 2004. Therefore the source of the VDPV type 2 is most likely the trivalent vaccine used before 2003.

The press release at polioeradication.org announcing the bivalent vaccine proclaims:

Of the three wild polioviruses (known as types 1, 2 and 3), type 2 has not been seen anywhere in the world since 1999.

The statement ignores the fact that there is vaccine-derived type 2 poliovirus in the world – and it can cause polio as well as ‘wild’ poliovirus. Such strains have been isolated in Nigeria as recently as October 2009. Why isn’t the type 2 vaccine being used in Afghanistan when it is very likely that vaccine-derived type 2 poliovirus is still circulating? Just because we haven’t isolated type 2 poliovirus recently doesn’t mean that it’s gone. No type 2 poliomyelitis was detected in 1999, yet the vaccine-derived virus was silently circulating in humans.

What will be the WHO response to an outbreak of type 2 polio in Afghanistan? They will probably deploy trivalent vaccine, as was done in Nigeria in 2006. But this approach will simply lead to another cycle of eradication and emergence of type 2 polio. It’s time to begin using inactivated poliovirus vaccine, which I’ve been dreaming about for some time.

Poliovirus type 2 returns

polio-immunizationThe global battle to eradicate poliomyelitis is already 9 years behind schedule. To make matters worse, type 2 poliovirus, which was declared eradicated in 1999, has returned.

There are three serotypes of poliovirus, each of which causes poliomyelitis. The vaccine used by WHO in the global eradication effort is a trivalent preparation comprising all three serotypes. When type 2 poliovirus was eliminated, many countries began using monovalent type 1 and type 3 vaccines: one vaccine for type 1 and another for type 3. As a consequence of this immunization strategy, population immunity to type 2 poliovirus declined. But if type 2 poliovirus was eradicated, where has it come from?

It came from the poliovirus vaccine.

The trivalent vaccine that is used in the eradication effort is an infectious vaccine. The vaccine is ingested, the viruses replicate in the intestine, and immunity develops. Viruses of all three serotypes undergo genetic changes during replication in the alimentary tract. As a consequence, the vaccine recipient excretes neurovirulent polioviruses. These so-called vaccine-derived polioviruses (VDPV) can cause outbreaks of poliomyelitis in non-immune people, as described in Polio among the Amish.

The outbreak of type 2 poliovirus in Nigeria began in 2006. There have been 126 cases of paralytic disease reported so far in 2009. Before 2003, the year that Nigeria began a boycott of polio immunization, the trivalent vaccine was used. Immunization resumed with monovalent types 1 and 3 vaccine in 2004. Therefore the source of the VDPV type 2 is most likely the trivalent vaccine used before 2003.

The resurrection of type 2 polio highlights the difficulties involved in using an infectious viral vaccine to eradicate the disease. In reality, type 2 poliovirus was not eradicated in 1999, because that virus was still present in the trivalent vaccine that was being used. Clearly the virus was still circulating in humans, despite the fact that no type 2 poliomyelitis was observed.

In response to the type 2 outbreak in Nigeria, trivalent vaccine is being used again. It’s not difficult to imagine that this will lead to another cycle of eradication and emergence of type 2 polio. What’s the solution to this apparently endless circle? Use inactivated poliovirus vaccine, which I’ve been dreaming about for some time.

Roberts, L. (2007). Vaccine-Related Polio Outbreak in Nigeria Raises Concerns Science, 317 (5846), 1842-1842 DOI: 10.1126/science.317.5846.1842

Roberts, L. (2009). Type 2 Poliovirus Back From The Dead in Nigeria Science, 325 (5941), 660-661 DOI: 10.1126/science.325_660

Jegede, A. (2007). What Led to the Nigerian Boycott of the Polio Vaccination Campaign? PLoS Medicine, 4 (3) DOI: 10.1371/journal.pmed.0040073